immunohematology - american red cross · immunohematology, volume 23, number 1, 2007 1 this issue...

ImmunohematologyJ O U R N A L O F B L O O D G R O U P S E R O L O G Y A N D E D U C A T I O N

V O L U M E 2 3 , N U M B E R 1 , 2 0 0 7

ImmunohematologyJOU R NA L O F B L O O D G RO U P S E RO L O G Y A N D E D U C AT I O N

VO L U M E 2 3 , N U M B E R 1 , 2 0 0 7

C O N T E N T S

1Dedication in memoriam

R.R.VASSALLO

2HPA-1a/b(PlA1/A2, Zwa/b): the odyssey of an alloantigen system

R.H.ASTER AND P.J. NEWMAN

9Neonatal alloimmune thrombocytopenia: a 50-year story

C. KAPLAN

14Platelet additive solutions: current status

H. GULLIKSSON

20ABO and platelet transfusion therapy

L. COOLING

34Scott Murphy’s contribution in the early years of posttransfusion purpura: a remembrance

M.KEASHEN-SCHNELL

38Differences in ABO antibody levels among blood donors: a comparison between past and present

Japanese, Laotian, and Thai populations

T.MAZDA, R.YABE, O. NATHALANG,T.THAMMAVONG,AND K.TADOKORO

42 43COMMUN I C AT I O N I N M EMO R I A M

Missing Scott Murphy, a platelet “maestro” Katherine M. Beattie

P. REBULLA

44 45E R R AT UM A N NO UN C E M E N T S

46 50A DV E R T I S E M E N T S I N S T RU C T I O N S F O R AU T HO R S

EDITORS-IN-CHIEF MANAGING EDITORSandra Nance,MS,MT(ASCP)SBB Cynthia Flickinger,MT(ASCP)SBB

Philadelphia, Pennsylvania Philadelphia, Pennsylvania

Connie M.Westhoff, MT(ASCP)SBB, PhDPhiladelphia, Pennsylvania

TECHNICAL EDITORS SENIOR MEDICAL EDITORChristine Lomas-Francis,MSc Geralyn M.Meny,MD

NewYork City, NewYork Philadelphia, Pennsylvania

Dawn M. Rumsey,ART(CSMLT)Glen Allen, Virginia

ASSOCIATE MEDICAL EDITORSDavid Moolton,MD Ralph R.Vassallo,MD

Philadelphia, Pennsylvania Philadelphia, Pennsylvania

EDITORIAL BOARD

EMERITUS EDITORIAL BOARDSandra Ellisor,MS,MT(ASCP)SBB Delores Mallory,MT(ASCP)SBB

Anaheim, California Supply, North Carolina

EDITORIAL ASSISTANT PRODUCTION ASSISTANTJudith Abrams Marge Manigly

COPY EDITOR PROOFREADER ELECTRONIC PUBLISHERMary L.Tod Lucy Oppenheim Paul Duquette

Immunohematology is published quarterly (March, June, September, and December) by the American Red Cross, National Headquarters,Washington, DC 20006.

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Copyright 2007 by The American National Red CrossISSN 0894-203X

Patricia Arndt,MT(ASCP)SBBPomona, California

James P.AuBuchon,MDLebanon, New Hampshire

Martha R. Combs,MT(ASCP)SBBDurham, North Carolina

Geoffrey Daniels, PhDBristol, United Kingdom

Anne F. Eder,MDWashington, District of Columbia

George Garratty, PhD, FRCPathPomona, California

Brenda J. Grossman,MDSt. Louis, Missouri

W. John Judd, FIBMS,MIBiolAnn Arbor, Michigan

Christine Lomas-Francis,MScNewYork City, NewYork

Gary Moroff, PhDRockville, Maryland

John J.Moulds,MT(ASCP)SBBShreveport, Louisiana

Paul M.Ness,MDBaltimore, Maryland

Joyce Poole, FIBMSBristol, United Kingdom

Mark Popovsky,MDBraintree, Massachusetts

Marion E. Reid, PhD, FIBMSNewYork City, NewYork

S. Gerald Sandler,MDWashington, District of Columbia

Jill R. Storry, PhDLund, Sweden

David F. Stroncek,MDBethesda, Maryland

I M M U N O H E M A T O L O G Y, V O L U M E 2 3 , N U M B E R 1 , 2 0 0 7 1

This issue of Immuno-hematology is a very specialone. It is dedicated toDr. Scott Murphy, a uniquelygifted physician renowned forhis equanimity, who passedaway last year after a long andcourageous battle with lympho-ma. After lengthy service onImmunohematology’s editorialboard, Scott became its seniormedical editor in 2003,serving in

that capacity for almost 3 years, until his untimelydeath in April 2006. He was a driving force behind abold new direction for the journal, beginning with its20th anniversary celebration in 2004, marked by fourspecial issues devoted to review articles. As a result ofScott’s efforts, the journal has a new look. Every issueis packed with important reviews from leaders intransfusion medicine along with the primary researchreports that have always made the journal a valuableresource for blood group serologists and molecularbiologists.

After an internal medicine internship andresidency at the Peter Bent Brigham Hospital in Boston,Scott returned to his native Philadelphia to complete aresearch fellowship in hematology at the Presbyterian-University of Pennsylvania Medical Center. Hepublished his very first paper in the New EnglandJournal of Medicine the following year. A landmarkreport, this research launched warm platelet storage,making routine platelet support feasible and usheringin an era of aggressive cancer treatment and complexsurgical procedures. Scott served on the medical staffsof the Presbyterian-University of Pennsylvania MedicalCenter and the Thomas Jefferson University Hospital,finally accepting the position of chief medical officer atthe Penn-Jersey Region of the American Red CrossBlood Services in 1994. Throughout his career, heguided international efforts in platelet containerdesign, storage medium development, and plateletviability assessment. Most recently, he developed“Murphy’s Rule,” a proposal for an objective qualitystandard for stored platelets adopted by the Food andDrug Administration. The importance of Scott’s work

has been recognized with numerous honors, includingthe AABB’s Karl Landsteiner Award and the AmericanRed Cross’ Charles R. Drew Award for LifetimeAchievement. He was a member and past chairman ofthe Biomedical Excellence for Safer Transfusion (BEST)Collaborative, a prolific group of internationaltransfusion medicine leaders, an accomplishment ofwhich Scott was quite proud.

This issue, written by his peers and collaborators,contains reviews of interest to platelet serologists andhospital-based cellular therapists. Richard Aster andPeter Newman recount the noteworthy history of thediscovery of the HPA-1 polymorphism that has resultedin a better understanding of platelet alloimmunity andimprovements in the support of alloimmunizedpatients. Cecile Kaplan takes this further with a skillfuldescription of the consequences of platelet alloim-munization in fetuses and neonates, reviewing thecommon syndrome of fetomaternal/neonatal allo-immune thrombocytopenia. Hans Gulliksson providesa fascinating perspective on platelet additive solutionsthat promise to improve storage conditions and reducethe allogeneic plasma content of apheresis and wholeblood–derived platelets. Laura Cooling writes elegantlyabout the basic science underlying major and minorABO mismatches in platelet transfusion as well as theclinical consequences of the use of out-of-groupplatelets. Maryann Keashen-Schnell shares a personalperspective as one of Scott’s coworkers, investigating aperplexing case of posttransfusion purpura.

Finally, an original article and several noteworthycommunications complete this issue, dedicated to thelife and work of a dearly missed collaborator, mentor,leader, and friend, Scott Murphy.

Ralph R.Vassallo, MD, FACPAssociate Medical Editor and

Guest Editor of this issueChief Medical Officer

American Red Cross Blood ServicesPenn-Jersey Region

700 Spring Garden StreetPhiladelphia, PA 19123

Dedication in memoriam

2 I M M U N O H E M A T O L O G Y, V O L U M E 2 3 , N U M B E R 1 , 2 0 0 7

HPA-1a/b (PlA1/A2, Zwa/b): theodyssey of an alloantigen systemR.H.ASTER AND P.J. NEWMAN

The HPA-1a/b (PlA1/A2, Zwa/b) alloantigen system wasone of the first such systems to be identified on a cellother than the RBC. From the discovery of HPA-1a in1958 to the present day, studies of this antigen and itsallele, HPA-1b, have led to a remarkable number of“firsts” in immunobiology. In this review,we shall tracethe history of the HPA-1a/b system and will highlightselected observations made during the past 48 yearsthat have contributed not only to the field of plateletimmunology but also to immunohematology andtransfusion medicine in general.

Serologic Characterization of HPA-1a and 1b,the First Recognized Platelet-SpecificAlloantigen System

It is likely that an HPA-1a-specific alloantibody wasfirst identified by Zucker and colleagues1 in 1956 or1957 in studying a patient who developed profoundthrombocytopenia and hemorrhagic symptoms 5 daysafter receiving a blood transfusion. The patient’s serumcontained a strong antibody that producedagglutination (Fig. 1) and induced complement-dependent lysis of platelets from four normal subjectsbut not with the patient’s own platelets obtained afterrecovery, about 4 weeks after the acute episode.Although the antigen recognized by this antibody wasnot characterized further, this case was almost certainlythe first example of a syndrome that was laterdesignated “posttransfusion purpura” (PTP) associatedwith an antibody specific for HPA-1a. Shortlythereafter, van Loghem and colleagues2 identified aplatelet isoagglutinin in serum from a transfusedpatient and showed that it was specific for a markerthey designated “Zwa” that was present in about 98percent of the Dutch population and was inherited asa dominant trait. Their studies were facilitated by thefact that van Loghem himself possessed the rare Zwa-negative phenotype. At about the same time, Shulmanand colleagues3 at the National Institutes of Health in

Bethesda used platelet agglutination and quantitativecomplement fixation to characterize two examples ofan alloantibody specific for a high-frequencyalloantigen they called“platelet antigen 1” (PlA1). It wassoon found that PlA1 and Zwa were identical.4 Theexpected allele of Zwa (Zwb) was identified by van derWeerdt et al.5 as the target for an IgM plateletisoagglutinin found in a transfused patient. When aunified nomenclature was developed for platelet-specific alloantigens, Zwa/PlA1 and Zwb/PlA2 weredesignated “HPA-1a” and “HPA-1b,” respectively.6

Analysis of the allelic frequencies of this and otherplatelet alloantigen systems suggests that the HPA-1aepitope was present in the primordial allele of plateletmembrane glycoprotein (GP) IIIa (discussed in a later

Fig. 1. The first anti-HPA-1a–specific antibody was detected on the basisof its ability to cause complement-dependent lysis of normalplatelets. EDTA platelet-rich plasma was reconstituted with 0.1MMgCl2. Patient serum (above) but not normal serum (below)produced platelet fragmentation when the mixture wasincubated at 37°C.*

*Reprinted with permission from Zucker et al.1


HPA-1a/b(PlA1/A2, Zwa/b) alloantigen system

section), traveled with early humans out of Africaduring migrations to Northern Europe approximately40,000 years ago, and finally came to North Americawith European colonists about 500 years ago (Fig. 2).Today, the approximate gene frequencies of HPA-1a andHPA-1b in the latter two populations are 0.86 and 0.14,respectively. However,HPA-1b is much less common inAmerindians7 and African Americans8 and is extremelyrare in Asians.8,9

Early Studies Led to Identification of TwoThrombocytopenic Syndromes AssociatedWith Sensitization to HPA-1a

The two examples of anti-HPA-1a studied byShulman and his colleagues3 came from patients whohad received a blood transfusion and, 1 week later,

developed profound thrombocytopenia lasting about 3weeks (Fig. 3). They coined the term “posttransfusionpurpura” (PTP) to define the combination of clinicaland serologic findings typical of this condition. By1986, at least 75 well-documented cases of PTP, nearlyall associated with anti-HPA-1a antibodies, had beendescribed. Not long after their description of PTP,Shulman et al.4 identified two more examples of anti-HPA-1a in women who had given birth to infants withsevere neonatal alloimmune thrombocytopenia(NATP), and provided the first description of thatcondition involving maternal-fetal incompatibility for aplatelet-specific antigen. It is now recognized thatNATP occurs in about one of approximately every1000 newborns, and that maternal-fetal incompatibilityof HPA-1a is responsible in the majority of cases.11

HPA-1a-Negative Platelets Were Used for theFirst “Antigen-Compatible” PlateletTransfusions

The recognition of NATP as a clinical entity raisedthe issue of whether HPA-1a-negative (HPA-1b-positive)platelets could be transfused successfully to infantswith thrombocytopenia caused by maternal anti-HPA-1a. Because only 2 percent of the general population ishomozygous for HPA-1b and few blood banks werecapable of performing platelet typing in the 1960s,platelets from HPA-1a-negative donors were notgenerally available. However, Adner and colleagues12

showed that washed maternal platelets producedsatisfactory posttransfusion increments in an infantwith this condition. Because a mother’s platelets areinvariably compatible with her own antibody, thisapproach has subsequently been used for treatment ofmany infants born with NATP, regardless of thespecificity of the mother’s alloantibody.

Localization of HPA-1a to GPIIIa (theIntegrin β3 Subunit)

From studies done using techniques available in1960, Shulman and colleagues3 concluded that HPA-1amight reside on a relatively abundant plateletmembrane protein. An important clue to the actuallocalization of the antigen came from the finding byKunicki and Aster13 that platelets from patients withtype I Glanzmann thrombasthenia, known to lackglycoproteins IIb and IIIa, were invariably HPA-1anegative. Kunicki and Aster14 then used immuno-chemical methods to isolate GPIIIa, showed that it wasthe carrier protein for HPA-1a, and speculated that the

Fig. 2. Early mutations in the GPIIIa gene in Homo sapiens gave rise topolymorphisms responsible for creation of platelet alloantigenicepitopes.

Fig. 3. PTP occurring 6 days after a blood transfusion in a 43-year-oldwoman. Prednisone was without effect on the platelet count, butspontaneous recovery occurred after about 3 weeks. Acomplement-fixing antibody was detected on Day 11 anddeclined to undetectable levels at about the time of plateletrecovery.*

*Reprinted with permission from Shulman et al.3



antigen was held in a configuration suitable forantibody recognition by one or more disulfide bonds.Newman and colleagues15 soon thereafter localized theHPA-1a epitope to a 17-kd tryptic fragment of theGPIIIa and provided evidence that oligosaccharidemoieties were not required for immunogenicity.Recognition that GPIIIa was one of the subunits of theintegrin αIIbβ316 enabled others to show that HPA-1a/balloantigens are not restricted to platelets, but can alsobe expressed as part of the αvβ3 complex onendothelial cells,17,18 macrophages, and fibroblasts.19

Genetic Basis for the HPA-1a/bPolymorphism

Discovery of PCR in the mid-1980s made it possiblefor the first time to identify nucleotide and amino acidpolymorphisms assumed to be responsible for thecreation of platelet alloantigens. Initially, it appearedthat this approach might not be applicable to “plateletmolecular biology” as platelets both lack a nucleus andcontain only vanishingly small amounts of mRNA. Thisproblem was circumvented by showing that mRNAcould be extracted from circulating platelets,converted into cDNA, cloned, and then sequenced.20

Using this approach, Newman and colleagues21 thensequenced cDNAs encoding GPIIIa from serologicallyidentified HPA-1a-positive and HPA-1a-negativeplatelets and found a single nucleotide that accountedfor the generation of Leu33 (Pl

A1 = HPA-1a) and Pro33

(PlA2 = HPA-1b) allelic forms of GPIIIa. That theLeu33Pro polymorphism was not only associated withthis alloantigen system but was directly responsible forcreating the alloantigenic epitope was later shown byexpressing the Leu33 and Pro33 forms of GPIIIa inChinese hamster ovary cells and demonstrating thatanti-HPA-1a human alloantisera bound only the Leu33

allelic isoform, whereas anti-HPA-1b sera reacted withonly the Pro33 form of the glycoprotein.22 Thiscombined approach was subsequently used by us andothers to characterize nucleotide and amino acidsubstitutions responsible for the creation of otherclinically important platelet-specific alloantigens,23–25

making possible routine DNA typing for most of theplatelet antigen systems,26,27 an application that is nowcommonplace in transfusion medicine.

PlA1 at Last! Structural Characterization of aPlatelet-Specific Alloantigen

The discovery that the HPA-1a and -1b alloantigensystem was controlled by a polymorphism at amino

acid 33 of GPIIIa, together with a complex biochemicalanalysis of disulfide bond assignments within GPIIIa,28

facilitated generation of several models of the HPA-1aand HPA-1b epitopes.One such early model is shown inFigure 4, which depicts a circular peptide loop heldtogether by cysteine residues 26 and 38 that waspredicted to form the alloantigen. Frustratingly,however, when cyclic peptides expected to containHPA-1a/b were synthesized, they were found to beincapable of reacting with HPA-1a- and HPA-1b-specificantibodies.29 An explanation for this apparent anomalywas finally provided by Springer and colleagues,30 whodetermined the actual crystal structure of the N-terminal plextrin-semaphorin-integrin (PSI) homologydomain of GPIIIa—the domain that encompassespolymorphic residue 33. In addition to providing thecorrect disulfide bond assignments, this landmark studyprovided the coordinates of the PSI domain at the N-terminus of β3 integrin and made it possible tovisualize for the first time the region of GPIIIa thatcontrols formation of the HPA-1a and HPA-1b epitopes(Fig. 5)—some 45 years after the initial description ofthis platelet alloantigen system! More work remains,however, because the actual surface in GPIIIarecognized by HPA-1a-specific antibodies appears to becomplex, as Valentin and coworkers31 found that

Fig. 4. Schematic diagrams from the early 1990s of the hypotheticalstructure of the GPIIb-IIIa complex (upper left). The aminoterminus of GPIIIa, including a small disulfide-bonded loop thatwas thought at that time to be formed by amino acids 26 and 38and to encompass polymorphic amino acid residue 33 (PlA1/PlA2

controlling region) is shown in the upper right (encircled).Bottom: an early molecular model of the Leu33 and Pro33 forms ofthe AA 26–38 loop thought to encompass the PlA1 and PlA2

alloantigenic epitopes. Modeling was performed by Jack Gorski,Blood Research Institute, BloodCenter of Wisconsin, using SYBILmolecular modeling software running on a Silicon GraphicsWorkstation.


HPA-1a/b(PlA1/A2, Zwa/b) alloantigen system

certain mutant forms of GPIIIa are recognized by some,but not all, HPA-1a-specific antibodies, and devised theterms “type 1” and “type 2” to describe those thatrequire a cysteine residue at amino acid residue 435and those that do not, respectively. It seems likely thattype 1 antibodies recognize a peptide sequencecontaining Leu33 plus a noncontiguous peptidesequence near Cys435.

The Immune Response to HPA-1a is UniqueAs blood banks and other laboratories acquired the

ability to detect HPA-1a-specific antibodies, it soonbecame apparent that not all HPA-1a-negativeindividuals challenged with HPA-1a-positive plateletsbecame sensitized to this antigen. In a series of studiesbegun in the 1980s,32,33 it was found that HPA-1a isunique among blood group antigens in inducing animmune response that is almost invariably linked to theclass II HLA marker DRB3*010134 or DQB1*02.35 In thecase of NATP, the relative risk for a fetus that is HPA-1aincompatible with its mother is approximately 141 ifthe woman is positive for DRB3*0101, roughly thesame as the risk of developing ankylosing spondylitis inHLA-B27-positive individuals.36 The allele HPA-1binduces antibodies much less often than HPA-1a, andthe response to this marker is not HLA-linked.37

A potential explanation for the remarkableassociation between the immune response to HPA-1aand HLA was provided by Gorski and associates.36,38 Inone series of studies, they showed that T cells from twowomen who had produced anti-HPA-1a antibodies arespecifically stimulated by a cyclic peptide containingthe polymorphism that controls HPA-1a expression(residues 27–37 of the HPA-1a allele of GPIIIa, β3integrin) but not by the same set of peptides from theHPA-1b allele (Leu to Pro at amino acid residue 33).36

Interestingly, a common complementarity determiningregion motif was identified in responding T cells fromtwo different women, despite use of genes fromdifferent V beta families to encode the peptiderecognition domain. These findings showed that theresponse to HPA-1a is unusual in that the same peptideis recognized by both B-cell and T-cell receptors. Thesame group then found that a peptide containing Leu33

bound tightly to recombinant DRB3*1010 whereas theone containing Pro33 was nonreactive, thus providing amolecular explanation for the unidirectional nature ofthe immune response to HPA-1 antigens.38

The HPA-1a/b Alloantigen System inHemostasis

A new and still evolving chapter in the HPA-1a/bstory began with observations by Weiss andcoworkers39 that persons admitted to an intensive careunit with myocardial infarction were more likely to bepositive for HPA-1b than individuals from the generalpopulation. This association appeared to be particu-larly significant in younger individuals. This report setoff a series of epidemiological and biochemical studiesthat are remarkable for lack of consensus in nearly 100published reports. For example, one study found thatfibrinogen binds more readily to GPIIb/IIIa onactivated HPA-1b-positive platelets than on HPA-1b-negative platelets,40 but no difference was found bytwo other groups.41,42 Various reports suggested thatHPA-1b-positive individuals are at higher risk forthrombosis or premature atherosclerosis42,43 and evenrenal transplant rejection.44 However, a retrospectiveanalysis of DNA samples from almost 15,000individuals found no association between HPA-1b andthrombosis, atherosclerosis, or stroke.45 HPA-1b-positive platelets were found to be hypersensitive tothe agonist ADP in one study46 and resistant toactivation by thrombin receptor activating peptide inanother.47 Recent work has provided a certain amountof objective biochemical evidence that HPA-1b-positive

Fig. 5. PlA1 at last! The amino terminal PSI domain of GPIIIa, based onthe actual crystal structure of the molecule, is shown in fourdifferent rotated views. Amino acid 33 is portrayed as a ball andstick structure at the base of a disulfide-bonded loop formed byresidues 16 and 38. The structure is complicated by the presenceof two additional cysteine residues within this loop—one atamino acid 23, and the other at residue 26—making mimickingthe epitope for potential therapeutic purposes an extremelychallenging task. Amino acid strands connecting the PSI domainto the hybrid domain of GPIIIa are shown at the top of eachstructure.



platelets may be “hyperfunctional” in severalrespects.48,49 Perhaps further work along these lineswill provide an explanation for apparently discordantobservations made at the clinical level.

ConclusionSince the alloantigen system now designated HPA-

1a/b was discovered about 1960, serologic,biochemical, epidemiologic, and molecular biologicalstudies of this diallelic alloantigen system havecontributed importantly to our understanding ofalloimmune thrombocytopenia, the cellular andhumoral basis of the alloimmune response, thestructural basis of alloantigenicity, and plateletpathophysiology. As more is yet to be learned, it is tobe hoped that further examination of this remarkablealloantigen system will be similarly rewarding.

AcknowledgmentSupported by Grants HL-13629 and HL-44612 from

the National Heart, Lung, and Blood Institute.

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31. Valentin N,Visentin GP, Newman PJ. Involvementof the cysteine-rich domain of glycoprotein IIIa inthe expression of the human platelet alloantigen,PlA1: evidence for heterogeneity in the humoralresponse. Blood 1995;85:3028–33.

32. Reznikoff-Etievant MF, Muller JY, Julien F, PatereauC. An immune response gene linked to MHC inman.Tissue Antigens 1983;22:312–4.

33. Mueller-Eckhardt C, Mueller-Eckhardt G, Willen-Ohff H, et al. Immunogenicity of and immuneresponse to the human platelet antigen Zwa isstrongly associated with HLA-B8 and DR3. TissueAntigens 1985;26:71–6.

34. Valentin N, Vergracht A, Bignon JD, et al. HLA-DRw52a is involved in alloimmunization againstPL-A1 antigen. Hum Immunol 1990;27:73–9.

35. L’Abbe D, Tremblay L, Goldman M, Decary F,Chartrand P. Alloimmunization to platelet antigenHPA-1a (Zwa): association with HLA-DRw52a isnot 100%.Transfus Med 1992;2:251.

36. Maslanka K, Yassai M, Gorski J. Molecularidentification of T cells that respond in a primarybulk culture to a peptide derived from a plateletglycoprotein implicated in neonatal alloimmunethrombocytopenia. J Clin Invest 1996;98:1802–8.

37. Kuijpers RW, von dem Borne AE, Kiefel V, et al.Leucine33-proline33 substitution in humanplatelet glycoprotein IIIa determines HLA-DRw52a (Dw24) association of the immuneresponse against HPA-1a (Zwa/PlA1) and HPA-1b(Zwb/PlA2). Hum Immunol 1992;34:253–6.

38. Wu S, Maslanka K, Gorski J. An integrinpolymorphism that defines reactivity withalloantibodies generates an anchor for MHC classII peptide binding: a model for unidirectionalalloimmune responses. J Immunol 1997;158:3221–6.

39. Weiss EJ,Bray PF,Tayback M,et al.A polymorphismof a platelet glycoprotein receptor as an inherited



risk factor for coronary thrombosis. N Engl J Med1996;334:1090–4.

40. Vijayan KV, Liu Y, Souza S, Thiagarajan P, Bray PF.Fibrinogen and prothrombin binding is enhancedto the Pro33 isoform of purified integrinalphaIIbbeta3. J Thromb Haemost 2006;4:905–6.

41. Bennett JS,Catella-Lawson F,Rut AR, et al. Effect ofthe Pl(A2) alloantigen on the function of beta(3)-integrins in platelets. Blood 2001;97:3093–9.

42. Meiklejohn DJ, Urbaniak SJ, Greaves M. Plateletglycoprotein IIIa polymorphism HPA 1b (PlA2):noassociation with platelet fibrinogen binding. Br JHaematol 1999;105:664–6.

43. Gardemann A, Humme J, Stricker J, et al.Association of the platelet glycoprotein IIIaPlA1/A2 gene polymorphism to coronary arterydisease but not to nonfatal myocardial infarctionin low risk patients. Thromb Haemost 1998;80:214–7.

44. Salido E, Martin B, Barrios Y, et al. The PlA2polymorphism of the platelet glycoprotein IIIAgene as a risk factor for acute renal allograftrejection. J Am Soc Nephrol 1999;10:2599–605.

45. Ridker PM, Hennekens CH, Schmitz C, StampferMJ, Lindpaintner K. PlA1/A2 polymorphism ofplatelet glycoprotein IIIa and risks of myocardial

infarction, stroke, and venous thrombosis. Lancet1997;349:385–8.

46. Feng D,Lindpaintner K,Larson MG,et al. Increasedplatelet aggregability associated with plateletGPIIIa PlA2 polymorphism: the FraminghamOffspring Study. Arterioscler Thromb Vasc Biol1999;19:1142–7.

47. Lasne D, Krenn M, Pingault V, et al. Interdonorvariability of platelet response to thrombinreceptor activation: influence of PlA2

polymorphism. Br J Haematol 1997;99:801–7.48. Vijayan KV, Liu Y, Dong JF, Bray PF. Enhanced

activation of mitogen-activated protein kinase andmyosin light chain kinase by the Pro33polymorphism of integrin beta 3. J Biol Chem2003;278:3860–7.

49. Vijayan KV, Liu Y, Sun W, Ito M, Bray PF.The Pro33isoform of integrin beta3 enhances outside-insignaling in human platelets by regulating theactivation of serine/threonine phosphatases. J BiolChem 2005;280:21756–62.

Richard H. Aster, MD, and Peter J. Newman, PhD,Blood Research Institute of BloodCenter ofWisconsin,Inc., and Departments of Medicine and Cell Biology,Medical College ofWisconsin, Milwaukee,Wisconsin.

IMPORTANT NOTICE ABOUT MANUSCRIPTS

FOR

IMMUNOHEMATOLOGY

Please e-mail all manuscripts for consideration to Marge Manigly at [email protected]


Since the first description of the immunologicmechanisms in neonatal and thrombocytopenicpurpura1 and the first report of a maternal antibodydirected against a platelet alloantigen inherited fromthe father,2 in the 1950s, much has been learnedconcerning fetal and neonatal alloimmunethrombocytopenia (FNAIT), but questions are stillunanswered. FNAIT has been regarded as the plateletcounterpart of HDN, but, in contrast to HDN, the firstinfant is affected in 50 percent of cases. This condition,which has been estimated to have an incidence of 1 in800 to 1 in 1000 live births,3,4 can cause severebleeding in the fetus and newborn, and allincompatible fetuses will be at risk for subsequentpregnancies. Therefore, it is important to diagnoseFNAIT and to manage the subsequent pregnancies toprevent the consequences of severe fetalthrombocytopenia.

1953–2007: the Platelet Alloantigen StoryOver the years, considerable progress has been

made in the characterization of platelet-specificalloantigens. Improvements in serologic methods forthe detection of maternal alloantibodies—includingthe use of antigen-capture assays, the monoclonalantibody-specific immobilization of platelet antigenstechnique,5 or monoclonal antigen capture ELISA6—the development of immunochemical techniques, andthe advent of molecular biological techniques have ledto the description of 24 platelet-specific alloantigens.A human platelet antigen (HPA) nomenclature wasadopted in 1990 to replace the lab-specificnomenclature that was used previously. The antigenicsystems are numbered in order of the date ofdiscovery; the high-incidence allele is called“a,”and thelow-incidence allele is called “b” in the originalpopulation in which the alleles were identified.7 Underthe auspices of the International Society of BloodTransfusion and the International Society ofThrombosis and Haemostasis, a Platelet Nomenclature

Committee has published tables that will be updated;these include the list of the HPA antigens, the antigengenetic basis, and the platelet antigen alleles defined bysequencing. The HPA nomenclature will still be usedfor clinical and scientific purposes.8 The reader isreferred to the table of numbered HPAs, theirglycoprotein location, and approximate Caucasianphenotype frequencies in “Scott Murphy’s contri-butions in the early years of posttransfusion purpura: aremembrance” in this issue of Immunohematology.

HPA antigens are expressed on different integrinsplaying a role in cellular interactions. α2bβ3 (GPIIb−IIIa)is the major platelet integrin and is restricted toplatelets, whereas the αvβ3 integrin is expressed morewidely. α2bβ1, also known as GPIa-IIa, is the secondmost important platelet integrin and is also found onlymphocytes. Antigens located on β3 (GPIIIa) havebeen found on other cells, such as endothelial cells, andon activated T lymphocytes when located on α2; thismay play a pathophysiologic role.

Frequencies of platelet antigens vary amongdifferent populations. In Caucasians, HPA-1a is by farthe most common antigen implicated in FNAIT,9

followed at much lower frequency by HPA-5b,10 thenHPA-3.11 In contrast, in Asians, FNAIT is essentiallyassociated with HPA-4 and HPA-5b. FNAIT has beenreported involving rare or private antigens.8 Recentstudies have shown that these low-frequency antigensare not restricted to single families,12 especially anti-HPA-9bw, which could account for up to 2 percent ofconfirmed cases13,14; therefore they must not be ignoredin the screening for FNAIT with a negative initiallaboratory investigation. The humoral maternalresponse is not uniform in this condition and must betaken into account when undertaking serologicdiagnosis.15 Further study of the characteristics of thematernal alloantibodies and their relevance to theclinical condition would be of interest.16

Neonatal alloimmunethrombocytopenia: a 50-year storyC.KAPLAN


C. KAPLAN

The genetic basis for maternal alloimmunizationhas been investigated. Alloimmunization to the HPA-1aantigen appears to be associated with HLA class IIalleles: DRB3*0101 and DQB1*0201 (odds ratio 24.9and 39.7, respectively).17,18 An anti HPA-1b responsewas not associated with either DRB3*0101 or anyknown HLA class II molecules.19 This finding impliesthe Leu33/Pro33 substitution on the plateletglycoprotein IIIa plays a role in antigen presentation.Data have shown that the binding of peptides from theLeu33/Pro33 dimorphic region to HLA-DR3*0101 isallele specific with stimulation of specific T cells20,21

providing help to B cells for generating alloantibodies;however, the positive predictive value is only 35percent, and utility for screening is therefore limited.

The immune response to HPA-5b antigen isstrongly associated with a particular DRB1 genesequence encoding residues Glu-Asp at positions 69and 70 of the DRβ chain.22 For other antigens, becauseof the low number of cases, statistical analyses are notsignificant when compared with the generalpopulation, as seen for HPA-6b and DRB1*1501,DQA1*0102, DQB1*0602 haplotypes shared byimmunized mothers.23

1953–2007: the Fetal and NeonatalAlloimmune Thrombocytopenia Story

Natural historyFNAIT has been known for decades as “neonatal

alloimmune thrombocytopenia” (NAIT). The usualpresentation is a full-term neonate exhibiting petechiaeor widespread purpura at birth, or a few hours afterbirth, to a healthy primiparous mother. Otherwise, thisinfant is well with no clinical signs of infection(hepatosplenomegaly) or malformation (hemangioma,absence of radii). Visceral hemorrhages such asgastrointestinal bleeding or hematuria are less commonthan purpura or hematoma. The thrombocytopenia isisolated. Coexisting anemia is caused by hemorrhage.

Anti-HPA-1a and anti-HPA-3a immunization inducesevere neonatal thrombocytopenia with plateletcounts less than 50 × 109/L in most cases.9,11 NAITlinked to HPA-5b incompatibility seems to be lesssevere than HPA-1a NAIT.10 The most seriouscomplication is fetal or neonatal intracranialhemorrhage (ICH; 25.5% of cases for HPA-1a, 24% forHPA-3a, 15% for HPA-5b)24 leading to death in up to 10percent or neurologic sequelae in up to 20 percent ofreported cases.

The risk of life-threatening hemorrhagenecessitates prompt diagnosis and effective therapy.Phenotyped platelet transfusion is the best postnatalmanagement. Because of the logistic difficulties inobtaining such platelets in emergency situations,random platelet transfusions with or without IVIG havebeen proposed.25 However, compatible platelets givebetter results.26 On the other hand, thrombocytopeniamay be asymptomatic and pass unnoticed unless thereis a routine platelet count performed. Therefore,unexpected or unexplained neonatal thrombo-cytopenia or severe early onset thrombocytopenia inboth preterm and term babies should raise thepossibility of NAIT and guide investigationsaccordingly. The fetal thrombocytopenia tends toworsen in subsequent incompatible pregnancies.

Fetal blood samplingIn 1983, a major advance in the diagnosis of NAIT

was the use of ultrasound-guided fetal blood sampling(FBS), which led to a better understanding of the fetalstatus.27 The mean platelet count has been evaluated tobe more than 150 × 109/L by the end of the firsttrimester of pregnancy in healthy fetuses and similar tothe adult platelet count later on. Therefore, thrombo-cytopenia has been defined in the fetus and theneonate as a platelet count less than 150 × 109/L,irrespective of the gestational age.28

In 1984, the first fetal alloimmune thrombo-cytopenia case was documented with FBS at 32 weeksof gestation, and in utero maternal platelet transfusionbefore delivery was proposed as therapy to avoidperinatal ICH.29 Severe fetal thrombocytopenia wasthen documented early during pregnancy when FBS,as part of the antenatal management protocol, wascarried out at 21 weeks of gestation for subsequentpregnancies in women with a previously affectedinfant.30 A retrospective survey of 5194 fetal bloodsamplings showed that fetal thrombocytopeniaresulting from maternal alloimmunization was themost severe thrombocytopenia observed among thedifferent disorders encountered, including chromo-somal malformations, infections, or maternalautoimmune thrombocytopenia.31

Development of antenatal managementThe rationale for antenatal management is the

high rate of recurrence for subsequent incompatiblefetuses who usually experience more severe thrombo-cytopenia. The first attempts to prevent severe


Neonatal alloimmune thrombocytopenia

thrombocytopenia in the preterm period and thus ICHduring delivery were in utero platelet transfusionsbefore delivery.30 However, this management could notprevent in utero ICH, which has been reportedprimarily before 30 weeks of gestation.24 Because ofthe short survival of transfused platelets, weekly fetalplatelet transfusions have been proposed and shown tobe effective in preventing ICH in a number of cases.32

The risks of in utero platelet transfusion, bleeding, andfetal loss have been estimated to be 1 to 2 percent perprocedure and 8 percent per pregnancy.33 Fetal cardiacarrhythmia (prolonged bradycardia) has also beenreported. Less invasive therapy has also beenproposed, involving maternal administration of IVIG,steroids, or both.

The mechanism of action of IVIG is complex,including inhibition of the transplacental passage ofmaternal alloantibodies and modification of theimmune response. Corticosteroids may modulate thematernal immune response. Different IVIG protocolshave been developed and differences in results werereported mainly because of the definition of asuccessful response.34–36 Low-dose corticosteroids, assole therapy, have been given to a limited number ofpatients, and response to therapy appears highlyvariable.35

The optimal management remains to bedetermined, and an international forum in 2003demonstrated the absence of standardization.37

The decision regarding therapy depends ondifferent factors, among which the fetal status plays acentral role. There is no reliable and sensitiveparameter for predicting which fetuses will be severelyaffected and which ones will respond to therapy. Theonly way to assess the fetal status is to perform FBS,butthe risk of serious adverse events from this techniqueis high, up to 10 percent.38 Most protocols favormaternal therapy with less invasive strategies39,40 withstratification according to the previously affectedsibling’s status.41

Serial in utero platelet transfusions are consideredonly as salvage therapy after failure of maternaltreatment.

Antenatal screeningNo systematic screening for FNAIT exists at the

moment, and this condition is still underdiagnosed.42

Reduction of neonatal death and disability is a publichealth issue, and recent progress in the prevention ofcurrent causes of neonatal disorders has been efficient

at reducing the risk of neonatal disorders goingundiagnosed and untreated. Prospective studies haveshown that although 2 to 3 percent of women are atrisk for developing anti-HPA immunization, only 1 in800 to 1000 newborns will be affected.3,4,43 It has beenfound that 26 percent of infants born to immunizedmothers are nonthrombocytopenic. Until proceduresare found that predict which women will have anaffected fetus, maternal screening has a low sensitivity.Only identifying thrombocytopenic newborns at birthincreases the sensitivity but will miss fetuses severelyaffected during pregnancy who should have beentreated. Cost-effectiveness analysis indicated screeningnewborns was more cost-effective than screeningprimiparous women.3 A consensus on routine investi-gation and optimal antenatal management has yet to bereached.

Future DirectionsAlthough real progress has been achieved in the

more accurate diagnosis and management of FNAIT,further studies must focus on improvements inantenatal management and on mechanisms ofmaternal sensitization. A recent study has shown thathigh maternal anti-HPA-1a alloantibody concentrationsmay provide an indication of severely affected fetuses44

and this may contribute to a less invasive antenataltherapeutic strategy. However, the follow-up of mater-nal anti-HPA-1a concentration during pregnancy shouldnot be considered as an indicator of therapeuticeffectiveness.

A murine model has also been recently developed,which may contribute to a better understanding of thiscondition in the future.45

References1. Harrington WJ, Sprague CC, Minnich V, et al.Immunologic mechanisms in neonatal andthrombocytopenic purpura. Ann Intern Med1953;38:433–69.

2. Moulinier J. Alloimmunisation maternelleantiplaquettaire “Duzo.” Proc 6th Congr Eur SocHaematol 1953;817–20.

3. Durand-Zaleski I, Schlegel N, Blum-Boisgard C, etal. Screening primiparous women and newbornsfor fetal/neonatal alloimmune thrombocytopenia:a prospective comparison of effectiveness andcosts.Am J Perinatol 1996;13:423–31.

4. Williamson LM, Hackett G, Rennie J, et al. Thenatural history of fetomaternal alloimmunization


to the platelet-specific antigen HPA-1a (PlA1, Zwa)as determined by antenatal screening.Blood 1998;92:2280–7.

5. Kiefel V, Santoso S,Weisheit M,Mueller-Eckhardt C.Monoclonal antibody-specific immobilization ofplatelet antigens (MAIPA): A new tool for theidentification of platelet-reactive antibodies.Blood1987;70:1722–6.

6. McMillan R. Antigen-specific assays in immunethrombocytopenia. Transfus Med Rev 1990;4:136–43.

7. von dem Borne AEGKr,Decary F.Nomenclature ofplatelet specific antigens. Br J Haematol 1990;74:239–40.

8. Metcalfe P, Watkins NA, Ouwehand WH, et al.Nomenclature of human platelet antigens. VoxSang 2003;85:240–5.

9. Mueller-Eckhardt C, Kiefel V, Grubert A, et al. 348cases of suspected neonatal alloimmune thrombo-cytopenia. Lancet 1989;1:363–6.

10. Kaplan C, Morel-Kopp MC, Kroll H, et al. HPA-5b(Br(a)) neonatal alloimmune thrombocytopenia:clinical and immunological analysis of 39 cases.BrJ Haematol 1991;78:425–9.

11. Glade-Bender J, McFarland JG, Kaplan C, PorcelijnL, Bussel JB. Anti-HPA-3a induces severe neonatalalloimmune thrombocytopenia. J Pediatr 2001;138:862–7.

12. Kroll H,Yates J, Santoso S. Immunization against alow-frequency human platelet alloantigen in fetalalloimmune thrombocytopenia is not a singleevent: characterization by the combined use ofreference DNA and novel allele-specific cell linesexpressing recombinant antigens. Transfusion2005;45:353–8.

13. Kaplan C, Porcelijn L, Vanlieferinghen Ph, et al.Anti-HPA-9bw (Maxa) feto-maternal alloimmuni-zation, a clinically severe neonatal thrombo-cytopenia: difficulties in diagnosis and therapy,report on 8 families. Transfusion 2005;45:1799–803.

14. Peterson JA, Balthazor SM, Curtis BR, McFarlandJG, Aster RH. Maternal alloimmunization againstthe rare platelet-specific antigen HPA-9b (Max) isan important cause of neonatal alloimmunethrombocytopenia.Transfusion 2005;45:1487–95.

15. Morel-Kopp MC, Daviet L, McGregor J, Kaplan C.Drawbacks of the MAIPA technique incharacterising human antiplatelet antibodies.Blood Coagul Fibrinolysis 1996;7:144–6.

16. Kroll H, Penke G, Santoso S. Functional hetero-geneity of alloantibodies against the humanplatelet antigen (HPA)-1a.Thromb Haemost 2005;94:1224–9.

17. Valentin N, Vergracht A, Bignon JD, et al. HLA-DRw52a is involved in alloimmunization againstPL-A1 antigen. Hum Immunol 1990;27:73–9.

18. L’Abbe D, Tremblay L, Filion M, et al.Alloimmunization to platelet antigen HPA-1a (PlA1)is strongly associated with both HLA-DR3*0101and HLA-DQB1*0201. Hum Immunol 1992;34:107–14.

19. Kuijpers RWAM,von dem Borne AEGKr,KiefelV,etal. Leucine33-proline33 substitution in humanplatelet glycoprotein IIIa determines HLA-DRw52a (Dw24) association of the immuneresponse against HPA-1a (Zwa/PlA1) and HPA-1b(Zwb/PlA2). Hum Immunol 1992;34:253–6.

20. Maslanka K, Yassai M, Gorski J. Molecularidentification of T cells that respond in a primarybulk culture to a peptide derived from a plateletglycoprotein implicated in neonatal alloimmunethrombocytopenia. J Clin Invest 1996;98:1802–8.

21. Wu S, Maslanka K, Gorski J. An integrinpolymorphism that defines reactivity withalloantibodies generates an anchor for MHC classII peptide binding: a model for unidirectionalalloimmune responses. J Immunol 1997;158:3221–6.

22. Semana G, Zazoun T, Alizadeh M, et al. Geneticsusceptibility and anti-human platelet antigen 5balloimmunization. Role of HLA class II and TAPgenes. Hum Immunol 1996;46:114–9.

23. Westman P, Hashemi-Tavoularis S, Blanchette V, etal. Maternal DRB1*1501, DQA1*0102, DQB1*0602haplotype in fetomaternal alloimmunizationagainst human platelet alloantigen HPA-6b (GPIIIa-Gln489).Tissue Antigens 1997;50:113–8.

24. Spencer JA,Burrows RF.Feto-maternal alloimmunethrombocytopenia: a literature review andstatistical analysis. Aust N Z J Obstet Gynaecol2001;41:45–55.

25. Kiefel V, Bassler D, Kroll H, et al. Antigen-positiveplatelet transfusion in neonatal alloimmunethrombocytopenia (NAIT). Blood 2006;107:3761–3.

26. Allen D,Verjee S, Rees S, Murphy MF, Roberts DJ.Platelet transfusion in neonatal alloimmunethrombocytopenia. Blood 2007;109:388–9.

C. KAPLAN


Neonatal alloimmune thrombocytopenia

27. Daffos F, Capella-Pavlosky M, Forestier F. A newprocedure for fetal blood sampling in utero:preliminary results. Am J Obstet Gynecol 1983;146:985–7.

28. Forestier F, Daffos F, Galacteros F, et al.Hematological values of 163 normal fetusesbetween 18 and 30 weeks of gestation.Pediatr Res1986;20:342–6.

29. Daffos F, Forestier F, Muller JY, et al. Prenataltreatment of alloimmune thrombocytopenia.Lancet 1984;2:632.

30. Kaplan C,Daffos F,Forestier F,et al.Management ofalloimmune thrombocytopenia: antenatal diag-nosis and in utero transfusion of maternalplatelets. Blood 1988;72:340–3.

31. Hohlfeld P, Forestier F,Kaplan C,Tissot JD,Daffos F.Fetal thrombocytopenia: a retrospective survey of5,194 fetal blood samplings. Blood 1994;84:1851–6.

32. Murphy MF, Pullon HW, Metcalfe P, et al.Management of fetal alloimmune thrombo-cytopenia by weekly in utero platelettransfusions.Vox Sang 1990;58:45–9.

33. Overton TG, Duncan KR, Jolly M, Letsky E, FiskNM. Serial aggressive platelet transfusion for fetalalloimmune thrombocytopenia: platelet dynamicsand perinatal outcome. Am J Obstet Gynecol2002;186:826–31.

34. Bussel JB, Berkowitz RL, Lynch L, et al. Antenatalmanagement of alloimmune thrombocytopeniawith intravenous gamma-globulin: a randomizedtrial of the addition of low-dose steroid tointravenous gamma-globulin.Am J Obstet Gynecol1996;174:1414–23.

35. Kaplan C, Murphy MF, Kroll H, Waters AH. Feto-maternal alloimmune thrombocytopenia: ante-natal therapy with IvIgG and steroids—morequestions than answers. Br J Haematol 1998;100:62–5.

36. Birchall JE, Murphy MF, Kaplan C, Kroll H, onbehalf of the European Fetomaternal AlloimmuneThrombocytopenia Study Group. Europeancollaborative study of the antenatal managementof feto-maternal alloimmune thrombocytopenia.Br J Haematol 2003;122:275–88.

37. Engelfriet CP, Reesink HW, Kroll H, et al. Prenatalmanagement of alloimmune thrombocytopenia ofthe fetus.Vox Sang 2003;84:142–9.

38. Paidas MJ,Berkowitz RL,Lynch L,et al.Alloimmunethrombocytopenia: fetal and neonatal lossesrelated to cordocentesis. Am J Obstet Gynecol1995;172:475–9.

39. Radder CM, Brand A, Kanhai HH. A less invasivetreatment strategy to prevent intracranialhemorrhage in fetal and neonatal alloimmunethrombocytopenia. Am J Obstet Gynecol 2001;185:683–8.

40. Kanhai HHH,van den Akker ESA,Walther FJ,BrandA. Intravenous immunoglobulins without initialand follow-up cordocentesis in alloimmune fetaland neonatal thrombocytopenia at high risk forintracranial hemorrhage. Fetal Diagn Ther2006;21:55–60.

41. Berkowitz RL,Kolb EA,McFarland JG, et al. Parallelrandomized trials of risk-based therapy for fetalalloimmune thrombocytopenia. Obstet Gynecol2006;107:91–6.

42. Turner ML, Bessos H, Fagge T, et al. Prospectiveepidemiologic study of the outcome and cost-effectiveness of antenatal screening to detectneonatal alloimmune thrombocytopenia due toanti-HPA-1a.Transfusion 2005;45:1945–56.

43. Dreyfus M, Kaplan C,Verdy E, et al. Frequency ofimmune thrombocytopenia in newborns: aprospective study. Blood 1997;89:4402–6.

44. Bertrand G, Martageix C, Jallu V,Vitry F, Kaplan C.Predictive value of sequential maternal anti-HPA-1a antibody concentrations for the severity of fetalalloimmune thrombocytopenia. J ThrombHaemost 2006;4:628–37.

45. Ni H, Chen P, Spring CM, et al. A novel murinemodel of fetal and neonatal alloimmunethrombocytopenia: response to intravenous IgGtherapy. Blood 2006;107:2976–83.

Cecile Kaplan, MD, Platelet Immunology Laboratory,Institut National de la Transfusion Sanguine (INTS),6 Rue Alexandre Cabanel, 75015 Paris, France.


During the last two decades, there has been greatinterest in developing and using platelet additivesolutions (PASs) for the storage of platelets.1–5 Atpresent, such additive solutions are in use fortransfusion in several countries.PAS is generally used asa substitute for plasma to (1) reduce the amount ofplasma transfused with platelets and to recover plasmafor other purposes, primarily fractionation into plasmaproducts; (2) avoid transfusion of large volumes ofplasma with possible adverse reactions and circulatoryoverload; (3) make possible photochemical treatmentfor the inactivation of bacteria and other pathogens inplatelets; and (4) improve storage conditions as wasdiscussed in a previous review.6

A basic principle is that aging of platelets after invitro storage at 22°C is significantly slower than agingof platelets in vivo at 37°C, a situation that may makeextended storage of platelets possible.7 Threeapproaches were suggested to be of specific impor-tance to improve storage conditions of platelets6: (1)reducing the activation of platelets during collection ofblood and the preparation and storage of platelets; (2)reducing the metabolic rate in terms of glucoseconsumption and lactate production; and (3) ensuringthat glucose will be available in the storage mediumduring the entire storage period. The activation ofplatelets can be counteracted either by the addition ofplatelet activation inhibitors or of certain componentssuch as magnesium to the PAS.

The use of PAS offers the possibility of includingcomponents with specific effects on platelets in thestorage solution that are not present in plasma or in theanticoagulant. A number of effects have been observedthat can be assigned to certain components.4,5

Reducing platelet activation and inclusion of keycomponents in the platelet storage environment, suchas acetate, citrate, glucose, potassium, and magnesium,were suggested to be useful tools to optimize plateletstorage conditions. The compositions of some presentPASs are presented in Table 1. The purpose of this

review is to describe some events of the last severalyears as a complement to the knowledge presented inprevious reviews.1–6

In Vivo CharacteristicsResults from a number of in vivo studies using PASs

have been published during the last decade. In the firstpatient transfusion studies in the 1990s using PASssuch as PAS-II (T-Sol, Baxter) or Plasma Lyte A (Table 1),the results were not consistent.8–14 In some studies,satisfactory CCIs were found in patient transfusionstudies,9,11 in some studies significantly lower CCIswere observed than for platelets stored in plasma.12,14

In a recent study, encouraging platelet recovery andsurvival data were found,comparing platelets preparedby apheresis and stored in PAS-II for 1 versus 7 days.15

Mean recovery was 69 percent and 53 percent, andsurvival was 8.2 and 5.1 days at Days 1 and 7,respectively. The ratios of Day 7 to Day 1 were 0.80and 0.65 for recovery and survival, respectively. Aproposal by Murphy16 of a new standard of efficacy forthe evaluation of platelets for transfusion has createdconsiderable interest. This concept implies thatacceptably stored platelets on the last day of storagewould demonstrate at least two-thirds the recovery andone-half the survival of platelets collected from thesame subject and then retransfused as reference“fresh”platelets. The ratios in this study15 met the proposedcriteria for 7-day storage, although the use of a Day 1control may not totally have fulfilled the requirementsof the reference indicated.16

Comprehensive patient transfusion studies usingpathogen-reduced platelets have added significantknowledge about platelets stored in more recent PASs,particularly PAS-III (InterSol, Baxter). A photochemicaltreatment method using a novel psoralen, amotosalenHCl, in combination with ultraviolet light has beendeveloped to inactivate viruses, bacteria, protozoa, andWBCs in platelets.17 Two major studies wereconducted in Europe and in the United States, namely

Platelet additive solutions:current statusH.GULLIKSSON


Platelet additive solutions

the euroSPRITE trial in Europe and the SPRINT trial inthe United States.18–21 In the euroSPRITE trial,comparable platelet CIs and CCIs and a comparablesafety profile for photochemically treated andreference untreated pooled buffy-coat (BC) plateletunits in 103 oncology patients with thrombocytopeniawere demonstrated. In the SPRINT study, whichincluded 645 transfused patients, the hemostaticefficacy of photochemically treated and referenceuntreated platelets was found to be equivalent.19

Transfusion reactions were significantly fewer, andadverse events and overall safety profiles werecomparable for treated and reference platelets.20 In asecond evaluation of data from the euroSPRITE patienttransfusion study using BC-derived platelets, referencegroups with untreated platelets were compared.22

Reference platelets were stored in either T-Sol (n=35)or plasma (n=16). The results indicated that 1-hour and24-hour mean platelet CIs and posttransfusionhemostatic scores were not significantly different forpatients receiving platelet components suspended in100 percent plasma and patients receiving platelets ina T-Sol environment. In a review in 2003, thepreliminary conclusion had been that additional in vivodata, from studies of clinical outcome after transfusionof platelets stored in either PAS or plasma, wereneeded/required.6 Today, comprehensive clinical datasupport the routine use of PAS as an equivalentalternative to plasma to suspend platelets.

In Vitro Platelet Aging at 22°C Versus In VivoAging at 37°C

Because the average survival of human platelets inthe circulation, after release from megakaryocytes, isknown to be 9 to 10 days, it may be questionedwhether it is at all meaningful to extend the shelf lifeof platelet concentrates up to or even beyond 7 days.23

Removal of platelets from the circulation isbelieved to be age-dependent and isprobably associated with failure to maintainnormal hemostasis. Transfusion of agingplatelets may be of little help to stopbleeding because they would be removedfrom the circulation.

Studies comparing aging of platelets invivo with in vitro storage at 22°C, usingisotope labeling, suggested that aging ofplatelets after in vitro storage for 5 days at22°C was similar to aging of platelets during2.1 days in vivo at 37°C.7 The relative aging

factor was found to be 0.42 (2.1 days divided by 5.0days). In a subsequent study, a similar relative agingfactor was found (0.44).7 These differences wereassociated with a much higher turnover rate of ATP, themajor energy carrier in platelets, at 37°C than at 22°C.The calculated ATP turnover ratio of 0.48 at those twotemperatures was of the same magnitude as thecalculated relative aging factor, indicating that therelative decrease in aging of platelets at 22°C comparedwith that at 37°C is similar to the relative decrease inmetabolic rate at this temperature. The rate of aging ofplatelets during storage at 22°C may be less than half ofthat found in vivo at 37°C. If this ratio is applied to thenormal lifespan of platelets, 9 to 10 days in thecirculation would correspond to at least 18 to 20 daysof in vitro storage at 22°C. These data may providemeaningful objectives to develop methods and storageenvironments for extended storage of platelets.

The Use of PAS in Combination WithDifferent Methods for the Preparation ofPlatelets for Transfusion

In general, the various PASs can be used forapheresis as well as for BC platelets. The storagemedium normally is composed of a mixture of plasma(generally 20–40%) and PAS (60–80%). The maindifference is that in apheresis platelets, ACD is oftenpreferred to CPD anticoagulant, because resuspensionof platelets after preparation is facilitated. In someapheresis equipment, platelets are kept in thecentrifugation chamber during the entire apheresiscycle; in other equipment, platelets are continuouslytransferred to a platelet storage container. Thesedifferences may result in significant variation in plateletactivation. Although apheresis equipment originallymay have been designed for the preparation ofplatelets suspended in plasma,most equipment can be

Table 1. Composition of some current PASs (in mmol/L) including commercialdesignations*

Plasma PAS-II PAS-III Composol PAS-III MLyte A (T-Sol, (InterSol, (Fresenius) (SSP+,

Baxter) Baxter) MacoPharma)

NaCl 90 116 77 90 69

KCl 5 – – 5 5

MgCl2 3 – – 1.5 1.5

Na3-citrate – 10 10 11 10

NaH2PO4/Na2HPO4 – – 26 – 26

Na-acetate 27 30 30 27 30

Na-gluconate 23 – – 23 –

*The compositions of commercial solutions may be slightly different from basic compositions.


H. GULLIKSSON

used for platelet collection in a small final resuspensionvolume of plasma, which is necessary to allow theaddition of PAS. Satisfactory results were obtained byRingwald et al.24,25 when evaluating the suspension ofhigh concentrations of apheresis platelets (4000–5000× 109 platelets/L) in PAS.

BC-derived platelets suspended in PAS are generallyprepared in pools from several donors. There areprimarily variations in the number of BCs included inthe pools (generally 3–7 BCs) and in the time ofholding of whole blood preceding the preparation ofBCs. Either BCs can be prepared on the day ofcollection, generally within 8 hours, and storedovernight for the preparation of platelets on thefollowing day, or whole blood can be stored overnightfor the preparation of BCs and platelets on thefollowing day.

The platelet rich plasma (PRP) method is generallynot used for the routine preparation of platelets in PAS.However, in a recent study by Sweeney et al.,26 plateletswere prepared from individual donors by the PRPmethod in a first step and then pooled as platelets fromseveral donors and suspended in either plasma or PAS.Platelets were stored for 7 days for in vitro evaluation.Good preservation of platelet quality in bothenvironments was reported.

Effects on Metabolism and Platelet FunctionAssociated With Components in PAS

The results of early studies by Holme et al.27 andGulliksson et al.28 on the effects of PAS indicated thatthe presence of glucose during the entire plateletstorage period is crucial for platelet metabolism.Effects observed after depletion of glucose involvedrapid decrease in adenine nucleotide levels, cessationof lactate production, and finally disintegration ofplatelets. Depletion of glucose is generally associatedwith an increased rate of platelet metabolism and fall inpH levels. The results suggested that depletion ofglucose, not the pH level alone, is detrimental toplatelets during storage. In contrast to plasma, the fallin pH during storage of platelets in PAS will stop at asignificantly higher level than pH 6.0,often at about pH6.5 as a result of the limited amount of glucosegenerally available in PAS.28 Because the bufferingcapacity of PAS is approximately half that of plasma,PAS is more susceptible to increased production oflactic acid by platelets.29,30 On the other hand,metabolism of acetate present in PAS significantlystabilizes the pH level. In parallel with glucose, acetate

is used as a substrate for the oxygen-dependent plateletmetabolism, and enters into the tricarboxylic acidcycle, and is further oxidized in the respiratorychain.29–32 The end products are carbon dioxide fromthe first step and water from the second step. Byformation of bicarbonate from the carbon dioxideproduced by acetate, very stable pH levels aremaintained during platelet storage.29,30,32 A possiblethird substrate for platelet metabolism may be fattyacids.33

Generally, phosphate has two possible roles duringstorage of platelets, as a stimulant of platelet glycolysisto increase production of lactic acid and as a buffer toprevent a fall in pH. These two effects theoreticallycompete and may neutralize each other. There are noindications of net utilization or production ofphosphate during storage of platelets.29

The new PASs designated Composol and PAS-IIIMas well as the early Plasma Lyte A all containmagnesium and potassium. Composol and PAS-IIIMalso contain citrate, in contrast to Plasma Lyte A. Thethree components: magnesium, potassium, and citrateare associated with complexity of effects andinterdependence. Effects on platelet membranefunction and platelet activation as well as rate ofglycolysis have been described and the various effectsmay even be combined.

Increased concentration of extracellularmagnesium ions significantly inhibits exposure of P-selectin, decreases binding of fibrinogen to ADP-activated platelets, and significantly decreases ADP-induced platelet aggregation.34 Magnesium may alsomodify calcium influx into the platelets. In addition,magnesium, calcium, and the concentration of citratehave an effect on potassium permeability of theplatelet membrane and the intracellular concentrationof potassium.35 Citrate also heightens plateletresponsiveness to some activating agents such as ADP.36

In addition, effects on the rate of glucose consumptionand lactate production related to the concentration ofcitrate and the presence of potassium and magnesiumin PAS have been observed. Platelets stored in mediumwith a citrate concentration of 8 mmol/L producedonly half the quantity of lactate produced by plateletsin a similar medium with a citrate concentration of 14to 26 mmol/L.37 Inasmuch as no negative effects onadenine nucleotide levels were observed, the resultssuggested that synthetic media preferably shouldinclude citrate at low concentrations to avoid excessivelactate production and an acid pH. On the other hand,


increased production of lactate associated with higherconcentrations of citrate can be neutralized by theaddition of acetate.37 Again, this situation illustrates thecomplexity of effects and interdependence associatedwith those components. In parallel, the combination ofmagnesium and potassium ions present in PAS has asimilar effect on platelets, i.e., significantly reducedmetabolic rate in terms of glucose consumption andlactate production and also reduced plateletactivation.38–40

When platelets are shipped between differentcenters,agitation may be interrupted for a considerableperiod of time. A previous study using a plasmaenvironment suggests that possible storage timewithout agitation is strongly affected by plateletconcentration.41 The level of pH should be kept above6.5 to avoid negative effects on functional in vitrofactors such as hypotonic shock response (HSR).41 In arecent study, the effects of interruption of agitation onplatelets stored in two PAS alternatives, namely PAS-IIIM and Composol, were evaluated.42 At a plateletconcentration of approximately 1000 × 109/L, plateletscould be stored for 4 days in PAS-IIIM withmaintenance of HSR and pH. This was not possiblewith Composol, suggesting that the presence ofphosphate is of importance to maintaining pH andother in vitro characteristics during interruption ofagitation. Similar effects were not observed duringcontinuous agitation.

Future PerspectivesTo conclude, present knowledge and experience

support platelet storage only at 22°C. The use of PASinstead of plasma as the platelet storage environmentwould provide benefits to patients and facilitate theinclusion of certain components with proven favorableeffects on platelets that are present in neither plasmanor anticoagulant. The present knowledge of effectsassociated with different approaches discussed abovesuggests that reducing platelet activation incombination with the inclusion of key components inthe platelet storage medium, such as acetate, citrate,glucose,potassium,magnesium,and additional possiblefuture ingredients, should be the tools to optimize thestorage conditions and maintain the function ofplatelets. Additional in vivo studies of recovery andsurvival as well as count increments in patients withthrombocytopenia will be needed.

References1. Holme S. Effect of additive solutions on plateletbiochemistry. Blood Cells 1992;18:421–30.

2. Holme S. Platelet storage in liquid environment.Transfus Sci 1994;15:117–30.

3. Högman CF. Aspects of platelet storage. TransfusSci 1994;15:351–5.

4. Murphy S. The efficacy of synthetic media in thestorage of human platelets for transfusion.Transfus Med Rev 1999;13:153–63.

5. Gulliksson H.Additive solutions for the storage ofplatelets for transfusion. Transfus Med 2000;10:257–64.

6. Gulliksson H. Defining the optimal storageconditions for the long-term storage of platelets.Transfus Med Rev 2003;17:209–15.

7. Holme S, Heaton WA. In vitro platelet ageing at22°C is reduced compared to in vivo ageing at37°C. Br J Haematol 1995;91:212–8.

8. Oksanen K, Ebeling F, Kekomäki R, et al. Adversereactions to platelet transfusions are reduced byuse of platelet concentrates derived from buffycoat.Vox Sang 1994;67:356–61.

9. van Rhenen DJ,Vermeij K,Kappers-Klunne R,et al.Evaluation of a new citrate-acetate-NaCl plateletadditive solution for the storage of white cell-reduced platelet concentrates obtained from half-strength CPD pooled buffy coats. Transfusion1995;35:50–3.

10. Eriksson L,Kristensen J,Olsson K, et al. Evaluationof platelet function using the in vitro bleedingtime and corrected count increment of transfusedplatelets: comparison between plateletconcentrates derived from pooled buffy coats andapheresis.Vox Sang 1996;70:69–75.

11. Strindberg J, Berlin G. Transfusion of plateletconcentrates: clinical evaluation of twopreparations. Eur J Haemotol 1996;57:307–11.

12. Turner VS, Mitchel SG, Hawker RJ. More on thecomparison of Plasma-Lyte A and PAS-2 as plateletadditive solutions.Transfusion 1996;36:1033–4.

13. Högman CF, Eriksson L,Wallvik J, et al. Clinical andlaboratory experience with erythrocyte andplatelet preparations from a 0.5 CPD Erythro-solOpti System.Vox Sang 1997;73:212–9.

14. de Wildt-Eggen J, Nauta S, Schrijver JG, et al.Reactions and platelet increments aftertransfusion of platelet concentrates in plasma oran additive solution: a prospective, randomisedstudy.Transfusion 2000;40:398–403.



15. Shanwell A, Diedrich B, Falker C, et al. Paired invitro and in vivo comparison of apheresis plateletconcentrates stored in platelet additive solutionfor 1 versus 7 days.Transfusion 2006;46:973–9.

16. Murphy S.Radiolabeling of PLTs to assess viability:a proposal for a standard. Transfusion 2004;44:131–3.

17. Lin L, Cook DN, Wiesehahn GP, et al.Photochemical inactivation of viruses andbacteria in platelet concentrates by use of a novelpsoralen and long-wavelength ultraviolet light.Transfusion 1997;37:423–35.

18. van Rhenen, D, Gulliksson H, Cazenave JP, et al.Transfusion of pooled buffy coat plateletcomponents prepared with photochemicalpathogen inactivation treatment: the euroSPRITEtrial. Blood 2003;101:2426–33.

19. McCullough J, Vesole DH, Benjamin RJ, et al.Therapeutic efficacy and safety of platelets treatedwith a photochemical process for pathogeninactivation. The SPRINT Trial. Blood 2004;104:1534–41.

20. Snyder E, McCullough J, Slichter JS, et al. Clinicalsafety of platelets photochemically treated withamotosalen HCl and ultraviolet A light forpathogen inactivation: the SPRINT trial.Transfusion 2005;45:1864–75.

21. Janetzko K, Cazenave JP, Klüter H, et al.Therapeutic efficacy and safety of photo-chemically treated apheresis platelets processedwith an optimized integration set. Transfusion2005;45:1443–52.

22. van Rhenen, DJ, Gulliksson H, Cazenave JP, et al.Therapeutic efficacy of pooled buffy-coat plateletcomponents prepared and stored with a plateletadditive solution.Transfus Med 2004;14:289–95.

23. Gewirtz AM, Schick B. Platelet production andfunction: megakaryocytopoiesis. In: Colman RW,Hirsh J, Marder VJ, et al., eds. Hemostasis andthrombosis. 3rd ed. Philadelphia: JB Lippincott,1994:353–96.

24. Ringwald J, Walz S, Zimmermann R, et al.Hyperconcentrated platelets stored in additivesolution: aspects on productivity and in vitroquality.Vox Sang 2005;89:11–8.

25. Ringwald J, Duerler T, Frankow O, et al. Collectionof hyperconcentrated platelets with Trima Accel.Vox Sang 2006;90:92–6.

26. Sweeney J, Kouttab N, Holme S, et al. Storage ofplatelet-rich plasma-derived platelet concentratepools in plasma and additive solution.Transfusion2006;46:835–40.

27. Holme S, Heaton WA, Courtright M. Plateletstorage lesion in second-generation containers:correlation with platelet ATP levels. Vox Sang1987;53:214–20.

28. Gulliksson H,Sallander S,Pedajas I, et al. Storage ofplatelets in additive solutions: a new method forstorage using sodium chloride solution.Transfusion 1992;32:435–40.

29. Shimizu T, Murphy S. Roles of acetate andphosphate in the successful storage of plateletconcentrates prepared with an acetate-containingadditive solution.Transfusion 1993;33:304–10.

30. Bertolini F, Murphy S, Rebulla P, et al. Role ofacetate during platelet storage in a syntheticmedium.Transfusion 1992;32:152–6.

31. Guppy M, Whisson ME, Sabaratnam R, et al.Alternative fuels for platelet storage: a metabolicstudy.Vox Sang 1990;59:146–52.

32. Murphy S. The oxidation of exogenously addedorganic anions by platelets facilitates maintenanceof pH during their storage for transfusion at 22°C.Blood 1995;85:1929–35.

33. Cesar J, DiMinno G,Alam I, et al. Plasma free fattyacid metabolism during storage of platelet con-centrates for transfusion. Transfusion 1987;27:434–7.

34. Gawaz M, Ott I, Reininger AJ, et al. Effects ofmagnesium on platelet aggregation and adhesion.Thromb Haemost 1994;72:912–8.

35. Weis-Fogh US. The effect of citrate, calcium, andmagnesium ions on the potassium movementacross the human platelet membrane.Transfusion1985;25:339–42.

36. Kinlough-Rathbone RL, Packham MA, Mustard JF.Platelet aggregation. In: Harker LA, ZimmermanTS, eds.Methods of hematology:measurements ofplatelet function.NewYork:Churchill Livingstone,1984:64–91.

37. Gulliksson H. Storage of platelets in additivesolutions: the effect of citrate and acetate in invitro studies.Transfusion 1993;33:301–3.

38. deWildt-Eggen J, Schrijver JG,Bins M,et al. Storageof platelets in additive solutions: effects ofmagnesium and potassium. Transfusion 2000;42:76–80.

H. GULLIKSSON



39. Gulliksson H, AuBuchon JP, Vesterinen M, et al.Storage of platelets in additive solutions: a pilotstudy of the effects of potassium and magnesium.In vitro studies.Vox Sang 2002;82:131–6.

40. Gulliksson H, AuBuchon JP, Cardigan R, et al.Storage of platelets in additive solutions: amulticentre study of the in vitro effects ofpotassium and magnesium. Vox Sang 2003;85:199–205.

41. Hunter S, Nixon J, Murphy S. The effect of theinterruption of agitation on platelet quality duringstorage for transfusion. Transfusion 2001;41:809–14.

42. van der Meer PF, Gulliksson H,AuBuchon JP, et al.Interruption of agitation of platelet concentrates:effects on in vitro parameters. Vox Sang 2005;88:227–34.

Hans Gulliksson, PhD, Department of ClinicalImmunology and Transfusion Medicine C2 66,Karolinska University Hospital, Huddinge, SE-141 86Stockholm, Sweden.

Phone, Fax, and Internet Information: If you have any questions concerning Immunohematology,Journal of Blood Group Serology and Education, or the Immunohematology Methods and Proceduresmanual,contact us by e-mail at [email protected] information concerning the National ReferenceLaboratory for Blood Group Serology, including the American Rare Donor Program, please contact SandraNance, by phone at (215) 451-4362, by fax at (215) 451-2538, or by e-mail at [email protected]


ABO and platelet transfusiontherapyL.COOLING

It is appropriate to take steps to optimize the efficacyand safety of transfusion therapy. Perhaps we shouldstudy the role of ABO compatibility in platelettransfusion therapy more carefully so that furtherimprovements may be made.

Scott Murphy, Transfusion Editorial 19881

The relative importance of ABO compatibility andplatelet transfusion has been a matter of debate formore than 50 years. Since the early studies of RichardAster,2 it has been recognized that transfusion of ABO-incompatible platelets can be associated with decreasedplatelet increments after transfusion. In the last 10 to 15years, there has been increasing discussion and concernregarding minor-incompatible platelet transfusions thathave been linked to acute hemolytic transfusionreactions, venoocclusive disease, and increasedmorbidity in allogeneic transplants.3–5 This review willbriefly summarize the biochemistry, regulation, clinicalpractice, and considerations surrounding ABOcompatibility and platelet transfusions.

ABO Glycoconjugates on PlateletsABO antigens are expressed on several endogenous

platelet glycoproteins and glycolipids. Platelet-specificglycoproteins (GP) known to express ABH antigensinclude GPIIb/IIIa, GPIb/IX, GPIa/IIa, GPIc, GPIV, GPV,CD31 (PECAM), and CD109.6–10 GPIIb/IIIa, inparticular, appears to be a significant contributor anddeterminant of ABH expression on platelets. TheGPIIb/IIIa complex numbers nearly 250,000 moleculesper platelet and possesses between five and eight bi-and triantennary N-glycans capable of displaying ABHepitopes.10,11 GPIIb/IIIa is the major target of humananti-A and anti-B in vitro.7 Likewise, a linear correlationbetween A antigen on GPIIb and intact plateletmembranes (R = 0.91) was reported by Cooling et al.10

Translocation and ongoing synthesis of GPIIb/IIIa mayalso contribute to the increased antigenicity of

platelets during in vitro storage. Julmy et al.12 reporteda nearly 50 percent increase in ABH expression onplatelets during routine storage, accompanied byevidence of platelet activation and translocation of α-granule proteins (including GPIIb/IIIa) to the plateletmembrane. More recent studies in platelet proteomicsand the platelet transcriptome suggest ongoingtranslation and synthesis of GPIIb/IIIa during storage aswell.13

Platelets also express a small population ofglycosphingolipids with ABH activity. Early studiesshowed the ability of platelets to adsorb and elutesoluble type 1 chain ABH and Lewis antigens fromplasma, leading to the speculation that all ABH-activeglycolipids were entirely of exogenous origin.14

Subsequent studies using chain-specific monoclonalantibodies showed that platelets expressed type 2chain glycolipids and glycoproteins, indicating thatmost ABH-active glycolipids were of megakaryocyte orendogenous origin.6,10,15,16 This was confirmed byCooling et al.10 who identified ABH-active glycolipids inplatelets from Le(a+b–) individuals who should lacksoluble type 1 chain antigen. The ABH-activeglycosphingolipids of platelet are predominantlysimple, linear structures (6–8 oligosaccharides)although a few complex sialylated structures arepresent.10,17,18 In group A1 donors, type 4 chain orglobo-ABH structures have also been identified.16,19

This is in sharp contrast to RBCs,which express a richvariety of ABH-active glycolipids, including complex,highly branched polyglycosylceramides.19,20 Thecontribution of ABH-active glycolipids to overall ABHexpression on platelets appears to be minor.10

ABO Expression on MegakaryocytesAlthough recent studies indicate some residual

protein synthesis in young platelets,13 the majority ofABH synthesis occurs at the level of the megakaryocyte.ABH antigens have been identified on immortalized


ABO and platelets

megakaryoblastic cell lines and cultured bone marrowmegakaryocytes.21,22 In the latter, ABH exhibitsdevelopmental and distinct clonal variation, withindividual megakaryocyte colonies differing widely inthe amounts of A, B, and H antigens expressed.21 Clonalvariation may underlie the heterogeneity observed oncirculating platelets.10,21 In addition, there is a directcorrelation between A and H on megakaryocytes, withcolonies either positive or negative for both H and Aantigens. Cooling et al.10 subsequently showed a direct,linear relationship between H and A antigens oncirculating platelets by two-color flow cytometry (R =0.95). On the basis of their findings,Dunstan et al.21 andCooling et al.10 speculated that H antigen synthesis is arate-limiting step regulating ABH expression inmegakaryocytes and platelets.

H antigen appears to be developmentally regulatedin early hematopoiesis. H, LeY, and FUT1 mRNA areexpressed by CD34+ cells and likely early mega-karyocyte progenitors.22–24 During megakaryocyticdifferentiation, both H and LeY antigens areprogressively lost with increasing ploidy andproplatelet formation.22 The presence of H antigen onearly megakaryocyte progenitors may facilitateadhesion to stromal fibroblasts,25 a necessary step formegakaryocytic preservation and expansion.26,27

Schmitz and colleagues25 were able to inhibitmegakaryocyte adhesion to stromal fibroblasts withfucosylated bovine albumin, soluble H antigen, andfucose-specific lectins. Although the precise lectin-ligand pair involved in fucose-mediated adhesion is notyet identified, several megakaryocyte proteins criticalto megakaryocyte development and adhesion canexpress ABH antigens, including α4β1,α5β1 GPIIb/IIIa,GPIb, and PECAM.28,29 H antigen on integrins may beparticularly important: H and LeY expression areassociated with increased fibronectin binding, celladhesion, and resistance to apoptosis in epithelialtissues.30–32 An increase in fucose-mediated adhesion,with delayed megakaryocyte maturation, may accountfor delayed platelet engraftment in some group Oautologous transplant patients.33

Platelet ABO and Genetic VariationThere are dramatic differences in the amount of

ABO expressed on platelet membranes betweenindividual donors. Genetic differences in ABOglycosyltransferase alleles underlie some of thesedifferences. In group A donors, expression of A antigenon platelet membranes, glycoproteins, and glycolipids

is linked to an A1 RBC phenotype.7,10,16,34,35 In contrast,A2 donors express little or no A antigen and can beconsidered group O compatible.10,35 Lewis andsecretor phenotype appear to play little role in eitherthe presence or strength of ABO antigens on platelets:A2, Le(a–b+) donors are also negative for A antigen byflow cytometry.10 Because they lack A antigen onplatelets,A2 individuals can develop an anti-A1 and ABH-specific refractoriness.34

Among group A1 donors, there is a wide variation inplatelet A expression (Fig. 1E).7,10,35 In individualdonors, the percentage of circulating platelets positivefor A antigen by two-color flow cytometry (CD41+,HPA+) can range from less than 5 percent to 87percent (population average = 40%).10 Even within asingle donor, there is distribution of positive andnegative platelets (Fig. 1B). Despite the latter, theaverage percentage of platelets positive for A antigen isrelatively stable over time and may represent a unique,donor-specific characteristic.10 As shown in Figure 1F,the percentage of A antigen–positive platelets, in pairedsamples (S1, S2) collected from the same donor at twoseparate times, is nearly identical.

Unlike A1 donors, most group B donors weaklyexpress ABO antigens on platelets.7,10,36 In our ownstudies, only 20 percent of platelet donors werepositive for B antigen by flow cytometry.10 Similarfindings are observed with platelet crossmatching, inwhich 83 to 100 percent of group B platelet donorswere crossmatch-compatible with group O and groupA serum, respectively.36 Despite these results, group Bantigen can be identified on platelet glycoproteins andglycosphingolipids in most group B donors (80–90%).10

The density or antigenicity on individual plateletglycoproteins, however, may be decreased relative tomost A1 donors (< 50%),on the basis of solid phase andradioimmunoassay studies with human anti-B.7

High-Expressor PhenotypeApproximately 4 to 8 percent of A1 and B donors

are ABO high expressors (HXP),defined as a 2- to 3-foldincrease in platelet ABO expression (> 2 SD).10,34,37 ABOHXPs can be further subclassified as type 1 and type 2on the basis of their flow cytometry profiles.10,35 Type1 donors demonstrate a bimodal population of stronglypositive and moderately to weakly positive platelets,akin to the bimodal population of positive and negativeplatelets observed in most A1 donors (Fig. 1B). Incontrast, type 2 donors have an essentially uniformpopulation of strongly positive platelets (Fig. 1C).


L. COOLING

Transfusion of ABO-HXP platelets to a group Orecipient has resulted in profound transfusion failures,even with HLA-matched platelets.37 Platelets fromABH-HXP donors should be reserved for ABO-identicalrecipients only.

The molecular origin of the ABO-HXP phenotype isstill unknown. Family studies indicate that the trait isautosomal dominant.37 There is no relation betweenthe ABO-HXP and a secretor phenotype.10,37 ElevatedABO glycosyltransferase activity has been noted in thesera of some donors35,37;however, sequencing studies ofthe ABO gene have not revealed any mutations.35

Cooling et al.10 suggested that the ABO-HXP mightrepresent clonal upregulation of the H or FUT1 gene.Future studies focusing on transcriptional regulation ofH and ABO genes in megakaryocytes, including

epigenetic phenomena, may eventually unravel thegenetic mystery behind the ABO-HXP phenotype.

ABO Compatibility and Platelet TransfusionResponse

The influence of platelet ABO and the post-transfusion response is highly variable, reflecting bothdonor and recipient factors. Recipient factors thatmay influence the transfusion response include ABOtype, sex, and parity; isohemagglutinin titers; andimmunosuppressive drugs. Platelet and donor factorscan include ABO type (A1, A2, B), donor-specificdifferences in ABO strength, platelet activation, andstorage time. As noted in the previous section, ABOexpression varies widely between individual donorsand ABO types.10,35

Fig. 1. Donor-specific differences in ABO expression. A–C: Sample histograms of platelets from A2 (A),A1 (B), and an A1 high-expressor donor (A1-HXP,C).Platelets were dual labeled with a platelet-specific monoclonal antibody (CD41) and the anti-A lectin,Helix pomatia (HPA). For analysis, plateletswere gated on CD41 and the percent CD41+, HPA+ platelets determined by two-color flow cytometry. D and E: Expression of A antigen onplatelets in A2 (D) and A1 donors (E). Note the wide distribution of A antigen-positive platelets in A1 donors, including 5 percent with highexpression (A1-HXP). Little or no A antigen was detected in A2 and 6 percent A1 donors. F: Stable expression of A antigen on platelets.The percentHPA+ platelets in individual donors was compared in two different samples (S1, S2) at 1- to 4-month intervals.


ABO and platelets

The earliest studies examining the effect of ABOand platelet transfusion were performed by RichardAster in 1965.2 Using radiolabeled platelets,Aster et al.reported a 60 to 90 percent decrease in the 1-hourposttransfusion platelet increments after transfusion ofgroup A and AB platelets to group O recipients. Thiswas substantiated in an elegant study by Jimenez etal.,38 who compared the posttransfusion recovery inpaired recipients of split apheresis products from thesame donor. The authors found similar plateletrecoveries when both recipients were ABO compatiblewith the donor (R = 0.8); however, transfusion to anABO-incompatible recipient was accompanied bymarked, significant decreases in posttransfusionrecovery at 1 (p < 0.001), 4 (p < 0.004), and 24 hours(p < 0.04). Improved platelet recovery with ABO-identical and ABO-compatible platelets has also beenobserved in large clinical trials, including the Trial toReduce Alloimmunization to Platelets.39,40

There are several examples of plateletrefractoriness caused by ABO in the literature.34,37,41,42

In a small randomized trial of ABO-identical and ABO-unmatched platelets, the vast majority of patientsreceiving ABO-unmatched platelets had a decrease inposttransfusion recovery, with 37 percent having clearevidence of ABO-specific refractoriness.43 In a secondstudy, clinical refractoriness was significantly higher inpatients receiving ABO-mismatched platelet trans-fusions (69% vs. 8%) and was typically heralded by asudden acute rise in isoagglutinin titers.44 In a morerecent study, ABO-incompatible platelet transfusionsstimulated isohemagglutinin titers in 40 to 50 percentof patients after only one to two transfusions.45 Notsurprisingly, patients with documented ABO refrac-toriness often have high titer isohemagglutinins(> 1:500–1:1000). Platelet-associated antibody in thesepatients can approach 30,000 molecules of IgG perplatelet after transfusion of an ABO-incompatibleplatelet.7 In contrast, IgM binding to platelets isminimal (< 350 molecules/platelet).7

Although most case reports and studies havefocused on ABO major-incompatible transfusions, poorplatelet recoveries and clinical refractoriness can alsobe observed with ABO minor-incompatible or out-of-group transfusions.39,46,47 According to one study,plasma-incompatible transfusions were associated withan 18 percent decrease in posttransfusion recoverywhen compared with ABO-identical transfusions.39 Theadverse effect of incompatible-plasma infusion may becaused by the formation of immune complexes

composed of donor ABO antibodies and soluble ABOsubstances in blood.46,47 These immune complexes maythen bind to either complement or Fc receptors onplatelets, leading to accelerated immune clearance.46

Among refractory patients, 40 percent have elevatedlevels of circulating immune complexes.Among groupA patients transfused with ABO-mismatched platelets,80 percent had evidence of immune complexescontaining IgG anti-A of donor origin.47

AlloimmunizationRoutine transfusion of ABO-incompatible platelets

can promote HLA alloimmunization. In randomizedtrials, patients routinely transfused with ABO-mismatched platelets were more likely to develop bothHLA and platelet-specific antibodies, accompanied byan earlier onset and higher incidence of clinicalplatelet refractoriness.44,48 ABO incompatibility canalso act synergistically with HLA and crossmatchcompatibility to further decrease the posttransfusionresponse.36,39,49

In a retrospective study of 51 refractory patients,Blumberg and colleagues39 demonstrated decreasedplatelet recoveries in 60 percent of all ABO-incompatibletransfusions, regardless of platelet crossmatch results.Among patients receiving crossmatch-compatibleplatelets, an ABO mismatch was associated with a 40percent decrease in posttransfusion recovery. Theimpact was more dramatic with ABO-mismatched,crossmatch-incompatible platelets (85% decrease).Overall, ABO-identical platelets were clinically equiv-alent or better than ABO-incompatible, HLA-matched,or crossmatch-compatible platelets.

The synergism between ABO, HLA, and clinicalrefractoriness can have direct economic conse-quences. The University of Rochester reported a 20percent decrease in platelet utilization and a 40percent decrease in HLA-matched platelets afterinstituting a policy requiring ABO-identical plateletsfor their leukemic patients.50–52 This policy has led toimproved survival and decreased morbidity based onthe results of a small randomized trial (n = 40 patients).Leukemia patients who received only ABO-identicalplatelets had longer remissions and improved survivalrelative to patients transfused with ABO-nonidenticalplatelets.53 There was also a decrease in major bleedingepisodes (5%) with 70 percent of patients having noevidence of clinical bleeding.54

The impact and cost effectiveness of ABOcompatibility in other patient populations is still


L. COOLING

debated. In a retrospective study of cardiovascularsurgery patients, Blumberg et al.55,56 found that patientstransfused with ABO-identical platelets had decreasedmorbidity and mortality, as measured by fever,antibiotic usage, transfusion support, and hospitaladmission days. These differences corresponded to anaverage savings of $10,000 to $15,000 in direct costsand patient charges. These findings have beenchallenged by Lin et al.,57 who found no significantdifferences in either morbidity or mortality in a similargroup of cardiovascular patients. In the latter study, theauthors concluded that the use of ABO-nonidenticalplatelets is “an acceptable and safe practice” in surgicalpatients.

ABO, Platelets, and TransplantationThere is increasing concern regarding platelets and

ABO compatibility in bone marrow transplant patients,particularly in patients receiving ABO-incompatibleallogeneic transplants who have complex transfusionneeds.58–60 Because of the adverse effect of ABOincompatibility on erythroid engraftment, it is apractice to transfuse plasma and platelets that arecompatible with the stem cell or marrow donor.61 Inmost instances, this may require transfusion of an ABOmajor-incompatible or minor-incompatible platelet.Several studies have linked increased platelettransfusions and transfusion of plasma-incompatibleplatelets with increased morbidity, venoocclusivedisease, multiorgan failure, and death.4,5,62 Organdysfunction usually follows a 1- to 2-week period ofrising platelet transfusion requirements, suggesting acausal relationship.62

It is hypothesized that donor ABO antibodies mayresult in immune complex formation in the host,leading to systemic inflammation. Alternatively, pas-sively infused antibodies may recognize and bind ABOantigens on endothelial cells, leading to an increase intreatment-related toxicity and microvasculaturedamage.5,62 Some transplant centers provide ABO-compatible, plasma-reduced products for their ABO-mismatched, allogeneic transplant patients.4,5

Little is known regarding the effect of ABO-incompatible platelets in the setting of solid organtransplantation, particularly ABO-incompatible kidneyand heart transplants. Current regimens require severalrounds of plasmapheresis or immunoadsorptioncombined with immunosuppression to decrease iso-agglutinin titers.63,64 To avoid passive transfusion ofisoagglutinins harmful to the graft, platelets should be

plasma compatible with the donor in the immediateperioperative period. However, transfusion of ABOmajor-incompatible platelets could also presentproblems, particularly postoperatively. As discussed ina previous section, ABO-incompatible platelets canstimulate isoagglutinin titers in some recipients.44,45 Inone small study,ABO-incompatible platelet transfusionsfrom the intended kidney donor were used to test theimmune responsiveness of the recipient.45 Of sevenpatients, one developed HLA antibodies and threedemonstrated a rise in isohemagglutinin titers. Onlypatients with little or no response to platelettransfusion underwent transplantation with goodoutcomes 4 to 7 years after transplant.

Acute Hemolytic Transfusion ReactionsCurrently, the greatest discussion around ABO and

platelet transfusion is the risk of acute hemolytictransfusion reactions (HTR) with ABO minor-incompatible or out-of-group platelets (Table 1).65–88

Although most queried transfusion services provideABO-identical platelets or plasma-compatible plateletswhen available,89,90 it is estimated that 10 to 40 percentof transfusions are plasma incompatible with therecipient.73,76 The most common reasons for trans-fusing out-of-group platelets are limited inventory ofABO type-specific platelets and HLA-matched plateletsand to minimize product wastage.90 To date, only ahandful of severe HTRs with platelet transfusion havebeen reported in the literature and Internet chatrooms.

Despite the transfusion of more than 2 millionplatelet concentrates per year (including potentially200,000 to 400,000 out-of-group transfusions), the truerisk and incidence of hemolysis with plasma-incompatible platelet transfusions is still a matter ofconjecture. Facility-specific rates for platelet-associatedHTR range from less than 1 in 1000 to less than 1 in25,000 for all platelet transfusions and less than 1 in100 to less than 1 in 9000 for minor-incompatibleplatelet trans-fusions.73,76,91,92 This wide range mayreflect heterogeneity in the type of platelet productstransfused (apheresis vs. random), index of clinicalsuspicion, and severity of the reaction. On the basis ofthe yearly number of transfusions in the United States,it is clear that severe HTRs after out-of-group platelettransfusions are relatively rare events. However, mildhemolysis may be more common than appreciated.Oza et al.92 found mild to moderate evidence ofhemolysis with elution of ABO antibodies in 1 to 2


ABO and platelets

Table 1. Acute hemolytic transfusion reactions with platelets

Donor ABO type Platelet donor and product Outcome†

Reference Age* Sex Diagnosis Patient Donor(s) Saline AHG Product Volume ↓Hb (%)

Zoes65 44 F AML-M4 AB O Anti-A:256‡ NR Random-pool 500 mL NRAnti-B:64

McLeod66 45 M AML-M6 A O 1280 10,240 Random-pool 50 6 (43%)O 640 10,240 Random-pool 50

Conway67 15 F Transplant A O 8192 NR Apheresis-HLA 200 NR

Pierce68 2.5 F ALL A O 512 32,000 Apheresis 200 5.8 (50%)58 F Angioplasty B O 512 16,384 Random 50 6.1 (43%)

Ferguson69 66 M AML A O 256 >4000 Random 50 2.6

Reis70 56 M Aplastic anemia B O NR 4096 Apheresis NR 6.1 (46%)

Murphy71 30 F AML A O 512 2048 Apheresis-HLA 225§ 4.0 (40%)256 1024 Apheresis-HLA 448§ 5.4 (47%)

Chow72 18 F AML-M4 AB O 1024‡ 520‡ Apheresis, pooled 1200¶ 5.5 (50%)

Mair73 28 M Neuroblastoma A O,A,AB 128 NR Apheresis 225 2.6 (31%)

McManigal74 72 F Cardiac surgery AB O,B,A NR NR Apheresis 3380¶

Larsson76 44 F AML A O Anti-A:16384 NR Apheresis 371 2.3 (30%)

Duguid75 0.1 M Cardiac surgery A O NR NR Random-pool 100 4.5 (33%)0.02 F Cardiac surgery A O NR NR Random-pool 100 NR

Valbonesi77 51 F Breast cancer A O >8000 NR Double apheresis 37 3.5 (40%)16 F Aplastic anemia A (dry platelet) 37 3.2 (37%)

Sauer-Heilborn78 35 M Transplant B O 4096 NR Apheresis 526 5.3 (50%)

Anonymous79 36 F Hodgkin’s A O NR 2048 Apheresis 214 2.5 (22%)45 M AML A O NR 4096 Apheresis 241 1.6 (15%)

Gresens80 29 M Trauma A O 1024 1024 Apheresis 260 NR

Ozturk82 21 M RAEB A O NR NR Apheresis 600 5.7 (36%)

Yeast81 0** NR NAIT B O NR NR Apheresis 15 10 (83%)

Anonymous83 NR NR NR AB O NR NR Apheresis NR 5.3 (53%)

Josephson84 Adult NR Leukemia A O 256 8192 Apheresis 50 3Adult NR Leukemia A O NR 1024 Apheresis 50 3

Angiolillo85 0.7 M Histiocytosis A O 128 NR Apheresis 107 3.5 (47%)

Sapatnekar86 2 F Medulloblastoma A O 2048 16384 Apheresis 145 3.9 (32%)

Sadani87 65 F AML-M0 A O 160 1280 Apheresis NR 3.8 (49%)

Mean 32.5 2403 7162 350 5.8

Range 0–72 128–16,384 1024–32,000 82%-Apheresis 15–3380 1.6–10

AHG = antihuman globulin;ALL = acute lymphocytic leukemia;AML = acute myelocytic leukemia; NAIT = neonatal alloimmune thrombocytopenia; NR = not reported; RAEB =refractory anemia with excess blasts.

* Age in years.

† Fall in Hb by g/dL and percent decrease (%) from pretransfusion levels.

‡ Isoagglutinin titers in the recipient after transfusion of ABO minor-incompatible platelets.

§ Same HLA-matched donor, transfusions 1 month apart.

¶ Total volume of incompatible plasma infused.

Active bleeding.

** Intrauterine transfusion.


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percent of all ABO-mismatched apheresis platelets,which accounted for 6.7 percent of all transfusionreactions associated with platelets. An increased rate ofadverse reactions with ABO minor-incompatibletransfusions was also noted by Wagner and Adamo.93

Out-of-group platelet transfusions, however, have aminimal impact on RBC utilization. Two studies havefound no difference in the level of anemia or RBCutilization in oncology patients receiving plasma-incompatible transfusions.73,94

A summary of severe HTRs associated with plasma-incompatible platelets is shown in Table 1. In nearly allcases, hemolysis followed the transfusion of group Oplatelets to a non-group O patient. Clinically, theunfortunate recipient displayed classic symptoms of anacute HTR immediately or shortly after transfusion,leading to rapid recognition and diagnosis. In a fewinstances, the diagnosis was only suspected hours ordays later after the onset of hemoglobinemia,hemoglobinuria, or unexplained anemia.66,68,75,77,85,87

Although most patients recovered with hydration andsupportive care, a handful of deaths are known,including at least five deaths in the United States.68,78 Invery rare cases, plasma exchange, RBC exchange withgroup O RBCs, or both were performed to stem thehemolysis.77

Laboratory studies in these patients were allconsistent with an acute HTR. Patients’ RBCs typicallybecame strongly positive in the DAT (IgG, C3) withelution of ABO antibodies from the RBCs. Laboratoryevidence of intravascular hemolysis was alwayspresent, including an abrupt drop in hemoglobin,hemoglobinemia, hemoglobinuria, decreased hapto-globin, spherocytes, hyperbilirubinemia, and elevatedlactate dehydrogenase. Disseminated intravascularcoagulation has been reported in two cases with fataloutcomes.68,78 The degree of hemolysis can beimpressive, with a drop in hemoglobin ranging from 2to 10 g/dL. In many cases, the patient will show anappropriate increment in platelet count aftertransfusion.70,74,81 An investigation of the implicatedplatelet donors revealed unusually high isoagglutinintiters in most cases. Several of the group O donorsin published reports were women, and includeda directed donation from a group O mother to her 2-year-old daughter.68,77 Women with a history of a priorABO-incompatible pregnancy are at increased risk fordeveloping high-titer isoagglutinins.95

Several factors may raise the risk for an HTR afteran out-of-group platelet transfusion. Apheresis

platelets,which contain 200 to 400 mL of plasma froma single donor, carry a higher risk of causing hemolysisbecause of a high-titer donor than do pooled platelets.This is substantiated by a review of the literature, inwhich 23 of 28 (82%) platelet-associated HTRs wereassociated with apheresis platelets. This risk issubstantially lessened with pooled platelets because ofthe smaller volume of plasma per donor (50 mL),whichis subsequently diluted 4- to 6-fold in the final product.The use of pooled platelets does not eliminate the riskof an HTR: four HTRs caused by plasma-incompatiblepooled random platelets have been reported, in twoneonates75 and two adults.65,66 In the latter, two high-titer donors were pooled and infused.66 The totalvolume of ABO-incompatible plasma must also beconsidered, including residual plasma present intransfused group O RBCs.74,87,96 McManigal et al.74

reported severe hemolysis in a patient who receivedmultiple out-of-group platelet transfusions, resulting inthe transfusion of 3380 mL of incompatible plasma(106% of patient’s blood volume) during a period of 4days.74

Patient factors can also contribute to the risk of anHTR after an ABO-incompatible transfusion. As shownin Table 1, there is an increased risk of hemolysis whenhigh-titer isoagglutinins are transfused to group A andAB recipients (23 of 28)—a finding not surprising toblood bank staff. Very young children may also be atincreased risk because of their relatively small bloodvolume.75,81,85,87 It has been surmised that youngchildren and nonsecretors may also have an increasedsusceptibility because ABO substances capable ofadsorbing and neutralizing ABO antibodies aredecreased in or absent from their blood. Amongpublished cases, only two investigated the Lewis typeof the recipient.65,78 One recipient was a nonsecretor[Le(a+b–)])65; however, the second patient wasLe(a–b+),78 suggesting that soluble ABO antigens mayprovide only limited protection. Finally, other ill-defined patient factors could also play a role. Look-back investigations of two high-titer group O apheresisplatelet donors failed to identify any additional HTRs in70 prior transfusion recipients.76,78

Approaches to Minimize Hazards of Plasma-Incompatible Platelet Transfusions

The increased use of apheresis platelets, in whichall the plasma is from a single donor, has heightenedawareness and concern regarding out-of-groupplatelet transfusions. Several strategies have been


ABO and platelets

implemented. Most transfusion services have policiesdictating the transfusion of ABO-identical or plasma-compatible platelets when available.89,90 Moreproactive strategies are discussed in the followingsections.

Identification of “dangerous donors”Although only 2 percent of U.S. centers routinely

screen group O donors,90 the practice is relativelycommon in Europe.89 With rare exceptions, donorsidentified as high titer are still permitted to donate;however, their products are segregated and labeled ashigh titer with a warning to only transfuse to group Opatients. The cost effectiveness of prospectivescreening is still unproven, given the relative rarity ofHTR with out-of-group platelets. Sadani et al.87 recentlyreported a severe HTR in a group A patient aftertransfusion of group O apheresis platelets that initiallytested as low titer by automated testing. Subsequenttesting revealed a high-titer anti-A IgG (Table 1).87 Incontrast, the New York Blood Center, which routinelyscreened group O apheresis donors until 1998,has hadno reports of acute HTRs in the 5 years, and more than25,000 platelet transfusions, since screening wasdiscontinued.91

A persistent problem that plagues widespreaddonor screening is the absence of a recognizedstandard method for testing. Several manual andautomated methods are currently used for donorscreening (tube, gel, and solid phase) with a variety ofend point measurements: saline titers, AHG titers, orhemolysis.84,87,89,97 Even with a single method, there is awide range in results (Fig. 2A). Improvements andincreased use of automated blood group testing, suchas gel and solid phase methods,may eliminate much ofthe perceived variation in donor isoagglutinin testingwith minimal add-on cost to the final product. This hasbeen demonstrated in England,where high-throughputtesting is performed using a single plasma or serumdilution (1 in 100) on an automated blood analyzer.89 Acost analysis by Emory University, which implementedtesting of all group O apheresis donors by gel method,estimated an additional cost of $1.20 per apheresisplatelet.84

A second problem with donor screening isdetermining a critical titer for “safe” versus high-titer,“dangerous” donors. As shown in Figure 2A, thepercentage of donors classified as high titer can rangefrom 3 to 50 percent, depending on the populationtested and the method and critical titer used.76,84,89,97 A

method and critical titer that classifies 40 to 50 percentof group O donors as high titer could significantlyhamper platelet availability to non-group O patients,particularly during times of severe shortages. A surveyof current practice shows a cutoff titer of 32 to 200 forsaline testing and 250 to 512 for AHG (Table 2). For

Fig. 2. ABO titers in platelet donors. A: Distribution and variability in“critical” titers (anti-A) among group O donors. Critical titers bysaline (black circles) and with AHG (squares) for several studiesare shown. B and C: Distribution of anti-A titers in a randomsampling of group O platelet donors (hatched columns) andgroup O donors implicated in acute HTRs (black,Table 1). Datafor normal donors taken from Josephson et al.84 by gel. Note that38 percent of normal donors had titers less than the initial titerof 32.84 Shown are saline titers (B) and after AHG,C.


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this review, we compared the distribution ofisoagglutinin titers from normal group O apheresisdonors84 and group O donors implicated in HTRs(Table 1). As shown in Figures 2B and 2C,many of thecutoffs used (32–64) are quite conservative.

Plasma reduction and platelet additive solutionsA popular method to reduce the risk of ABO HTR

caused by plasma-incompatible transfusions combinesplasma reduction with resuspension in either additivesolutions or group AB donor plasma. In the mostcommon method, platelet concentrates are preparedand stored in plasma (30–40%) diluted with a plateletadditive storage solution. Although not licensed in theUnited States, a selection of storage solutions arecommercially available or undergoing trials inEurope.98,99 The use of additive solutions has severalattractive advantages for increasing transfusion safetyincluding a decreased risk of HTRs as well as TRALIand allergic and febrile reactions caused by residualproteins, cytokines, and antibodies present in donorplasma.93,98–100 In Germany,Wagner and Alamo93 noteda marked decrease in transfusion reactions withplasma-reduced platelet concentrates (0%) comparedwith routine pooled platelets (27%). Plasma-reducedplatelets reportedly have equivalent posttransfusionrecovery at 1 hour although there is reduced 24-hoursurvival.89,98

Plasma reduction and use of platelet additivesolutions may reduce, but will not completelyeliminate, the risk of an acute HTR caused by a high-titer donor. An enlightening case was recentlypublished by Valbonesi et al.,77 who described severe

HTRs in two recipients of a split, group O apheresisunit (Table 1). Apheresis platelets were collected “dry”with only minimal plasma carryover (74 mL), followedby resuspension in 400 mL of a platelet additive (T-Sol).An investigation identified extremely high isoagglutinintiters (> 8000) in the female donor. The authors notedthat this was the first severe HTR encountered in morethan 16,000 “dry” platelet collections in the last 10years.

Limiting plasma infusionSome transfusion services monitor and limit the

total volume of incompatible plasma that a recipientcan receive.79,83,89,90,101 Policies vary betweeninstitutions, ranging from 300 to 500 mL of plasma perday to 1000 mL of plasma per week (Table 3). In theUnited States, approximately 10 percent of polledinstitutions (255) limited the amount of incompatibleplasma transfused to adult patients.

In neonates and young children with small bloodvolumes, there is a greater concern regarding the riskof out-of-group platelets.102 As a consequence, mostU.S. hospitals serving neonatal populations provideonly ABO-identical or plasma-compatible platelets(60%). Very few institutions (1%, 29 institutions) havepolicies limiting incompatible plasma infusion toneonates and children.

Identification of A2 donors as “universal plateletdonors”

Because A2 donors have little or no A antigen onplatelet membranes, they are compatible with group Aand O donors. Donor screening to identify group A2

donors could increase the inventory of group Ocompatible from 44 to 52 percent, increasing thenumber of apheresis platelets available for plateletcrossmatching,10 which requires ABO compatibilitybetween donor and patient.36 It could also benefitselection and survival of HLA-matched platelets.39,40 Toour knowledge, only Norway has policies foridentifying A2 platelet donors as “universal donors.”

89

A2 platelets are also an ideal component for patientsundergoing ABO-mismatched allogeneic bone marrowtransplantation. In ABO major-incompatible transplants,present guidelines recommend that transfused plateletsbe plasma compatible with the marrow or stem celldonor to minimize delays in erythroid engraftment.46 Asa consequence, platelets will often be ABOincompatible with the patient’s isohemagglutinins. Ingroup A (donor) to group O (patient) mismatched

Table 2. Strategies for screening apheresis platelet donors for high-titerisoagglutinins

Screen Critical %Country donors Method titer Donors

United States84,90,91 No (2%)90 Tube, gel 1:50–1:200 3–28%

England89 Yes Automated 1:100 3–10%Tube 1:128

Scotland87 Yes 1:50

Italy89 Yes Gel 1:64IAT 1:256

Germany89 Yes Tube, saline 1:64 5%

Czech Republic89 Yes Tube, saline 1:64

Norway89 Yes IAT 1:250

Sweden89 Yes Tube, saline 1:100IAT 1:400

Switzerland89 Yes Hemolysis 1:16

Finland89 Yes Tube, saline 1:32 5.7%

Japan89 Yes IAT 1:512


ABO and platelets

transplants, this incompatibility can be overcome bytransfusion of A2 platelets, which are ABO compatiblewith the patient’s isoagglutinins and plasma compatiblewith the donor graft.10 A2 platelets might also be aproduct of choice in group O (donor) to group A(patient) minor-mismatch transplants. Likewise, A2

platelets are an ideal product for group O patientsreceiving group A-incompatible solid organ transplants.In these patients, transfusion of incompatible plasmaand platelets should be avoided because of the risk ofacute humoral rejection and immune stimulation,respectively.45,60 It is my fervent wish that A subtypingof apheresis platelet donors become a standard ofpractice by blood collection centers.

SummaryPlatelets express ABO antigens on a large number

of platelet glycoproteins and glycolipids. The amountof ABO antigen expressed on individual donor plateletsis heterogeneous and determined by genetic andepigenetic factors. Routine transfusion of ABO-incompatible platelets can be associated withcumulative adverse effects including decreasedposttransfusion recovery, increased platelet utilization,incompatible platelet crossmatches, HLA alloim-munization, and ABH-specific refractoriness. Out-of-group platelet transfusion can also be associatedwith adverse effects including decreasing plateletincrements and, rarely, severe HTRs. Out-of-groupplatelet transfusion may have more severeconsequences in ABO-mismatched bone marrow

transplant patients, who are at increased risk forsignificant morbidity and mortality because of ABOincompatibility.

ABO-identical platelets should be providedwhenever possible, particularly for patients requiringlong-term transfusion support. In patients undergoingABO-mismatched transplants, provision of A2 plateletsmay be preferable. In patients requiring HLA-antigennegative and HLA-matched platelets,HLA matching haspriority; however, the ABO compatibility of HLAplatelets should be recorded and considered whenassessing the success or failure of HLA platelets. ABOcompatibility should always be considered whenevaluating and monitoring patients with clinicalrefractoriness, particularly group O patients who havehigher mean isohemagglutinin titers.

In neonates and young children, only ABO-identicalor plasma-compatible platelets should be transfused.102

In surgical and other patients with short-termtransfusion needs, ABO compatibility is less of aconcern. ABO-identical or plasma-compatible plateletsshould be transfused if available; however, an activelybleeding patient should never be denied a platelettransfusion because of ABO compatibility. The risks ofbleeding and the need to achieve hemostasis outweighthe potential adverse consequences of transfusing anABO-incompatible platelet. One potential exception isABO-incompatible organ transplantation, in whichplasma containing donor-reactive isoagglutinins shouldbe avoided. In the postoperative period, donor-compatible platelets may be considered to minimizeimmune stimulation of recipient isoagglutinins.

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Table 3. Transfusion strategies for minimizing HTR caused byincompatible-plasma infusion

ABO-identical platelets

ABO plasma-compatible platelets

Plasma-reduced

Resuspension in storage additive solution, group AB plasma

Pooled platelets

Screened “low-titer” apheresis platelets

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Children79

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Neonates: 0 mL, plasma-compatible only


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44. Carr R, Hutton JL, Jenkins JA, Lucas GF, AmphlettNW. Transfusion of ABO-mismatched plateletsleads to early platelet refractoriness.Br J Haematol1990;75:408–13.

45. Ishibashi M,Oshida M,KurataY,et al. Immunologiclow-responder in ABO-incompatible renal trans-plant recipients explored by donor-specificplatelet transfusion. Transplant Proc 1998;30:2298–9.

46. Heal JM, Masel D, Blumberg N. Interaction ofplatelet Fc and complement receptors withcirculating immune complexes involving the ABOsystem.Vox Sang 1996;71:205–11.

47. Heal JM, Masel D, Rowe JM, Blumberg N.Circulating immune complexes involving the ABOsystem after platelet transfusion. Br J Haematol1993;566–72.

48. Duquesnoy RJ, Anderson AJ, Tomasulo PA, AsterRH.ABO compatibility and platelet transfusions ofalloimmunized thrombocytopenic patients. Blood1979;54:595–9.

49. Ellinger PJ, Morgan LK, Malecek AC, Chaplin H.Effect of ABO mismatching on a radioim-munoassay for platelet compatibility. Successfuladsorption of ABO alloantibodies with synthetic Aand B substance.Transfusion 1989;29:134–8.

50. Blumberg N, Heal JM, Rapoport A, et al. Effect ofABO-identical platelets and leukodepletion onblood utilization and costs of autologous marrowtransplantation (abstract). Transfusion 1993;33:A15.

51. Heal JM, Rowe JM, Blumberg N. ABO and platelettransfusion revisited. Ann Hematol 1993;66:309–14.

52. Blumberg N, Heal JM, Kirkley SA, et al.Leukodepleted-ABO identical blood componentsin the treatment of hematologic malignancies: acost analysis.Am J Hematol 1995;48:108–15.

53. Heal JM, Kenmotsu N, Rowe JM, Blumberg N. Apossible survival advantage in adults with acute


L. COOLING

leukemia receiving ABO-identical platelettransfusions.Am J Hematol 1994;45:189–90.

54. Heal JM, Blumberg N. Optimizing platelettransfusion therapy. Blood Rev 2004;18:149–65.

55. Blumberg N, Heal JM, Hicks GL, Risher WH.Association of ABO-mismatched platelet trans-fusions with morbidity and mortality in cardiacsurgery.Transfusion 2001;41:790–3.

56. Blumberg N, Heal JM. ABO-mismatched platelettransfusions and clinical outcomes after cardiacsurgery (letter).Transfusion 2002;42:1527–9.

57. Lin Y, Callum JL, Coovadia AS, Murphy PM.Transfusion of ABO-nonidentical platelets is notassociated with adverse clinical outcomes incardiovascular surgery patients. Transfusion2002;42:166–72.

58. Badros A,Tricot G,Toor A,et al.ABO mismatch mayeffect engraftment in multiple myeloma patientsreceiving nonmyeloablative conditioning. Trans-fusion 2002;42:205–9.

59. Benjamin RJ,McGurk S, Ralston MS,ChurchillWH,Antin JH. ABO incompatibility as an adverse riskfactor for survival after allogeneic bone marrowtransplantation.Transfusion 1999;39:179–87.

60. Stussi G, Halter J, Schanz U, Seebach JD.ABO-histoblood group incompatibility in hematopoieticstem cell and solid organ transplantation.TransfusApheresis Sci 2006;35:59–69.

61. Brecher ME, ed. Technical manual, 15th ed.Bethesda, MD: American Association of BloodBanks, 2005.

62. Gordon B, Tarantolo S, Ruby E, et al. Increasedplatelet transfusion requirement is associatedwith multiple organ dysfunctions in patientsundergoing hematopoietic stem cell transplan-tation. Bone Marrow Transplant 1998;22:999–1003.

63. West L, Pollock-Barziv SM, Diphanc AI, et al. ABO-incompatible heart transplantation in infants. NEngl J Med 2001;344:793–800.

64. Stegall MD, Dean PG, Gloor JM.ABO-incompatiblekidney transplantation. Transplantation 2004;78:635–40.

65. Zoes C, Dube VE, Miller HJ,Vye MV.Anti-A1 in theplasma of platelet concentrates causing ahemolytic reaction.Transfusion 1977;17:29–32.

66. McLeod BC, Sassetti RJ,Weens JH,Vaithianathan T.Haemolytic transfusion reaction due to ABOincompatible plasma in a platelet concentrate.Scand J Haematol 1982;28:193–6.

67. Conway LT, Scott EP. Acute hemolytic transfusionreaction due to ABO incompatible plasma in aplatelet apheresis concentrate (letter).Transfusion1984;24:413–4.

68. Pierce RN, Reich LM, Mayer K. Hemolysisfollowing platelet transfusions from ABO-incompatible donors.Transfusion 1985;25:60–2.

69. Ferguson DJ.Acute intravascular hemolysis after aplatelet transfusion. Can Med Assoc J 1988;138:523–4.

70. Reis MD, Coovadia AS. Transfusion of ABO-incompatible platelets causes severe haemolyticreaction. Clin Lab Haematol 1989;11:237–40.

71. Murphy MF, Hook S, Waters AH, et al. Acutehaemolysis after ABO-incompatible platelettransfusions (letter). Lancet 1990;335:974–5.

72. Chow M-P,Yung C-H,Hu H-Y,Tzeng C-H.Hemolysisafter ABO-incompatible platelet transfusion. ChinMed J (Taipei) 1991;48:131–4.

73. Mair B, Benson K. Evaluation of changes inhemoglobin levels associated with ABO-incompatible plasma in apheresis platelets.Transfusion 1998;38:51–5.

74. McManigal S, Sims KL. Intravascular hemolysissecondary to ABO incompatible platelet products.Am J Clin Pathol 1999;111:202–6.

75. Duguid JKM,Minards J, Bolton-Maggs PHB. Lessonof the week: incompatible plasma transfusionsand haemolysis in children. Br Med J 1999;318:176–7.

76. Larsson LG,Welsh VJ, Ladd DJ. Acute intravascularhemolysis secondary to out-of-group platelettransfusion.Transfusion 2000;40:902–6.

77. Valbonesi M, De Luigi MC, Lercari G, et al. Acuteintravascular hemolysis in two patients transfusedwith dry-platelet units obtained from the sameABO incompatible donor. Int J Artif Organs2000;23:642–6.

78. Sauer-Heilborn A, Jahagirdar B, Burns L, Scofield T,Nollet KE. Passive antibody, aggressive hemolysis:an ABO-incompatible platelet transfusion (ab-stract).Transfusion 2002;42(Suppl):SP304.

79. Reducing supernatant plasma of pooled plateletsbefore administration to ABO-incompatible recip-ients. CBBS, e-network forum. Available at: http://www.cbbsweb.org/enf/2002/pltpoolplasma.html.

80. Gresens C, Gloster E, Wang L, Dimaio T. Acutehemolysis in a group A trauma patient whoreceived a group O plateletpheresis unit (ab-stract).Transfusion 2003;43(Suppl):SP234.


ABO and platelets

81. Yeast JD, Plapp F. Fetal anemia as a response toprophylactic platelet transfusion in the manage-ment of alloimmune thrombocytopenia. Am JObstet Gynecol 2003;189:874–6.

82. Ozturk A, Turken O, Sayan O, Atasoyu EM. Acuteintravascular hemolysis due to ABO-incompatibleplatelet transfusion. Acta Haematol 2003;110:211–2.

83. What is the frequency of hemolytic reactionsassociated with transfusion of plateletpheresisunits containing ABO incompatible plasma. CBBS,e-network forum. Available at: http://www.cbbsweb.org/enf/2003/plt_aborxnfreq.html.

84. Josephson CD, Mullis NC, Van Demark C, HillyerCD. Significant numbers of apheresis-derivedgroup O platelet units have “high-titer” anti-A/A,B:implications for transfusion policy. Transfusion2004;44:805–8.

85. Angiolillo A, Luban NLC. Hemolysis following anout-of-group platelet transfusion in an 8-month-old with Langerhans cell histiocytosis. J PediatrHematol Oncol 2004;26:267–9.

86. Sapatnekar S, Sharma G, Downes KA,Wiersma S,McGrath C, Yomtovian R. Acute hemolytictransfusion reaction in a pediatric patient follow-ing transfusion of apheresis platelets. J ClinApheresis 2005;20:225–9.

87. Sadani DT,Urbaniak SJ, Bruce M,Tighes JE. RepeatABO-incompatible platelet transfusions leading tohaemolytic transfusion reaction. Transf Med2006;16:375–9.

88. Lozano M, Cid J. The clinical implications ofplatelet transfusions associated with ABO orRh(D) incompatibility.Transfus Med Rev 2003;17:57–68.

89. Pietersz RNI, Engelfriet CP. Transfusion of apher-esis platelets and ABO groups.Vox Sang 2005;88:207–21.

90. College of American Pathology. TransfusionMedicine Survey (J-A). 2005.

91. Schwartz J, Delpalma H, Kapoor K, Hamilton T,Grima K. Anti-A titers in group O single donorplatelets: to titer or not to titer (abstract).Transfusion 2003;43(Suppl):SP247.

92. Oza KK.ABO mismatched platelet transfusion andacute intravascular hemolysis (abstract). Trans-fusion 2002;42(Suppl):SP308.

93. Wagner F, Adamo W. Reduced rate of adversereactions to plasma reduced pooled platelet units:possible role of minor incompatible transfusion(abstract).Vox Sang 2006;91(Suppl 3):P508.

94. Shanwell A, Ringden O, Wiechel B, Rumin S,Akerblom O. A study of the effect of ABOincompatible plasma in platelet concentratestransfused to bone marrow transplant recipients.Vox Sang 1991;60:23–7.

95. Mollison PL, Engelfriet CP, Contreras M, ed. Bloodtransfusion in clinical medicine. 10th ed. Oxford:Blackwell Science, 1997.

96. Boothe G, Brecher ME, Root M, Robinson J, HaleyR. Acute hemolysis due to passively transfusedhigh-titer anti-B causing spontaneous in vitroagglutination. Immunohematology 1995;11:43–5.

97. The practice of transfusing out-of-type platelets.CBBS, e-network forum. Available at: http://www.cbbsweb.org/enf/2002/plttx_aboincompat.html.

98. deWildt-Eggen J,Gulliksson H. In vivo and in vitrocomparison of platelets stored in either syntheticmedia or plasma.Vox Sang 2003;84:256–64.

99. Ringwald J, Zimmerman R, Eckstein R. The newgeneration of platelet additive solution for storageat 22°C: development and current experience.Transfus Med Rev 2006;20:158–64.

100. deWildt-Eggen J,Nauta S,Schrijver JG,van MarwijkKooy M, Bins M, van Prooijen HC. Reactions andplatelet increments after transfusion of plateletconcentrates in plasma or an additive solution: aprospective, randomized study. Transfusion2000;40:398–403.

101. References on patient outcomes followingtransfusion of ABO incompatible platelets. CBBS,e-network forum. Available at: http//www.cbbsweb.org/enf/outcome_aboinc_plt.html.

102. Chambers LA.Transfusion of plasma and plateletsto neonates. In: Herman JH, Manno CS, eds.Pediatric transfusion therapy, 2002. Bethesda,MD:AABB Press, 2002:93–107.

Laura Cooling, MD, MS, Assistant Professor,Pathology, Associate Medical Director, TransfusionMedicine, University of Michigan Hospitals, 2F225UH, Box 0054, 1500 East Medical Center Drive, AnnArbor, MI 48109-0054.


Dr. Scott Murphy always enjoyed a medical mysteryand his passion for the syndrome of posttransfusionpurpura (PTP) was one of his favorite topics. This“perfect storm”of immunologic events is characterizedby profound thrombocytopenia approximately 7 daysafter transfusion and includes the development of aplatelet-specific alloantibody which destroys anyantigen-positive transfused platelets as well as thepatient’s own platelets, which lack the correspondingantigen. Dr. Murphy’s interest in this syndrome beganin the early years of his research career. He alsoprovided our laboratory with the first antiserum,whichwe used to develop testing procedures and toinvestigate numerous cases of PTP. In the next fewpages of this edition dedicated in his honor, I wouldlike to provide a brief trip down memory lane toremember his valued contribution to the PlateletSerology Laboratory at the Penn-Jersey Region of theAmerican Red Cross (ARC) in Philadelphia, to outlinesome of the many advances that have taken place inthe investigation of platelet antibodies, and tosummarize the results of our laboratory investigation ofPTP at Penn-Jersey.

I first had the pleasure of meeting Scott Murphy inthe late 1970s while working as a research technologistat the Penn-Jersey Region of the ARC. At that time,wewere in the process of developing a procedure to typedonor platelets for the PlA1 antigen. Dr.Miriam Dahlke,our medical director, informed me that Dr.Murphy hadoffered to share a supply of anti-PlA1, which was theresult of a plasma exchange performed on a patientdiagnosed with PTP. Not wanting to wait for him tochange his mind, I headed to his laboratory at the otherend of town on that very hot, late spring day, armedwith a box and a small supply of ice.

I was greeted by a bearded 1970s version of ourown Scott Murphy, complete with heavy cotton lab

coat emblazoned with his name and“Cardeza” in brightred embroidery. He appeared supremely happy in hiscrowded research lab as he presented me with a bagcontaining nearly a full liter of plasma. I remember hisresponse after I thanked him and asked if ourlaboratory could do anything to return the favor. Hesaid, “just type a lot of platelets for our patients.” Iwould later learn that this was his way, always agracious gentleman who held his patients and others’as his first priority.

As I headed back with my precious cargo, I noticeda small split in the bag and was faced with the dilemmaof making it back to the ARC building before it thawed(and leaked all over Center City Philadelphia). The just-in-time arrival of an available taxi cab allowed me tobeat the clock and the weather. I have to admit thatthis was, unquestionably, the only time in my careerthat, while working late into the evening, the usuallyboring task of preparing a thousand tiny 1-mL vials waspure pleasure.

In 1975, while on the staff of the HematologyResearch Laboratory at the Presbyterian-University ofPennsylvania Medical Center in Philadelphia, Dr.Murphy coauthored a seminal publication callingattention to the variety of responsible antibodies andpatient profiles in the relatively newly described andclinically mysterious syndrome of PTP.1

Before the publication of this paper in 1975, all ofthe 14 described cases of PTP were attributed to thepresence of anti-PlA1 produced by PlA1-negative femalepatients. Two of the three patients described in thispaper (one PlA1-negative man who produced anti-PlA1

and a PlA1-positive woman with a demonstrable anti-platelet antibody of unknown specificity) served toexpand the definition of PTP to include both male andfemale patients with antibody specificities not limitedto anti-PlA1.

Scott Murphy’s contribution inthe early years of posttransfusionpurpura: a remembranceM.KEASHEN-SCHNELL


PTP is a relatively rare, but well-defined, syndromefirst described by Shulman et al. in 1961.2 The classicpatient profile describes an older, multiparous womanwith a precipitous, antibody-mediated drop in plateletcount occurring 7 to 10 days after transfusion. Unlikethe maternal antibodies responsible for neonatalalloimmune thrombocytopenia, the antibodiesproduced by patients diagnosed with PTP cause severethrombocytopenia in the antibody producer. PTP isself-limiting, but carries a significant risk of fatalhemorrhage. Similarities have been noted betweenPTP and the hemolysis of autologous RBCs duringdelayed hemolytic transfusion reactions caused bysome RBC antibodies.3 To date, there is no definitiveexplanation for the destruction of autologous cells ineither situation.

In the years after Dr. Murphy’s 1975 publicationand since the first description of a platelet-specificantigen by van Loghem in 1959,4 much has beenlearned about the nature of platelet antigens. Thenomenclature for platelet-specific antigens wasstandardized by a Working Party on Platelet Serology5

(Table 1) and adopted by the International Society ofBlood Transfusion. In this more orderly, numericsystem, the platelet antigens are designated as HPA(human platelet antigens) and are numbered (withArabic numbers) according to discovery date, followedby a lower case“a”or“b,”which denotes the high (a) orlow (b) frequency member of the antigen pair. As anexample of the need for simplification, the first antigenreported was referred to as Zwa in Europe and PlA1 inthe United States. Under the new nomenclature, it isnow called HPA-1a. Its lower frequency allele (PlA2 orZwb) is now HPA-1b.

We now know that six platelet-specific antigensystems (HPA-1 through 5 and HPA-15) are biallelicwith one high frequency and one low frequencyantigen or allele.6 For the remaining 10 antigens (HPA-6 through 14 and HPA-16), alloantibodies against thelow frequency but not the high frequency antigen havebeen observed. The molecular basis and location onthe platelet membrane has been resolved for antigenslisted in Table 1. With the exception of HPA-14, thedifferences in the alleles are the result of a single aminoacid substitution at a specific location on the geneencoding for the membrane glycoprotein. Theisoantigen,Naka,7 is now known to be located on GPIV(CD36).8,9

These advances, as well as the availability ofmonoclonal antibodies, which recognize specific

glycoprotein locations on the platelet surface, have ledto the development of serologic testing techniqueswith increased sensitivity10–14 as well as to thedevelopment of molecular techniques15 for plateletantigen typing.

Other specificities have since been identified aspathogenic antibodies in PTP using a variety ofinvestigative techniques. These include anti-PlA2 (HPA-1b),16 anti-Baka (HPA-3a),17 anti-Bakb (HPA-3b),18 anti-Pena (HPA-4a),19 anti-Bra (HPA-5b),20 and anti-Brb (HPA-5a).21,22

In 1992,Dr.Murphy became the medical director ofthe Penn-Jersey Region, and we had the honor ofworking with him for the next 14 years. His fascinationwith the syndrome of PTP and his concern for thepatients involved never waned. He was never too busyto make time in his schedule to learn the details of eachnew case. He would then consult directly with thepatient’s clinician to ensure that our center couldprovide appropriate testing, consultation, andtransfusion support. Options for transfusion support,

Scott Murphy’s contribution to PTP

Table 1. Human platelet antigen nomenclature

Phenotypicfrequency

New Old Glycoprotein (approx. %nomenclature nomenclature location Caucasians)*

HPA-1a PlA1 (Zwa) IIIa (CD61) 98

HPA-1b PlA2 (Zwb) IIIa (CD61) 29

HPA-2a Kob Ib (CD42b) > 99

HPA-2b Koa (Siba) Ib (CD42b) 13

HPA-3a Baka (Leka) IIb (CD41) 81

HPA-3b Bakb IIb (CD41) 70

HPA-4a Yukb (Pena) IIIa (CD61) > 99

HPA-4b Yuka (Penb) IIIa (CD61) < 0.1

HPA-5a Brb (Zavb) Ia (CD49b) 99

HPA-5b Bra (Zava, Hca) Ia (CD49b) 20

HPA-6bw Caa,Tua IIIa (CD61) 0.7

HPA-7bw Moa IIIa (CD61) 0.2

HPA-8bw Sra IIIa (CD61) < 0.1

HPA-9bw Maxa IIb (CD41) 0.6

HPA-10bw Laa IIIa (CD61) < 1.6

HPA-11bw Groa IIIa (CD61) < 0.3

HPA-12bw Iya Ib (CD42c) 0.4

HPA-13bw Sita Ia (CD49b) 0.3

HPA-14bw Oea IIIa (CD61) < 0.2

HPA-15a Govb CD109 74

HPA-15b Gova CD109 81

HPA-16bw Duva IIIa (CD61) < 1

*After Norton A,Allen DL,Murphy MF. Review: platelet alloantigens and antibodiesand their clinical significance. Immunohematol 2004;20:89–99.

when needed,would vary from case to case and mightinclude antigen-negative single-donor platelets orwashed or deglycerolized RBCs.

Since 1983, our laboratory has investigated 115patient samples submitted to rule out PTP. Sampleswere tested for the presence of platelet-specific andHLA antibodies using a variety of methodologies. Of115, 33 (28.6%; 31 from women, 2 from men)contained platelet-specific antibody(ies) and thesepatients were diagnosed with PTP based on thisfinding along with clinical history. Of these, 23demonstrated anti-HPA-1a, 4 anti-HPA-1b, 3 anti-HPA-3a,1 anti-HPA-1b and anti-HPA-3a,1 anti-HPA-5b,and 1 anti-HPA-5a (Fig. 1). Molecular testing was used to confirmthe antigen-negative status in five cases. Nineteen wereconfirmed using serologic techniques on postrecoveryplatelet samples. Nine were not submitted or wereunavailable for confirmation. Of note is the fact that 23of the 33 (69.6%) sera also demonstrated HLA class Iantibodies in combination with platelet-specificantibodies, indicating the necessity to use glycoprotein-based as well as standard whole-cell methods as part ofthe investigative protocol to ensure detection ofcoexisting antibodies. Of the remaining 82 samples,the majority either were negative or demonstrated HLAantibody only. These results, along with otherpublished data,support Dr.Murphy’s early report of theheterogeneous nature of PTP.

Thank you,Scott Murphy, for your support,sustainedenthusiasm, and undiminished desire to help patients inneed. You are missed by everyone here at the ARC. As Ianticipate further advances in the challenging fields ofplatelet testing and transfusion, I will not forget thathappy, bearded researcher of many years ago, and I amdelighted to report that the plasma handed to me 29

years ago remains in use in our laboratory and continuesto help so many of “our patients.”

References1. Zeigler Z, Murphy S, Gardner F. Post-transfusionpurpura: a heterogeneous syndrome. Blood 1975;45:529–36.

2. Shulman NR, Aster RH, Leitner A, Hiller MC.Immunoreactions involving platelets. V. Post-transfusion purpura due to a complement-fixingantibody against a genetically controlled plateletantigen. A proposed mechanism for thrombo-cytopenia and its relevance in “autoimmunity.” JClin Invest 1961;40:1597–620.

3. Garratty G. Review: platelet immunology—similarities and differences with red cellimmunology. Immunohematol 1995;11:112–24.

4. van Loghem JJ,Dorfmeijer J,Van Hart M,SchruederF. Serological and genetical studies on a plateletantigen (Zw).Vox Sang 1959;4:161–9.

5. von dem Borne AE, DeCary F. Nomenclature ofplatelet-specific antigens (letter). Transfusion1990;30:477.

6. Metcalfe P, Watkins NA, Ouwehand WH, et al.Nomenclature of human platelet antigens. VoxSang 2003;85:240–5.

7. Ikeda H,Mitani T,Ohnuma M, et al.A new platelet-specific antigen, Naka, involved in the refrac-toriness of HLA-matched platelet transfusion.VoxSang 1989;57:213–7.

8. Yamamoto N, Ikeda H,Tandon NN, et al.A plateletmembrane glycoprotein (GP) deficiency inhealthy blood donors: Naka-platelets lackdetectable GPIV (CD36). Blood 1990;76:1698–703.

9. Greenwalt DE, Lipsky RH, Ockenhouse CF, et al.Membrane glycoprotein CD36: A review of itsroles in adherence, signal transduction, andtransfusion medicine. Blood 1992;80:1105-15.

10. von dem Borne AE,Verheught FW, Oosterhof F, etal.A simple fluorescence test for the detection ofplatelet antibodies. Br J Haematol 1978;39:195–207.

11. Rachel JM, Sinor LT,Tawfik OW, et al.A solid-phasered cell adherence test for platelet cross-matching.Med Lab Sci 1985;42:194–5.

12. Nordhagen R,Flaathen ST.Chloroquine removal ofHLA antigens for platelets for the plateletimmunofluorescence test. Vox Sang 1985;48:156–9.


M.A. KEASHEN-SCHNELL

Fig. 1. Penn-Jersey ARC Platelet Serology Laboratory results: antibodyspecificities of PTP investigations 1983–2007.


Scott Murphy’s contribution to PTP

13. Kiefel V, Santoso S,Weisheit M,Mueller-Eckhardt C.Monoclonal antibody-specific immobilization ofplatelet antigens (MAIPA): a new tool for theidentification of platelet-reactive antibodies.Blood1987;70:1722–6.

14. Furihata K, Nugent DJ, Bissonette A, et al. On theassociation of the platelet specific alloantigen,Pena, with glycoprotein IIIa. Evidence forheterogeneity of glycoprotein IIIa. J Clin Invest1987;80:1624–30.

15. McFarland JG, Aster RH, Bussel JB, et al. Prenataldiagnosis of neonatal alloimmune thrombo-cytopenia using allele-specific oligonucleotideprobes. Blood 1991;78:2276–82.

16. Taaning E, Killmann SA, Morling N, et al. Post-transfusion purpura (PTP) due to anti-Zwb (-PlA2):the significance of IgG3 antibodies in PTP. Br JHaematol 1986;64:217–25.

17. Keimowitz RM, Collins J, Davis K, Aster RH. Post-transfusion purpura associated with alloimmu-nization against the platelet-specific antigen, Baka.Am J Hematol 1986;21:79–88.

18. Kickler TS, Herman JH, Furihata K, et al.Identification of Bakb, a new platelet-specificantigen associated with posttransfusion purpura.Blood 1988;71:894–8.

19. Simon TL, Collins J, Kunicki TJ, et al. Post-transfusion purpura associated with alloantibodyspecific for the platelet antigen Pen(a). Am JHematol 1988; 29:38–40.

20. Christie DJ, Pulkrabek S, Putnam JL, et al. Post-transfusion purpura due to an alloantibodyreactive with glycoprotein Ia/IIa (anti-HPA-5b).Blood 1991;77:2785–9.

21. Anolik JH, Blumberg N, Snider J, Francis CW.Posttransfusion purpura secondary to an alloanti-body reactive with HPA-5a (Br(b)). Transfusion2001;41:633–6.

22. Ziman A, Klapper E, Pepkowitz S, et al. A secondcase of post-transfusion purpura caused by HPA-5a antibodies: successful treatment with intra-venous immunoglobulin. Vox Sang 2002;83:165–6.

Maryann Keashen-Schnell, BS, Supervisor, Platelet/Neutrophil Serology Laboratory, Manager, ProcessImprovement, American Red Cross Blood Services,Heritage Division, Penn-Jersey Region Department ofTechnical Services, 700 Spring Garden Street,Philadelphia, PA 19123.

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Passively transfused blood group antibodies cause clinicalproblems. High titers of anti-A and anti-B seem to be one reason forhemolytic transfusion reactions and for ABO HDN. In Japan, anti-Aand anti-B titers notably decreased in the 15 years between 1986and 2001. At present, titers of more than 100,as measured using thesaline method, are rare. Differences in the level of anti-A and anti-Bamong ethnic populations have been reported; these differenceswere found to be the result of environmental factors rather thanhereditary factors. In the present study, the anti-A and anti-B titersof random donors in three Asian populations are compared. InThailand, the IgM anti-A and anti-B titers are low and are similar tothe Japanese titers reported in 2001, but the IgG anti-A and anti-Btiters are high and are similar to the Japanese titers reported in1986. In the Lao People’s Democratic Republic, both the IgM andthe IgG anti-A and anti-B titers are high and are similar to thosereported in Japan in 1986. In addition, anti-A and anti-B titers ofdifferent sex donors and of various age groups were also compared.High titers were found in 8.8 percent of the female donors in theyounger than 30 age group and in 36.7 percent of the femaledonors in the older than 50 age group. Immunohematology2007;23:38–41.

Key Words: ABO blood groups, anti-A, anti-B

Passively transfused blood group antibodies causeclinical problems.1 The effects of anti-A and anti-B inABO-mismatched platelet transfusions, especially inHLA-matched platelet transfusions, have long beendebated. Although reports are rare, intravascularhemolytic transfusion reactions have been known tooccur.2,3 These hemolytic transfusion reactions seem tobe caused by high titers of anti-A and anti-B. Suchreactions have been seen in groups A, B, and ABrecipients receiving group O plasma.

Anti-A and anti-B levels have been reported to differamong ethnic populations.4 Many previous studieshave suggested that ethnicity is a risk factor for ABO

HDN. However, the differences among populationswere found to be the result of environmental factorsrather than hereditary factors.5,6 Redman et al.6

addressed the question of variations in ABO antibodylevels in persons with different ethnic backgrounds bystudying antibody levels in Asian, Caucasian, andAfrican blood donors, all of whom were living in thenorth London area of the United Kingdom. Althoughthe highest levels of IgG anti-A and anti-B titers werefound among group O African female donors, theselevels were not significantly higher than those in theother group O donors who were tested.

In this report, we investigated the changes in theanti-A and anti-B titers in the Japanese populationduring the last 19 years. Both anti-A and anti-B titersdecreased greatly during this period. We alsocompared these titers with the anti-A and anti-B titersin Laotian and Thai populations.

Materials and MethodsSera were obtained from Japanese blood donors in

1986 (n = 106), 2001 (n = 107), and 2005 (n = 93) inTokyo, Japan. In 2001, sera were obtained from blooddonors in Vientiane, Lao People’s Democratic Republic(Lao PDR; n = 58); in 2005, sera were obtained fromblood donors in Bangkok,Thailand (n = 93). All serumsamples were obtained randomly from the generalpopulation of volunteer unremunerated blood donors.The sera were stored at –30°C until tested.

Sera from group O Japanese blood donors classifiedaccording to age (16–29 years old and 51–69 years old)

Differences in ABO antibodylevels among blood donors: acomparison between past andpresent Japanese, Laotian, andThai populationsT.MAZDA, R.YABE, O. NATHALANG,T.THAMMAVONG,AND K.TADOKORO


ABO antibody levels in Asians

and sex were also collected forthe present study. These serumsamples were separate fromthose collected from thegeneral population in 1986,2001, and 2005.

The sera were titrated usinga 1 to 1 dilution with 0.01MPBS, pH 7.2, containing 2% BSA.The IgM antibody titers weredetermined using the salinemethod with incubation atroom temperature for 15minutes. The IgG antibodytiters were determined by anIAT using LISS and anincubation of 30 minutes at37°C. Before the IAT, the serawere treated with 2-ME for 15minutes at 37°C using themethod described in the AABBTechnical Manual7; thistreatment is used to destroy IgMantibodies in the sera.

Testing was performed bythe same person to avoidvariations in technique and testinterpretation.

ResultsIn the Japanese population,

anti-A and anti-B titers in 1986were distributed between 64and 1024 (mode 256) for IgM(Table 1) and between 64 and2048 (mode 256–512) for IgG(Table 2). In 2001, they mark-edly decreased to between 16and 512 (mode 32–64) for bothIgM and IgG. In 2005, the IgMtiters further decreased tobetween 2 and 64 (mode 8–16).In the Laotian population in2001, the IgM titers were similarto those of the 1986 Japanesepopulation; the distributionwas between 128 and 1024(mode 256), whereas the IgGtiters were slightly higher thanthose of the 1986 Japanese

Table 1. Distribution of anti-A and anti-B IgM titers in three populations

Anti-A titer 2 4 8 16 32 64 128 256 512 1024 Total (n)

Laos 2001* – – – – – – 2 36 8 6 52

Thailand 2005* – 6 8 17 14 12 11 7 – – 75

Japan 1986* – – – – – 6 3 67 7 3 86

Japan 2001* – – – 2 16 51 8 1 1 – 79

Japan 2005* – 9 18 27 14 7 – – – – 75

Japan (group O)16–29 male 2 9 28 32 26 5 – – – – 10216–29 female 3 14 32 31 20 2 – – – – 10251–69 male 3 24 32 23 12 – – – – – 9451–69 female 10 9 13 9 6 2 – – – – 49

Anti-B titer 2 4 8 16 32 64 128 256 512 1024 Total (n)

Laos 2001* – – – – – – – 36 3 1 40

Thailand 2005* – 3 5 17 11 8 12 6 – – 62

Japan 1986* – – – – – 6 8 68 2 4 88

Japan 2001* – – – 11 69 9 2 2 – – 93

Japan 2005* 2 5 17 27 7 4 – – – – 62

Japan (group O)16–29 male – 12 43 23 19 5 – – – – 10216–29 female 3 37 30 23 7 2 – – – – 10251–69 male 11 19 49 14 1 – – – – – 9451–69 female 8 10 21 4 5 1 – – – – 49

* Data include group O and nongroup O donors

Table 2. Distribution of anti-A and anti-B IgG titers in three populations

Anti-A titer 2 4 8 16 32 64 128 256 512 1024 Total (n)

Laos 2001* – – – – – 13 15 16 6 2 52

Thailand 2005* 1 8 8 9 23 15 5 3 3 – 75

Japan 1986* – – – 1 3 39 21 20 2 – 86

Japan 2001* – 2 16 51 8 1 1 – – – 79

Japan 2005* 2 22 24 21 6 – – – – – 75

Japan (group O)16–29 male 2 14 33 28 14 11 – – – – 10216–29 female 2 22 23 34 9 11 1 – – – 10251–69 male – 6 18 35 32 3 – – – – 9451–69 female – 2 18 14 4 6 2 3 – – 49

Anti-B titer 8 16 32 64 128 256 512 1024 2048 4096 Total (n)

Laos 2001* – – – – – 1 23 11 3 2 40

Thailand 2005* 1 1 1 7 12 19 15 5 1 – 62

Japan 1986* – – – – 1 2 28 32 22 3 88

Japan 2001* – 11 69 9 2 2 – – – – 93

Japan 2005* – 17 20 16 8 1 – – – – 62

Japan (group O)16–29 male 3 28 37 27 5 2 – – – – 10216–29 female 1 26 39 17 6 4 4 5 – – 10251–69 male 2 15 40 29 8 – – – – – 9451–69 female – 4 15 4 3 5 10 8 – – 49

* Data include group O and nongroup O donors


T.MAZDA ET AL.

population: between 256 and 4096 (mode 512–1024).In the Thai population in 2005, IgM titers were similarto those of the 2001 Japanese population: the distri-bution was between 4 and 256 (mode 16), and IgGtiters were similar to those of the 1986 Japanesepopulation: the distribution was between 8 and 2048(mode 128–256).

In the Japanese group O blood donors, IgM titers ofanti-A and anti-B were low and no differences werenoted between the sexes. However, IgG titers of anti-Aand anti-B showed differences between the sexes andan increase with donor age. High titers (> 512) of anti-A, anti-B, or both were found in 8.8 percent (9 in 102)of the female donors in the younger than 30 age groupand in 36.7 percent (18 in 49) of the female donors inthe older than 50 age group, determined by theantihuman globulin test. No male donors were foundwith titers more than 256.

DiscussionIn the Japanese population, the anti-A and anti-B

titers markedly decreased within the 15-year periodbetween 1986 and 2001. At present, the Japanese anti-A and anti-B titers are relatively low, compared withthose seen in the Laotian and Thai populations.Schwartz et al.8 showed that only 3.3 to 3.6 percent ofAmerican group O blood donors had high anti-A andanti-B titers (> 100). People with high ABO systemantibody titers are now rare among the Japanesepopulation, and the mean titers are lower than those ofblood donors in NewYork.

The mechanisms responsible for the reductions inJapanese anti-A and anti-B titers are unknown.However, previous studies have suggested thatenvironmental factors may influence anti-A and anti-Btiters.5,6 Enteric bacteria and other parasites may affectthe production of anti-A and anti-B.4,6 Environmentalfactors can affect the prevalence, counts, and suscepti-bility of enteric bacteria.9,10 Because environmentalfactors are thought to affect anti-A and anti-B titers,wecompared the anti-A and anti-B titers in three Asiancountries: Japan, Lao PDR, and Thailand. Japan has longbeen a developed country, Lao PDR is a developingcountry, and Thailand has recently been givendeveloped country status.

The environment in Japan continues to change.Recently, the Japanese lifestyle has become morewesternized, especially with regard to food.11 Theprevalence of allergic diseases, heart disease, diabetes,and cancer has been increasing in Japan. In contrast,

Lao PDR remains relatively undeveloped, and Laotiansusually eat natural foods, whereas Thais and theJapanese eat more processed foods. Theseenvironmental differences may explain, at least in part,why the Laotian anti-A and anti-B titers were similar tothe titers observed in Japanese donors in 1986,whereas the IgM and IgG antibody titers among theThais were similar to those observed in the Japanesedonors in 2001 and 1986, respectively. Our datasupport the concept that anti-A and anti-B titers areaffected by environmental factors. Lao PDR is rapidlydeveloping and becoming more westernized. We planto continue our comparison of Laotian and Thaiantibody titers in the future.

AcknowledgmentWe thank Dr.W.L. Gyure, Seton Hall University, for

his support in the preparation of this manuscript.

References1. Garratty G. Problems associated with passivelytransfused blood group alloantibodies. Am J ClinPathol 1998;109:769–77.

2. McManigal S, Sims KL. Intravascular hemolysissecondary to ABO incompatible platelet products:an underrecognized transfusion reaction. Am JClin Pathol 1999;111:202–6.

3. Larsson LG,Welsh VJ, Ladd DJ. Acute intravascularhemolysis secondary to out-of-group platelettransfusion.Transfusion 2000;40:902–6.

4. Issitt PD,Anstee DJ.Applied blood group serology.4th ed. Durham: Montgomery Scientific Publi-cations, 1998:188.

5. Toy PTCY, Reid ME, Papenfus L,Yeap HH, Black D.Prevalence of ABO maternal-infant incompatibilityin Asians, Blacks, Hispanics and Caucasians. VoxSang 1988;54:181–3.

6. Redman M, Malde R, Contreras M. Comparison ofIgM and IgG anti-A and anti-B levels in Asian,Caucasian and Negro Donors in the North WestThames Region.Vox Sang 1990;59:89–91.

7. Brecher ME, ed. Method 2.11. Use of sulfhydrylreagents to disperse autoagglutination. TechnicalManual, 15th ed. Bethesda: American Associationof Blood Banks 2005:744–5.

8. Schwartz J, Depalma H, Kapoor K, Hamilton T,Grima K. Anti-A titers in group O single donorplatelets: to titer or not to titer (abstract).Transfusion 2003;43(Suppl):115A.


ABO antibody levels in Asians

9. Björkstén B, Naaber P, Sepp E, Mikelsaar M. Theintestinal microflora in allergic Estonian andSwedish 2-year-old children. Clin Exp Allergy1999;29:342–6.

10. Björkstén B, Sepp E, Julge K,Voor T, Mikelsaar M.Allergy development and the intestinal microfloraduring the first year of life. J Allergy Clin Immunol2001;4:516–20.

11. McCurry J. Japanese people warned to curbunhealthy lifestyles. Lancet 2004;363:1126.

Toshio Mazda, PhD, (corresponding author) andKenji Tadokoro, MD, PhD, Japanese Red Cross CentralBlood Institute, Tatsumi 2-1-67, Koto-ku, Tokyo 135-8521, Japan; Ryuichi Yabe BS, Japanese Red CrossWest Tokyo Blood Center, Tokyo, Japan; OytipNaThalang, PhD, Department of Pathology,Phramongkutklao College of Medicine, Bangkok,Thailand;Te Thammavong, MD, Lao Red Cross BloodTransfusion Centre, Vientiane, Lao PDR.

Attention SBB and BB Students: You are eligible for a free 1-year subscription to Immunohematology. Askyour education supervisor to submit the name and complete address for each student and the inclusive datesof the training period to Immunohematology, P.O. Box 40325, Philadelphia, PA 19106.


C O M M U N I C A T I O N

Missing Scott Murphy, a platelet “maestro”

The most recent occasion I had to see Scott before his untimely death was at the 29th BEST meeting, held inRome in March 2005. Early spring in Rome is unique, as the local environment provides an ideal link between thepast—witnessed by its gorgeous monuments—and future dreams and expectations inspired by the Ponentinobreeze caressing the seven hills of Rome.

I met Scott as he exited his hotel on the via Veneto. I knew his disease had started a new aggressive phase, buthis optimistic smile overcame my concerns when he addressed me with a warm “Buongiorno!” The meeting wasscientifically very successful, as usual for all BEST gatherings, but most notable was Scott’s piano performanceduring dinner, when he played four-hand Diabelli’s Sonatine with Georges Andreu, another musically giftedtransfusion medicine professional. This performance was the second in a series begun at an earlier BEST meetingin Paris, where Scott played Mozart’s Twinkle,Twinkle Little Star with the devoted help of his wife Joanie playingthe triangle during a dinner at the Musée D’Orsay.

While everyone knows of Scott’s scientific accomplishments, it was his blend of science and art that trulymade him a platelet “maestro.” Our scientific interests crossed on several occasions, mainly surrounding theEuropean method to produce platelets for transfusion—the buffy-coat technique—as compared with the“platelet-rich-plasma” procedure used in the United States. In particular, he was instrumental in studies developed byFrancesco Bertolini in his laboratory and ours, elucidating the role of acetate in platelet storage.

Finally, remembering Scott without mentioning his deep love and dedication to his wife, their children, andgrandchildren would be forgetting one of his most important qualities, his humanity. We, his friends, along withthe many patients who have and will continue to benefit from his work,will miss the scientist, the gentleman, andthe maestro.

Paolo Rebulla, MDDirector

Center of Transfusion Medicine, Cellular Therapy and CryobiologyOspedale Maggiore Policlinico, Mangiagalli e Regina Elena

Via Francesco Sforza 3520122 Milano MI

Italy


I N M E M O R I A M

Katherine M. Beattie, BS, MT(ASCP)SBB 1922–2007

Kay Beattie received her BS in medical technology fromWayne State Universityin Detroit, Michigan, in 1943 and began her career at Children’s Hospital in Detroit.She became the supervisor of the Blood Bank at Children’s in 1954. In 1957, shereceived her SBB certification, and in 1961, she became the co-director of theMichigan Community Blood Center in Detroit. In 1975, she joined the SoutheasternMichigan region of the American Red Cross (ARC) Blood Services as director of thereference laboratory and education, a position she held until she retired to NorthPalm Beach, Florida, in 1988.

During her 45-year career, Kay was active at the local, state, and national levels,and was honored often by her peers. She was a member of the Ontario Antibody

Club from 1966 to 1988; the Michigan Blood Council, as chairman of the Reference Laboratory Committeefrom 1977 to 1988; the Invitational Conference of Investigative Immunohematologists from 1976 to 1988;and the ARC Accreditation Reference Committee from 1984 and as chairman from 1987 to 1988. As amember of the Michigan Association of Blood Banks (MABB) since its founding in 1956, Kay had servedas chairman of two committees and in all offices, including the presidency. She also served on the AABBCommittees on Education,Reference Laboratories,and Quality Control as vice chairman;Scientific SectionCoordinating Committee as secretary; as an inspector in the Inspection Program; and on the Committeeon Self-Testing and Review.

Among her awards from the AABB, she received the Ivor Dunsford Memorial Award in 1972; theprestigious Emily Cooley Lectureship Award in 1980, which was given for the first time to a medicaltechnologist; and the Chapman-Franzmier Memorial Award in 1998. The MABB has honored Kay fourtimes, with the Merit Award in 1963, the Technical Award in 1971, the Founders Award in 1985, and theKay Beattie Lecture, a lecture given in her honor each year at the MABB meeting. Kay received theDistinguished Alumni Award in 1982 fromWayne State University.

Kay published 44 scientific papers and was the co-editor of the ARC Immunohematology Methodsand Procedures Manual. Her final paper, on resolving ABO discrepancies, was published in 1993 in theMethods Manual.

She was a superb reference technologist and had a phenomenal memory for patients and theantibodies they made. Those of us who knew her will remember her as a mentor who always encouragedus to try harder and do better. For example, she got us on committees and asked us to give talks. Sheintroduced us to famous people at meetings and could always explain something new in blood banking.Kay was always a great friend with a ready joke. When she retired, she did not look back. She took hergolf clubs and went at golf like she went at antibodies. When she had it figured out, she was the women’sclub champion for many years. Her humor, intelligence, generosity, friendship, honesty, and jokes will begreatly missed.

Delores Mallory


Attention: State Blood Bank Meeting OrganizersIf you are planning a state meeting and would like copies of Immunohematology for distribution, pleasecontact Cindy Flickinger,Managing Editor, 4 months in advance, by fax or e-mail at (215) 451-2538 [email protected].

E R R A T U M

Vol. 22, No. 4, 2006; page 176

Review:molecular basis of MNS blood group variants

The author has informed the editors of Immunohematology that there is an error on page 176, second column,second paragraph, third sentence. The sentence should read “Some of the classes of Miltenberger did not reactwith anti-Mia but reacted with one or more of the other three specific antisera, e.g., GP.Hil (Mi.V) RBCs did notreact with anti-Mia but did react with anti-Hil.”

Free Classified Ads and AnnouncementsImmunohematology will publish classified ads and announcements (SBB schools, meetings, symposia, etc.)without charge. Deadlines for receipt of these items are as follows:

Deadlines1st week in January for the March issue1st week in April for the June issue

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Notice to Readers: All articles published,including communications and book reviews,reflect the opinions of the authors and do notnecessarily reflect the official policy of theAmerican Red Cross.

A N N O U N C E M E N T S

Monoclonal antibodies available at no charge:The NewYork Blood Center has developed a wide range of monoclonal antibodies (both murine and humanized)that are useful for donor screening and for typing RBCs with a positive DAT. These include anti-A1, -M, -s, -U, -D, -Rh17, -K, -k, -Kpa, -Jsb, -Fy3, Wrb, -Xga, -CD99, -Dob, -H, -Ge2, -CD55, -Oka, -I, and anti-CD59. Most of the antibodies aremurine IgG and require the use of anti-mouse IgG for detection (Anti-K, k, -Kpa, and -Fya). Some are directlyagglutinating (Anti-M, -Wrb and -Rh17) and one has been humanized into the IgM isoform (Anti-Jsb). The antibodiesare available at no charge to anyone who requests them. Please visit our Web site for a complete list of availablemonoclonal antibodies and the procedure for obtaining them.

For additional information,contact:Gregory Halverson,NewYork Blood Center,310 East 67th Street,NewYork,NY10021; e-mail: [email protected]; or visit the Web site at http://www.nybloodcenter.org >research>immunochemistry >current list of monoclonal antibodies available.

Meetings!

June 6–8 Blood Banks Association of New York State (BBANYS)The 56th annual meeting of the Transfusion Medicine Symposium, Blood Banks Association of New York State(BBANYS), cosponsored by the NewYork State Department of Health,will be held June 6 through 8, 2007, at theDesmond Hotel and Conference Center in Albany, New York. For more information, contact Kevin Pelletier at(518) 485-5341or [email protected], or refer to theWeb site at http://www.bbanys.org.

June 7–8 Heart of America Association of Blood Banks (HAABB)The spring meeting of the Heart of America Association of Blood Banks (HAABB) will be held June 7 and 8, 2007,at the Embassy Suites in Kansas City,Missouri. For more information, refer to theWeb site at http://www.haabb.org.


A D V E R T I S E M E N T S

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Applications are invited from medical or science graduates to study for the MSc in Transfusion andTransplantation Sciences. The course is run jointly by The Department of Cellular & Molecular Medicine,University of Bristol and the Bristol Institute of Transfusion Sciences.

The course starts in October 2007 and can be studied full-time for 1 year or part-time over 2 or 3years by block release.

The course aims to develop your interest, knowledge and understanding of the theory, practical tech-niques and research relating to the science of transplantation and transfusion medicine.

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Teaching combines informal lectures, tutorials, practical laboratory experience and a research projectwith the bias on transfusion.

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The course is accredited by The Institute of Biomedical Sciences and directed byProfessor David Anstee and Dr Tricia Denning-Kendall.

For further details visit:

http://www.blood.co.uk/ibgrl/MSc/MScHome.htmor contact:Dr Tricia Denning-Kendall,University of Bristol, Geoffrey Tovey Suite,National Blood Service, Southmead Rd Bristol, BS10 5ND, England.TEL 0117 9912093, E-MAIL [email protected]


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Immunohematology for style. Double-space throughout the manuscript.Number the pages consecutively in the upper right-hand corner, beginningwith the title page.

II. SCIENTIFIC ARTICLE, REVIEW, OR CASE REPORT WITHLITERATURE REVIEWA. Each component of the manuscript must start on a new page in thefollowing order:1. Title page2. Abstract3. Text4. Acknowledgments5. References6. Author information7. Tables8. FiguresB. Preparation of manuscript1. Title page

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c. Materials and MethodsSelection and number of subjects, samples, items, etc. studied anddescription of appropriate controls, procedures, methods, equipment,reagents, etc. Equipment and reagents should be identified inparentheses by model or lot and manufacturer’s name, city, and state.Do not use patient’s names or hospital numbers.

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III. EDUCATIONAL FORUMA. All submitted manuscripts should be approximately 2000 to 2500words with pertinent references. Submissions may include:1. An immunohematologic case that illustrates a sound investigative

approach with clinical correlation, reflecting appropriate collaborationto sharpen problem solving skills

2. Annotated conference proceedingsB. Preparation of manuscript1. Title page

a. Capitalize first word of title.b. Initials and last name of each author (no degrees; all CAPs)

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Send all manuscripts by e-mail to [email protected]

ImmunohematologyJOURNAL OF BLOOD GROUP SEROLOGY AND EDUCATION

Instructions to the Authors


What is a certified Specialist in Blood Banking (SBB)?• Someone with educational and work experience qualifications who successfully passes the American Society forClinical Pathology (ASCP) board of registry (BOR) examination for the Specialist in Blood Banking.

• This person will have advanced knowledge, skills, and abilities in the field of transfusion medicine and blood banking.

Individuals who have an SBB certification serve in many areas of transfusion medicine:• Serve as regulatory, technical, procedural, and research advisors• Perform and direct administrative functions • Develop, validate, implement, and perform laboratory procedures• Analyze quality issues, preparing and implementing corrective actions to prevent and document issues• Design and present educational programs• Provide technical and scientific training in blood transfusion medicine• Conduct research in transfusion medicine

Who are SBBs?Supervisors of Transfusion Services Managers of Blood Centers LIS Coordinators EducatorsSupervisors of Reference Laboratories Research Scientists Consumer Safety OfficersQuality Assurance Officers Technical Representatives Reference Lab Specialist

Why be an SBB?Professional growth Job placement Job satisfaction Career advancement

How does one become an SBB?• Attend a CAAHEP-accredited Specialist in Blood Bank Technology Program OR• Sit for the examination based on criteria established by ASCP for education and experience

Fact #1: In recent years, the average SBB exam pass rate is only 38%.Fact #2: In recent years, greater than 73% of people who graduate from CAAHEP-accredited programs pass the SBB

exam.

Conclusion:The BEST route for obtaining an SBB certification is to attend a CAAHEP-accredited Specialist in Blood BankTechnology Program

Becoming a Specialist in Blood Banking (SBB)

Contact the following programs for more information:PROGRAM CONTACT NAME CONTACT INFORMATION

Walter Reed Army Medical Center William Turcan 202-782-6210;[email protected]

Transfusion Medicine Center at Florida Blood Services Marjorie Doty 727-568-5433 x 1514; [email protected]

Univ. of Illinois at Chicago Veronica Lewis 312-996-6721; [email protected]

Medical Center of Louisiana Karen Kirkley 504-903-2466; [email protected]

NIH Clinical Center Department of Transfusion Medicine Karen Byrne 301-496-8335; [email protected]

Johns Hopkins Hospital Christine Beritela 410-955-6580; [email protected]

ARC-Central OH Region, OSU Medical Center Joanne Kosanke 614-253-2740 x 2270; [email protected]

Hoxworth Blood Center/Univ. of Cincinnati Medical Center Catherine Beiting 513-558-1275; [email protected]

Gulf Coast School of Blood Bank Technology Clare Wong 713-791-6201; [email protected]

Univ. of Texas SW Medical Center Barbara Laird-Fryer 214-648-1785; [email protected]

Univ. of Texas Medical Branch at Galveston Janet Vincent 409-772-4866; [email protected]

Univ. of Texas Health Science Center at San Antonio Bonnie Fodermaier SBB Program: 210-358-2807,Linda Smith [email protected]

MS Program: 210-567-8869; [email protected]

Blood Center of Southeastern Wisconsin Lynne LeMense 414-937-6403; [email protected]

Additional information can be found by visiting the following Web sites: www.ascp.org, www.caahep.org, and www.aabb.org

Musser Blood Center700 Spring Garden StreetPhiladelphia, PA 19123-3594

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ImmunohematologyThe Journal of Blood Group Serology and Education

published quarterly by TheAmerican National Red Cross2007 Subscription Application

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ImmunohematologyMethods and Procedures

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