CONCURRENT 10: THE COMPLETE BLOOD COUNT 10: the complete blood count and beyond: ... the complete blood count and beyond: quality issues and reference methods ... further advances in automation

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<ul><li><p>CONCURRENT10:</p><p>THECOMPLETEBLOODCOUNTANDBEYOND:QUALITYISSUESANDREFERENCEMETHODS</p><p>SaturdayMay14th 20161:30 3:00PM(13.30 15.00hrs)</p><p>InternationalSocietyforLaboratoryHematology (ISLH)</p></li><li><p>CONCURRENT10:</p><p>THECOMPLETEBLOODCOUNTANDBEYOND:QUALITYISSUESANDREFERENCEMETHODS</p><p>CHAIRS:Dr.PieroCappelletti.SIPMEL,Pordenone, Italy</p><p>Dr.AlbertHuisman,UMCUtrecht,Utrecht,Netherlands</p><p>InternationalSocietyforLaboratoryHematology (ISLH)</p></li><li><p>VerificationandQualityControlofautomatedHematologyAnalyzers</p><p>Dr.AlbertHuisman,UMCUtrecht,Utrecht,Netherlands</p><p>InternationalSocietyforLaboratoryHematology (ISLH)</p></li><li><p>VerificationandQualityControlofautomatedHematologyAnalyzers</p><p>Dr.AlbertHuisman,UMCUtrecht,Utrecht,Netherlands</p><p>InternationalSocietyforLaboratoryHematology (ISLH)</p></li><li><p>Disclosure information:</p><p>Dr AlbertHuisman:Nothing to disclose</p><p>TheUMCUtrecht,department ofClinical ChemistryandHematology hasreceived fundingfor contractresearchfrom:</p><p>Abbott DiagnosticsBeckman-CoulterDiagnostic GrifolsMechatronics</p></li><li><p>VerificationandqualitycontrolofautomatedHematologyAnalyzers AutomatedCellAnalysis(CompleteBloodCount(CBC))</p><p> Including5partLeukocytedifferentialcount,reticulocytecountandnewparameters(ReticulocyteHemoglobincontentReticulatedPlatelets/ImmaturePlateletFraction,)</p><p> Validationofanewanalyzer Verificationofanewanalyzer</p><p> ISO15189 CLSI,ICSH,others Samples Precision Accuracyandcomparability Sensitivityandspecificity Referenceintervals</p><p> Qualitycontrol Daily/internal ExternalQualityAssesment (EQA/Proficiencytesting)</p></li><li><p>Why?</p><p>Why bother about verification and qualitycontrolofahighly automated analyzer?</p></li><li><p>Because:</p><p> CBCresults formthe starting pointofnumerous diagnosticschedules,treatments and interventions,for example: Erytrocyte /Platelet transfusions Work upofanemia (irondeficiency /thalassemia /) Hematological malignancies Generalwork-upinvarious disease states Etctetera</p></li><li><p>Because:</p><p> TheISO15189standard[Medicallaboratories Particularrequirementsforqualityandcompetence]requiresaverificationprocess</p></li><li><p>AdvantagesofAutomatedCellAnalysers:</p><p> Excellentanalytical performance Closed-tubeanalysis Nointer-observer variability Noslidedistribution error Eliminate statistical variations Potential ofreflextesting Availabilityofextraparameterse.g.MCV,RDW,%rP, Moreefficient (&gt;100analyses/hour)and cost effective than</p><p>manualmethod .</p></li><li><p>Automatedcellcounters:</p><p> CompleteBloodCount(CBC): Hemoglobin(Hb)concentration,RBCcount&amp;RBCindices(MCV,</p><p>MCH,MCHC),WBCcount,PLTcount WBCdifferentialcount(5normalWBCs) RDW,MPV Reticulocytecount NucleatedRBCcount(NRBCs) .Flagging,etc etc</p><p> Newparameters: AdvancedRBCparameters:%microcyticRBCs,Reticulocyte</p><p>Hemoglobincontent,.. ReticulatedPlatelets/ImmaturePlatelets .</p></li><li><p>Furtheradvancesinautomation</p><p> Multipleinlineanalysers Built-inSlide-Maker-Stainer Auto-validationinLIS Automatedreflex/morphologyetc.</p><p> Includingdigitalmorphologyorflow-cytometry solutions AllFullbloodanalysisinline(ESR,HbA1c,..)</p><p>TotalLaboratoryAutomation DigitalMorphology</p></li><li><p>Validation ofanewanalyzer:goal</p><p> Provision ofobjective evidence that ahematologyanalyzerfulfills specified requirements (where thespecifiedrequirements areadequatefor intended use).</p><p> Validation isprimarily amanufacturersresponsibility toensure that designgoalsaremetand performanceclaimsarestated (including safety and effectiveness).</p><p> Validation study (manufacturer): collectdatatosupportaregulatorysubmissionandtheregistrationofahematologyanalyzer.</p></li><li><p>Validation ofanewanalyzer:goal</p><p> Provision ofobjective evidence that ahematologyanalyzerfulfills specified requirements (where thespecifiedrequirements areadequatefor intended use).</p><p> Validation isprimarily amanufacturersresponsibility toensure that designgoalsaremetand performanceclaimsarestated (including safety and effectiveness).</p><p> Validation study (manufacturer): collectdatatosupportaregulatorysubmissionandtheregistrationofahematologyanalyzer.</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Objectives ofavalidation study: Generate datato assess safety and clinical efficacy from amedical perspective Develop performanceinformationfor labelingandmarketingpurposes Validate appropriate operationalperformancecharacteristics ofthe</p><p>hematology analyzerinatypical end-usersetting Develop datathat areused to supportsubmissionoftheproductfor approval</p><p>orclearanceby regulatory bodies</p><p>Source:CLSIH26A2</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Validation performancespecifications: Limitofblank(LoB,background) Carryover Imprecision (reproducibility),shorttermand long-term Analytical measuring interval(AMI)(linearity) Lower limitofdetection (LLoD)and lower limitofquantitation (LLoQ) Comparibility (correlation) Interferences Frequency and typeofdatainvalidations</p><p>Source:CLSIH26A2</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Performancespecifications:current referencemethods*: Selective microscopy for WBCdifferential RBCcount andWBCcount (impedance) Selective microhematocrit (PCV)for hematocrit (HCT) Hemoglobin by hemiglobincyanidemethod Selective PLTmonoclonalantibody (contemporary referencemethod) Reticulocytes by flowcytometry</p><p>Source:CLSIH26A2</p><p>*Standardized and independent ofmanufacturer</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Performancespecifications:current referencemethods: When noreferencemethod isavailable:usually comparisonwith previous</p><p>generation instrument</p><p>Source:CLSIH26A2</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Current referencemethods,problems,need for improvement: Current referencemethods aremoreorless outdated</p><p> e.g.:microscopic 400cell WBCdifferential count:elaborative and imprecise</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Current referencemethods,problems,need for improvement: Current referencemethods aremoreorless outdated</p><p> e.g.:microscopic 400cell WBCdifferential count:elaborative and imprecise Noreferencemethods available for clinically relevantparameters*:</p><p> MCV ExtendedRBCparameters Reticulocyte Hb content MPV Reticulated platelets/immatureplatelet fraction .</p><p> *Clinically relevantparametersarenot standardized! This may leadto differences between differentHematology analyzers:</p><p> Differences inclinical interpretation (confusion for clinicians) Mayprevent wider use Invalid aggregation ofdata</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>Validationofanewanalyzer(manufacturer)</p><p> Current referencemethods,problems,need for improvement: Current referencemethods aremoreorless outdated</p><p> e.g.:microscopic 400cell WBCdifferential count:elaborative and imprecise Noreferencemethods available for clinically relevantparameters:</p><p> MCV ExtendedRBCparameters Reticulocyte Hb content MPV Reticulated platelets/immatureplatelet fraction .</p><p> Urgentneed for improvement,role for professionalsocieties</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>Verification</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Theendusershould asseswhether the manufacturers claimsonperformanceofthe specific instrumentalso apply to theintended use criteriasetby the laboratory:Verification</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Theverification process includes performanceanalysisof: Accuracy Precision Reportable rangeoftestresults and reference intervals (normal ranges) Background(limitofblank) Carryover (sample) Lower limits ofdetection Quantitation Clinically reportable intervals (CRIs)</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Accordingto ISO15189acertain levelofverification ofanynew(hematology)analyzerhasto be done (preferably)according toaprofessionalstandard.</p><p> ISO15189also stimulates laboratories to takepatient riskfactorsinto consideration tomeetthesestandards</p><p> There arecurrenly 2internationaldocuments available forverification ofahematology analyzer: Consensusdocuments with recommendations (not based onstrong</p><p>evidence butrather onexpertopinion).</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Accordingto ISO15189acertain levelofverification ofanynew(hematology)analyzerhasto be done (preferably)according toaprofessionalstandard.</p><p> ISO15189also stimulates laboratories to takepatient riskfactorsinto consideration tomeetthesestandards</p><p> There arecurrenly 2internationaldocuments available forverification ofahematology analyzer: Clinical Laboratory StandardsInstitute (CLSI)(standardH26A2)</p><p>published in2010 InternationalCouncilonStandardisation inHematology (ICSH)</p><p>guidelinepublished in2014.</p><p>Huismanetal.Clin LabMed 2015</p></li><li><p>Consensusdocuments with recommendations</p><p>CLSI ICSH(publishedintheInternationalJournalofLaboratoryHematology,theofficialISLH journal</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHVerification Thelaboratory(enduser)</p><p>should followtheproceduresofmanufacturervalidation,buttheverificationmaybeabbreviated;thegoalistoverifythatthemanufacturersstatedperformance iscorrect</p><p>Verification isconformationoftheevaluationperformedbythemanufacturerorpublished intheliterature;theverificationmaybeabbreviated(ie,focusedtomeetspecificrequirementsatthetestsite)</p><p>Precision/imprecision(within-run reproducibility,closeness ofagreementbetween testresults</p><p>Should be performed withnormal samplesand samplesatmedical decision levelsavailable inclinicallaboratories.Nospecifications for number ofmeasurements and reporting</p><p>Single runof10measurements onthe samesample(all reportedparameters),3levels(normal, abnormal low,andabnormal high,aroundclinical decision points).Reported asSDand %CV</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHVerification Thelaboratory(enduser)</p><p>should followtheproceduresofmanufacturervalidation,buttheverificationmaybeabbreviated;thegoalistoverifythatthemanufacturersstatedperformance iscorrect</p><p>Verification isconformationoftheevaluationperformedbythemanufacturerorpublished intheliterature;theverificationmaybeabbreviated(ie,focusedtomeetspecificrequirementsatthetestsite)</p><p>Precision/imprecision(within-run reproducibility,closeness ofagreementbetween testresults</p><p>Should be performed withnormal samplesand samplesatmedical decision levelsavailable inclinicallaboratories.Nospecifications for number ofmeasurements and reporting</p><p>Single runof10measurements onthe samesample(all reportedparameters),3levels(normal, abnormal low,andabnormal high,aroundclinical decision points).Reported asSDand %CV</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHPrecision(betweenbatch,longterm</p><p>Should beperformedwithnormalsamplesandsamplesatmedicaldecision levelsavailableinclinicallaboratories.Nospecificationsfortimeperiodandnumberofmeasurements</p><p>Single sample,repeateddaily,foraperiodof2030d.Threelevels(allparameters,abnormallowandabnormalhigh,aroundclinicaldecisionpoints).Fixedblood(controlmaterial)mayberequired</p><p>Comparability(comparison betweenevaluation HAandcurrent HA)</p><p>Should be performedbutnot specified, ifdifferentmodesareavailable,whether anextensive mode-to modecomparability should beperformed</p><p>Atleast 250300samples,measured onboth HAwith sampleswith various disordersand with interferingsubstances</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHPrecision(betweenbatch,longterm</p><p>Should be performedwith normal samplesand samplesatmedicaldecision levelsavailableinclinical laboratories.Nospecifications fortimeperiod and numberofmeasurements</p><p>Single sample,repeateddaily,for aperiod of2030d.Threelevels(allparameters,abnormallowand abnormal high,around clinical decisionpoints).Fixed blood(controlmaterial)maybe required</p><p>Comparability(comparisonbetweenevaluationHAandcurrentHA)</p><p>Should beperformedbutnotspecified, ifdifferentmodesareavailable,whetheranextensivemode-tomodecomparabilityshouldbeperformed</p><p>Atleast 250300samples,measured onboth HAwith sampleswith various disordersand with interferingsubstances</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHAccuracy (closeness ofagreementbetweenmeasurement and truevalue)</p><p>Should be performed;not otherwise specified</p><p>Depending onavailabilityofreferencemethod,often notapplicable;inpracticeoften compared withcurrent HA</p><p>Referenceintervals Mustbe established orverified for all reportableparameters</p><p>Should be calculated forall components oftheCBC;atleast 120samplesfrom apparentlyhealthy individuals (60male,60female)</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>CLSI ICSHAccuracy (closeness ofagreementbetweenmeasurement and truevalue)</p><p>Should be performed;not otherwise specified</p><p>Depending onavailabilityofreferencemethod,often notapplicable;inpracticeoften compared withcurrent HA</p><p>Referenceintervals Mustbe established orverified for all reportableparameters</p><p>Should be calculated forall components oftheCBC;atleast 120samplesfrom apparentlyhealthy individuals (60male,60female)</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Samples: Bewareofpre-analytical variables!</p><p> Timeisone ofthe mostimportantvariablesdue to timedependentalterations ofcells (volumeand morphology)</p><p> Usually anonymous surplusmaterial Broad rangeofunderlyingpathology (garantee that all cells are</p><p>recognized) Results should encompass the entire analytical range(very low</p><p>(e.g.extremetrombocytopenia)to very highlevels(e.g.extremeleucocytosis /CML).</p><p> Ageand gender(pediatric samples?) All typesoftubesthatmay enterthe laboratory</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>HuismanetalClin LabMed 2015</p></li><li><p>InstrumentVerification by the enduserlaboratory:Precision</p><p>Accuracyistheproximityofameasurementresulttothetruevalueandmainlydependentonsystematicerror(thetermbiasshouldbeavoided);precisionisthereproducibilityofthemeasurementsandmainlydependentonrandomerror.</p><p>HuismanetalIJKH2016</p></li><li><p>Precision</p><p> Thedesired precision ofaCBCparameterisdependentonthe biological variation ofthis parameter Work ofGeorgeCembrowski etal</p><p> (Clin LabMed 2015;IJLH2016).</p></li><li><p>Cembrowski etalIJLH2016</p></li><li><p>Precision:</p><p> Inorderto generate (clinically)relevantresults the analyticalprecision (%CVreproducibility)ofaCBCresult should be lessthan halfand preferable less than ofthe biologicalvariation.</p><p> Stateofthe artpossibilities ofcurrentgeneration ofHA</p></li><li><p>InstrumentVerification by the enduserlaboratory</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>Deanalytical %CVdoesaddto the total %CV</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>Roomfor improvement:ReticulocytesWBCWBCdifferential</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Carry-over,isit important? Background,isit important?</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Carry-over,isit important? Example:carry over1%,</p><p> Sample1:platelet count 1000x109/L Sample2:platelet count 10x10exp9/L(subsequent sample) Acarry-overof1%will result inafalsely increased platelet count of</p><p>20x109/L(100%increase)</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p> Background,isit important? Abackground(for example intheWBCchannel)can leadto</p><p>false positiveresults inCSFsamples</p></li><li><p>InstrumentVerification by the enduserlaboratory</p><p>Reference:TracyGeorgeetal.</p></li><li><p>QualityControl</p><p> InternalQualityControl Performedonadailybasis Electronicbuiltinqualitycontrolflags</p><p> Aretheremechanical/electronical problems? Controlmaterialsoftenprovidedbymanufacturer</p><p> Usually3-controllevels(low-/medium-/high- level) ~Widerange</p><p> Oftenmanipulatedblood 1versusmultipletimes/daydependingonstabilityofanalyser Comparisonbetweendifferentanalysers possible Expensive 3...</p></li></ul>