JOURNAL OF CLINICAL MICROBIOLOGY, Mar. 1989, p. 474-4790095-1137/89/030474-06$02.00/0Copyright © 1989, American Society for Microbiology

Development and Evaluation of Immunoassay for Detection ofAntibodies to the Feline T-Lymphotropic Lentivirus (Feline

Immunodeficiency Virus)THOMAS P. O'CONNOR, JR.,l* SHERYL TANGUAY,' ROBIN STEINMAN,l ROBERTA SMITH,'MARGARET C. BARR,2 JANET K. YAMAMOTO,3 NIELS C. PEDERSEN,3 PHILIP R. ANDERSEN,'

AND QUENTIN J. TONELLI1IDEXX Corp., Portland, Maine 041011; School of Veterinary Medicine, University of California, Davis, California 956163;

and College of Veterinary Medicine, Cornell University, Ithaca, New York 148532Received 19 September 1988/Accepted 22 November 1988

The feline T-cell lymphotropic lentivirus (feline immunodeficiency virus) is a recently described feline-specific retrovirus that can produce chronic immunodeficiency-like disorders in cats. A microdilution plateformat enzyme-linked immunosorbent assay has been developed to detect the presence of antibody to the virusin feline serum or plasma. Temporal studies performed with experimentally infected animals show thatseroconversion can be demonstrated 3 to 4 weeks after exposure to the virus. Results of a serosurvey (n = 1,556samples) indicate that infection is fairly common in both clinic (5.2%) and sick cat (15.2%) populations.Western blot (immunoblot) and sodium dodecyl sulfate radioimmunoprecipitation assays were developed toconfirm microdilution plate test results and to identify peptides specific for the feline immunodeficiency virus.All microdilution plate test positive results and selected negative results were confirmed by one or both of theseprocedures. These data demonstrate that this microassay plate enzyme-linked immunosorbent assay is a verysensitive and specific test for detection of antibody to the feline immunodeficiency virus.

A recently identified retrovirus has been shown to be thecausitive agent of a chronic immunodeficiency-like syn-drome in cats (21). In initial reports the virus was namedfeline T-lymphotropic lentivirus to reflect the tropism of thevirus for feline T cells and the classification of the agent as amember of the lentivirus subfamily of retroviruses (12, 22).The name of the virus has been recently changed to felineimmunodeficiency virus (FIV) to conform with internationalnomenclature for immunodeficiency-linked viruses (24). TheFIV agent has a strong but not absolute tropism for the felineT-lymphocyte cell line, which may be responsible for theimmunosuppressive nature of the virus. Viral particle mor-phology and the Mg2+ metal requirement of the viral reversetranscriptase are supportive of the classification of the virusas a lentivirus.The virus is distinct from previously described feline

retroviruses and represents the first report of a feline-specificlentivirus (21). Members of the lentivirus subfamily infectingother species include the human immunodeficiency virus(HIV) (8, 15), visna virus in sheep (19), caprine arthritis-encephalitis virus (7), equine infectious anemia virus (20),bovine immunodeficiency virus (9), and simian immunodefi-ciency virus (3).The clinical course of FIV infection appears to be charac-

terized by a lengthy asymptomatic phase persisting severalmonths or perhaps years, during which viral infection can bedemonstrated but clinical symptoms are not apparent (24).This period of apparent viral latency is typical of lentiviralinfections and frequently precedes the development of clin-ical abnormalities (10). Clinical symptoms most commonlyassociated with FIV infection include rhinitis and gingivi-tis, anemia, diarrhea, pustular dermatitis, and generalizedlymphadenopathy (12, 22). Symptoms vary among infected

* Corresponding author.

animals but are characterized by a chronic and persistentnature.The FIV is infectious within feline populations and can be

transmitted after intimate and prolonged contact. Initialreports suggest that biting may be an important mode of viraltransmission (24). Isolates of FIV have been identified in theUnited States (21), the United Kingdom (11), and Japan (13).A limited serosurvey in the United States reported that 1 in18 healthy cats and 10 of 25 unhealthy cats were seropositivefor FIV infection (21).The continuous propagation of FIV in an established

Crandell feline cell line has permitted the isolation andpurification of large quantities of virus. The highly purifiedvirus was used to develop a sensitive microdilution plate-based enzyme-linked immunosorbent assay (ELISA) fordetection of feline antibody specific for the FIV agent. In thisreport we describe the development of the FIV microdilutionplate ELISA, partially characterize immunoreactive viralpeptides, and present data on the prevalence of viral infec-tion in several populations (n = 1,556). The results of thissurvey demonstrate that FIV infection is relatively commonand widespread in the United States.

MATERIALS AND METHODS

Virus and cell culture. The FIV was propagated in chron-ically infected Crandell feline kidney (Crfk) cells (6). Thevirus was concentrated from tissue culture fluids by precip-itation with polyethylene glycol (4) and purified by densitygradient centrifugation on glycerol gradients as previouslydescribed (18).The FIV microdilution plates were prepared by coating

microdilution wells with 100 ,ul of an inactivated detergent-disrupted preparation of FIV antigen (5.0 ,ug/ml). Samples offeline serum or plasma were diluted (1:100) in 20 mM sodiumphosphate-150 mM NaCl (pH 7.4) containing calf serum andbovine serum albumin. Diluted samples (100 pt) were ap-

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IMMUNOASSAY FOR ANTIBODIES TO FIV 475

plied to individual microdilution wells and incubated for 30min at room temperature. The contents of the microdilutionplates were aspirated, and the plates were washed (fivetimes) and incubated (30 min, room temperature) with goatanti-feline horseradish peroxidase conjugate (Kirkegaardand Perry). The contents of the microdilution plates wereaspirated, the plates were washed (five times), and substratesolution (100 ,ul) was added. The substrate solution wasprepared immediately before the assay by mixing equivol-ume portions of 0.1% 3,3',5,5'-tetramethylbenzidine in 60%methanol-40% glycerol with 0.04% hydrogen peroxide in 0.1M dibasic potassium phosphate-citrate (pH 4.8) containing0.01% Thimerosal. The enzymatic reaction was carried outfor 15 min at room temperature and then stopped by additionof 100 ,ul of dilute hydrofluoric acid solution (1:400). Indi-vidual microdilution well optical density values were deter-mined spectrophotometrically at 650 nm.To minimize interassay variability, ELISA optical density

values were normalized by using positive and negativecontrol reagents assayed on each microdilution plate. Thepresence or absence of antibody to FIV was determined byrelating the A650 of the sample to the A6,( of the positivecontrol reagent. The positive control contains a standardizedlevel of antibody to FIV in feline serum. The relative level ofFIV-specific antibody in the sample was determined bycalculating the sample-to-positive (S/P) ratio in the followingmanner: (sample A650 - negative control A650)!(positivecontrol A65O - negative control A650). Samples with S/Pratios less than 0.5 are classified negative. S/P ratios equal toor greater than 0.5 are positive for antibody to FIV.

Clinical samples. The FIV ELISA was used to screen atotal of 1,556 samples obtained from five sources: 223 fromthe Boston Refuge League (Boston, Mass.), 130 from theChicago Cat Clinic (Chicago, 111.), 459 from Clinipath Lab-oratories (Valparaiso, Ind.), 145 from the Veterinary Refer-ence Laboratory (Dallas, Tex.), and 599 from the Universityof California (San Diego). Samples from the Boston RefugeLeague, Chicago Cat Clinic, Clinipath, and the VeterinaryReference Laboratory were not selected on the basis ofprevious disease status and represent a collection of samplesreceived by these clinics and laboratories. Samples obtainedfrom the University of California were obtained from ani-mals with a previous unspecified disease or illness. This setof samples should be regarded as comprising a sick catpopulation.Samples for the seroconversion studies were obtained

from the Cornell Feline Health Center (Ithaca, N.Y.) and theUniversity of California (Davis). These were obtained fromspecific-pathogen-free animals after inoculation with eitherinfectious cell free tissue culture fluid (1 ml), FIV-infectedwhole blood (1 ml), or 0.44-,um-filtered plasma (2 ml) frominfected animals.Western blot assay. The Western blot (immunoblot) proto-

col used was a modification of the procedure initially de-scribed by Towbin et al. (23). Purified FIV was disruptedwith sodium dodecyl sulfate (SDS) and mercaptoethanol andseparated with a 10% SDS-polyacrylamide gel. Viral pro-teins were transferred to nitrocellulose sheets, which werethen blocked with detergent and calf serum. Individual stripswere cut from each sheet and incubated for 2 h with a 1:100dilution of sample prepared in a phosphate-buffered salinesolution containing 0.05% Tween 20, 1% bovine serumalbumin, and 30% calf serum. Strips were washed exhaus-tively with phosphate-buffered saline-0.05% Tween 20 andincubated for 1 h with goat anti-feline horseradish peroxi-dase conjugate. The wash cycle was repeated, and the

precipitating substrate 4-chloronaphthol was added. Reac-tions were stopped by washing with deionized H20, andresults were immediately interpreted. The presence of twoor more FIV viral bands was necessary to confirm a sampleas positive for antibody to FIV. The molecular weights ofFIV-reactive peptides were determined by comparison ofimmunoperoxidase-stained Western blot strips with Coo-massie blue-stained polyacrylamide gels and amido black-stained nitrocellulose strips (R. Steinman, T. O'Connor, Q.Tonelli, K. Lawrence, C. Seymour, J. Goodness, N. Peder-sen, and P. R. Andersen, submitted for publication). Unin-fected Crfk host cell extract was used to prepare Westernblot strips to identify reactive bands unrelated to FIVinfection. These were used in the identification of nonspe-cific bands appearing on FIV blot strips.RIPA-SDS-PAGE. For the radioimmunoprecipitation as-

say (RIPA) with SDS-polyacrylamide gel electrophoresis(PAGE), the FIV productively growing in cell culture wasmetabolically labeled with [35S]methionine and [35S]cysteineat 37°C for 4 h. Cells were lysed in 10 mM phosphate buffer(pH 7.5) containing 100 mM NaCI, 1% Triton X-100, 0.5%sodium deoxycholate, 0.1% SDS, 0.1 mM phenylmethylsul-fonyl fluoride, and 100 Kallikren inactivator units of aproti-nin per ml (lysis buffer). Cell lysates were clarified bycentrifugation at 100,000 x g for 30 min before use. Gener-ally, 100 ,ul of viral extract was mixed with 5 ,ul of serumsample and incubated for 18 h at 3°C. Samples were concen-trated by adding 0.2 ml of a 5% suspension of proteinA-Sepharose CL-4B (Pharmacia Fine Chemicals) in 10 mMphosphate (pH 7.5)-100 mM NaCl-1% Triton X-100-0.1%SDS and centrifuging. The pellet was washed three times inlysis buffer, heated in sample buffer, and applied to a 12%SDS-polyacrylamide gel (14). Gels were processed for fluo-rography and exposed at -70°C to Kodak XAR-5 film.

RESULTS

FIV ELISA detection of seroconversion in response toexperimental infection. Serum samples from a total of 14experimentally infected cats were obtained to assess thesensitivity of the microdilution plate test with respect toseroconversion. Temporal samples were obtained for eachanimal and assayed by the FIV ELISA. Western blots and/orRIPA-SDS-PAGE were used to confirm both positive andnegative results.

Seroconversion was demonstrated in all animals by 4weeks after the initial exposure. Microdilution plate data forcats 2429 and 2838 are shown in Fig. 1 and are typical of theentire series of experimentally infected animals. Westernblots and RIPA-SDS-PAGE confirmatory test data areshown in Fig. 2 through 5 and demonstrate seroconversionparalleling the microdilution plate test results. With theexception of one cat, which died during the study (week 14),all cats remained seropositive during the testing period.Most of the experimentally infected animals showed sim-

ilar clinical manifestations, including an initial neutropenialasting 4 to 5 weeks, fever lasting several days postinfection,and a generalized enlargement of peripheral lymph nodesoccurring 4 weeks postinfection and persisting for 2 to 9months. However, with the exception of the single cat citedabove, none of the experimentally infected animals devel-oped symptoms associated with late-stage FIV infections(24).The Western blot assay of the two temporal series showed

initial reaction to the FIV gag proteins piS and p26 (Fig. 2and 3). Reaction to additional FIV proteins (plO, p32, gp4O,

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476 O'CONNOR ET AL. J. CLIN. MICROBIOL.

FTLV TEMPORAL SAMPLE2429

0 r I0 0.5 1 1.5 2 2.5 3 S 7 9 il 13 15

MIONTHS AFTER INOCULATION

FTLV TEMPORAL SAMPLE2838

3.5ri -3

2-

15

0-o o.s 1 1.5 2 2.5 3 5 7 9 il 13 15

MONTHS AFTER INOCULATIONFIG. 1. Microdilution plate assay results of serum samples ob-

tained sequentially during the course of experimentally induced FIVinfection in cats 2429 and 2838. S/P ratios were calculated asdescribed in Materials and Methods.

p47, and p65) occurred during the course of infection(Steinman et ai., submitted). Serum samples from specific-pathogen-free cats with experimentally induced FIV infec-tion were flot reactive on immunoblots of HIV, HIV type 2,simian immunodeficiency virus, caprine arthritis-encepha-titis virus, or feline leukemia virus (24). The RIPA-SDS-PAGE assay showed positive results after approximately thesame time period as the microdilution plate test and Westernblot assay (Fig. 4 and 5). FIV proteins immunoprecipitatedby the temporal samples were identified as gpl3O, pilO, p47,gp4O, p36, and p22 (Steinman et ai., submitted).FIV ELISA: clinical evaluation. The frequency distribu-

tions for the clinic and sick cat populations are shown in Fig.6 and 7, respectively. Results are plotted as frequencyversus the S/P ratio. As described in Materials and Methods,an S/P ratio greater than 0.5 indicates that the samplecontains significant levels of antibody to FIV. The seropos-itive rate was 5.2% for the clinic population (n = 957) and15.2% for the sick cat population (n = 599).

A B C D E F G H K L

-r

p65

_lu

gp4Op32p26 _ : - _ _ _M

pl5 - - _n _. _ |-_

li.Li.LL ÈLGLLL LLFIG. 2. Western blot strips after assay of serum samples ob-

tained sequentially during the course of experimentally induced FIVinfection in cat 2429. Results are shown for samples obtained at thefollowing times (months) postinfection (lanes): A, preimmunizationbleed; B, 0.75; C, 1.0; D, 1.25; E, 2.0; F, 3.0; G, 4.0; H, 5.0; t, 7.0;J, 9.0; K, 11.0; L, 14.0.

Western blot and/or RIPA-SDS-PAGE confirmatory as-says were conducted on all samples that tested positive bythe microdilution plate test. Each of these samples waspositive by the confirmatory procedures. Western blot as-says of FIV antibody-positive samples typically showed astrong reaction to the FIV gag proteins p26 and pi5 andvariable reactivity to additional FIV proteins (10,000,32,000, 40,000, 47,000, and 65,000 daltons) (Steinman et al.,submitted). Staining of two or more FIV peptides wasrequired for positive result confirmation by Western blot-ting, whereas reaction to gpl3O was required for confirma-tion by RIPA-SDS-PAGE. The RIPA-SDS-PAGE assays ofpositive samples demonstrated variable reactions to proteinsof 130,000, 110,000, 47,000, 40,000, 36,000, 22,000, and15,000 daltons; however, all plate test-positive samplesimmunoprecipitated gpl3O. A small percentage (3%) of mi-crodilution plate test-positive samples were reactive to onlya single FIV peptide (p26 or piS) in the Western blot assay.These samples were positive when tested using the RIPA-SDS-PAGE assay. A set of 150 plate test-negative sampleswere negative when tested by the Western blot assay.

A B C D E F G H J K L

p65

gp4O

p264 _çw i-r

I â- à" _p15 - w'[plo ' I

FIG. 3. Western blot strips after assay of serum samples ob-tained sequentially during the course of experimentally induced FIVinfection in cat 2838. Results are shown for samples obtained at thefollowing times (months) postinfection (lanes): A, preimmunizationbleed; B. 0.5; C, 1.0; D, 1.5; E, 2.0; F, 2.5; G, 3.0; H, 4.0; I, 5.0; J,6.0; K. 8.0; L, 13.0.

I

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IMMUNOASSAY FOR ANTIBODIES TO FIV 477

A B C D E F G H J K L M N O P Q R

gpl30 ` i i - -p110 t w to

p47gp4O

p36

p22

FIG. 4. RIPA-SDS-PAGE results for serum samples obtainedsequentially during the course of experimentally induced FIV infec-tion in cat 2429. Results are shown for samples obtained at thefollowing times (months) postinfection (lanes): A, preimmunizationbleed; B, 0.25; C, 0.75; D, 0.80; E, 1.0; F, 1.25; G, 2.0; H, 2.25, I,3.0; J, 4.0; K, 5.0; L, 7.0; M, 9.0; N, 10.0; O, 11.0; P, 13.0; Q, 14.0;R, 16.0.

DISCUSSIONThe seroconversion studies demonstrate that a significant

antibody level to FIV can be measured 3 to 4 weeks afterinitial exposure to the virus. The long time course (13 to 14months) of continued antibody response shown for cats 2429and 2838 demonstrate the persistent nature of infection. Theextended disease-free period observed for 13 of 14 experi-mentally infected cats may reflect the long incubation periodof the virus. A similar disease-free state has been welldocumented after seroconversion to HIV in humans (16).Western blot assays show the initial reaction to the FIV

gag proteins (primarily piS and p26), whereas the RIPA-SDS-PAGE technique showed an initial reaction to the FIVenv proteins (gp130, pllO, gp40). Reaction to additional viralproteins was observed during the course of infection withboth techniques. This pattern of antibody response, and thediffering capacity of the Western blot and the RIPA-SDS-PAGE techniques to detect sets of virus-specific antibodies,

A B D E F 5 " K L M N 0 P a R

gpO30D1 10 htipt*q t t

p47gp40D36

... .. fa du m _ @o22

FIG. 5. RIPA-SDS-PAGE results for serum samples obtainedsequentially during the course of experimentally induced FIV infec-tion in cat 2838. Results are shown for samples obtained at thefollowing times (months) postinfection (lanes): A, preimmunizationbleed; B, 0.25; C, 0.50; D, 1.0; E, 1.25; F, 1.50; G, 1.75; H, 2.0; 1,2.25; J, 2.50; K, 3.0; L, 4.0; M, 5.0; N, 6.0; 0, 8.0; P, 9.75; Q, 13.0;R, 14.0.

450-4so-400

350-

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00

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150-

100-

50-

zw

I

w

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CLINIC POPULATIONFTLV NEGATIVE N _ 907

o 0.5 1 1.5 2 2.5 3 3.5 4

S/P RATIO

CLINIC POPULATIONFTLV POSIVE N =50

6-

4-

3

2-

o

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S/P RATIO

FIG. 6. FIV microdilution plate assay results showing frequencyplotted versus S/P ratio for the FIV-negative (n = 907) and FIV-positive (n = 50) clinic population. S/P ratios were calculated asdescribed in Materials and Methods. Samples with S/P ratios greaterthan 0.5 are classified as positive.

has been observed in HIV antibody studies in humans (1, 2,5). The naturally occurring antibody response to FIV in catsmay serve as a useful model for the similar response to HIVin humans.The FIV microdilution plate test can be evaluated by

examining the distribution of assay results for a collection ofpositive and negative samples relative to the assay cutoff.ELISA plate test data have been reported for a total of 1,556samples. The mean S/P ratio was 0.019 (standard deviation,0.048) for the combined negative population and 2.46 (stan-dard deviation, 1.18) for the overall positive population. Theassay cutoff value (S/P ratio, 0.5) is positioned 0.481 units or10.02 standard deviations above the mean of the S/P ratio forthe negative samples. Similarly, the assay cutoff is belowassay results for all 141 positive samples tested and substan-tially below the mean S/P ratio for the positive population.None of the samples included in this study was incorrectlyidentified by the microdilution plate test when assayed by

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478 O'CONNOR ET AL.

SICK CAT POPULATIONFTLV NEGATIVE N = 508

disease is as high as reported, the benefit of FIV screeningfor veterinary clinicians would be substantial.

K LITERATURE CITEDw l 1. Allain, J. P., D. A. Paul, Y. Laurian, and D. Senn. 1986.O - Serological markers in early stages of human immunodeficiency

M -virus infection in hemopheliacs. Lancet ii:1233-1236.

O l 2. Barin, F., M. F. McLane, J. S. Allan, T. H. Lee, J. E.O l Groopman, and M. Essex. 1985. Virus envelope protein of;o - HTLV-III represents major target antigen for antibodies inMo AIDS patients. Science 228:1094-1096.Io 3. Benveniste, R. E., L. O. Arthur, C.-C. Tsai, R. Sowder, T. D.

Copeland, L. E. Henderson, and S. Oroszlan. 1986. Isolation ofa lentivirus from a macaque with lymphoma: comparison with

1o HTLV-III/LAV and other lentiviruses. J. Virol. 60:483-490.o 4. Bishop, D. H. L., R. Ruprecht, R. W. Simpson, and S. Spiegel-o- man. 1971. Deoxyribonucleic acid polymerase of Rous sarcomaIo- L virus: reaction conditions and analysis of the reaction productnucleic acids. J. Virol. 8:730-741.

o- lTIl,.,,,,,,,1,,,,,,,|,,,,,2 2.5|Ul§|l§||||| 3.5llllllglll5. Burke, D. S., R. R. Redfield, P. Putman, and S. S. Alexander.O 0.5 I 1.5 2 2.5 3 3.5 4 1987. Variations in Western blot banding patterns of human

S/P RATIO T-cell lymphotropic virus type III/lymphadenopathy-associatedSICK CAT POPULATION virus. J. Clin. Microbiol. 25:81-84.6. Crandell, R. A., and E. Q. Despeau. 1959. Cytopathology ofFTLV POSmVE N = 91 feline viral rhinotracheitis virus in tissue cultures of feline renal

cells. Proc. Soc. Exp. Biol. Med. 154:1403-1418.7. Dahlberg, J. E., J. M. Gaskin, and K. J. Perk. 1981. Morpho-

s5- logical and immunological comparison of caprine arthritis en-cephalitis and ovine progressive pneumonia viruses. J. Virol.39:914-919.

4- 8. Gallo, R. C., S. Z. Salahuddin, M. Popovic, M. Kaplan, B. F.Haynes, R. Redfield, J. Oleske, B. Sofai, G. M. Shearer, J. T.Palker, G. White, P. Foster, and P. D. Markham. 1984. Frequent

3- detection and isolation of cytopathic retroviruses (HTLV-III)from patients with AIDS and at risk for AIDS. Science 224:500-502.

2-9. Gonda, M. A., M. J. Braun, S. G. Carter, T. A. Kost, J. W. Bess,Jr., L. G. Arthur, and M. J. Van der Maaten. 1987. Character-ization and molecular cloning of a bovine lentivirus related to

-1"i- | | |human immunodeficiency virus. Nature (London) 330:388-392.10. Haase, A. T. 1986. Pathogenesis of lentivirus infections. Nature

(London) 322:130-136.0- riTlrr r ntmlT r 11. Harbour, D. A., P. D. Williams, T. J. Gruffyd-Jones, J. Bur-G 0.5 1 1.5 2 2.5 3 3.5 4 bridge, and G. R. Pearson. 1988. Isolation of a T-lymphotropic

S/P RATIO lentivirus from a persistently leucopenic domestic cat. Vet. Rec.Y. 7. FIV microdilution plate assay results showing frequency 122:8Hd86.d versus S/P ratio for the FIV-negative (n = 508) and FIV- 12. Hardy, W. D. 1988. Feline T-lymphotropic lentivirus: retrovi-ve (n = 91) sick cat population. S/P ratios were calculated as rus-induced immunosuppression in cats. J. Am. Anim. Hosp.ibed in Materials and Methods. Samples with S/P ratios greater Assoc. 24:241-243.).5 are classified as positive. 13. Ishida, T., T. Washizu, K. Toriyabe, S. Motoyoshi, and I.Tomoda. 1988. Detection of feline T-lymphotropic lentivirus

(FTLV) infection in Japanese domestic cats. Jpn. J. Vet. Sci.50:39-44.

Western blot or RIPA-SDS-PAGE procedures. The 14. Laemmli, U. K. 1970. Cleavage of structural proteins during thevely large standard deviation in S/P ratio observed for assembly of the head of bacteriophage T4. Nature (London))ositive population reflects variation in antibody re- 227:680-685.se in individual animals. 15. Levy, J. A., A. D. Hoffman, S. M. Kramer, J. A. Lendis, J. M.results of this serosurvey indicate that FIV infection Shimabukuro, and L. S. Oshiro. 1984. Isolation of lymphocyto-pathic retroviruses from San Francisco patients with AIDS.despread in both the clinic (5.2%) and the sick cat Science 225:840-842.

%'o) populations. The substantially higher incidence of 16. Lui, K., D. N. Lawrence, W. M. Morgan, T. A. Peterman,infection in the sick cat population apparently reflects H. W. Haverkos, and D. J. Bregman. 1986. A model-basedisease-causing potential of the virus. The overall sero- approach for estimating the mean incubation period of transfu-ive rate is similar to the feline leukemia virus seropos- sion-associated acquired immunodeficiency syndrome. Proc.rate reported for similar populations (17). The identifi- Natl. Acad. Sci. USA 83:3051-3055.n of FIV as a transmissable disease-causing agent and 17. McMichael, J. C., S. Stiers, and S. Coffin. 1986. Prevalence ofemonstration of the extent of infection in cat popula- feline leukemia virus infection among adult cats at an animalhas serious implications in veterinary medicine. The control center: association of viremia with phenotype and.ositive rate found for .oth the clinic and the sick cat season. Am. J. Vet. Res. 47:765-768.positive rate ound nforbot the clinic and the sick cat 18. Montelaro, R. C., N. Lohrey, B. Parekh, E. W. Blakeney, and

nations suggests that infection by FIV will provide a C. J. Issel. 1982. Isolation and comparative biochemical prop-ingful characterization for previously undiagnosed dis- erties of the major internal polypeptides of equine infectioussyndromes in cats. If the potential of the virus to cause anemia virus. J. Virol. 42:1029-1038.

26

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IMMUNOASSAY FOR ANTIBODIES TO FIV 479

19. Narayan, O., D. E. Griffin, and J. Chase. 1977. Antigenic shift invisna virus in persistently infected sheep. Science 197:376-378.

20. Parekh, B., C. J. lssel, and R. C. Montelaro. 1980. Equineinfectious anemia virus, a putative lentivirus, contains polypep-tides analogous to prototype-C oncornaviruses. Virology 107:520-525.

21. Pedersen, N. C., E. W. Ho, M. L. Brown, and J. K. Yamamoto.1987. Isolation of a T-lymphotropic virus from domestic catswith an immunodeficiency-like syndrome. Science 235:790-793.

22. Sparger, E. E. 1988. Feline T-lymphotropic lentivirus infection.

Feline Med. (Vet. Learning Systems) 4:9-14.23. Towbin, H., T. Staehelin, and J. Gordon. 1979. Electrophoretic

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24. Yamamoto, J. K., E. Sparger, E. W. Ho, P. R. Andersen, T. P.O'Connor, C. P. Mandell, L. Lowenstine, R. Munn, and N. C.Pedersen. 1988. The pathogenesis of experimentally inducedfeline immunodeficiency virus (FIV) infection in cats. Am. J.Vet. Res. 49:1246-1258.

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