Analytical validation of a reference laboratory ELISA for the detection of feline leukemia virus p27 antigen

Buch, Jesse; Clark, Genevieve H; Cahill, Roberta; Thatcher, Brendon; Smith, Peter J. S.; Chandrashekar, Ramaswamy; Leutenegger, Christian M.; O’Connor, Thomas P.; Beall, Melissa J.

doi:10.1177/1040638717710451

Cited by 10 publications

(24 citation statements)

References 12 publications

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“…The manufacturer reports this test has a sensitivity of ≥94.9% and a specificity of ≥98.4% defined by combined results of other p27 antigen test brands and virus isolation. PetChek ® FeLV 15 ELISA was performed with sequential screening and confirmatory protocols, including a neutralization step to reduce nonspecific reactivity as described . The sensitivity and specificity of this modified assay were both reported to be 100% defined by the combined results of FeLV PCR and ViraCHEK ® /FeLV .…”

Section: Methodsmentioning

confidence: 99%

“…PetChek ® FeLV 15 ELISA was performed with sequential screening and confirmatory protocols, including a neutralization step to reduce nonspecific reactivity as described . The sensitivity and specificity of this modified assay were both reported to be 100% defined by the combined results of FeLV PCR and ViraCHEK ® /FeLV . A total of 10 samples were discarded because they were positive on PetChek ® and negative on ViraCHEK ® .…”

Section: Methodsmentioning

confidence: 99%

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Performance of 4 Point‐of‐Care Screening Tests for Feline Leukemia Virus and Feline Immunodeficiency Virus

Levy

Crawford

Tucker

2017

Veterinary Internal Medicne

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BackgroundMore than 3 million cats in the United States are infected with FeLV or FIV. The cornerstone of control is identification and segregation of infected cats.Hypothesis/ObjectivesTo compare test performance with well‐characterized clinical samples of currently available FeLV antigen/FIV antibody combination test kits.AnimalsSurplus serum and plasma from diagnostic samples submitted by animal shelters, diagnostic laboratories, veterinary clinics, and cat research colonies. None of the cats had been vaccinated against FIV. The final sample set included 146 FeLV+, 154 FeLV−, 94 FIV+, and 97 FIV− samples.MethodsProspective, blind comparison to a gold standard: Samples were evaluated in 4 different point‐of‐care tests by ELISA antigen plate tests (FeLV) and virus isolation (FIV) as the reference standards. All test results were visually read by 2 blinded observers.ResultsSensitivity and specificity, respectively, for FeLV were SNAP ® (100%/100%), WITNESS ® (89.0%/95.5%), Anigen® (91.8%/95.5%), and VetScan® (85.6%/85.7%). Sensitivity and specificity for FIV were SNAP ® (97.9%/99.0%), WITNESS ® (94.7%/100%), Anigen® (96.8%/99.0%), and VetScan® (91.5%/99.0%).Conclusions and Clinical ImportanceThe SNAP ® test had the best performance for FeLV, but there were no significant differences for FIV. In typical cat populations with seroprevalence of 1–5%, a majority of positive results reported by most point‐of‐care test devices would be false‐positives. This could result in unnecessary segregation or even euthanasia.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Performance of 4 Point‐of‐Care Screening Tests for Feline Leukemia Virus and Feline Immunodeficiency Virus

Levy

Crawford

Tucker

2017

Veterinary Internal Medicne

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“…Consequently, inclinic diagnosis of FeLV infection relies on antigen testing rather than antibody testing. 7,8,16,21,[40][41][42] FeLV PCR testing is not without its limitations, with false-positives caused by DNA contamination during PCR set up and falsenegatives caused by mutations in the targeted region of the virus being possible. Evaluation of FeLV PoC test kit performance is challenging because of the absence of a clearly defined 'gold standard' for diagnosing FeLV infection.…”

Section: Felv Diagnosis Using Whole Blood and Poc Antigen Test Kitsmentioning

confidence: 99%

“…14,39 For more detail about other diagnostic methods of FeLV detection, including virus isolation, immunofluorescence for detection of FeLV cell-associated p27 antigen in circulating leukocytes and platelets, and most recently a laboratory-based ELISA to detect p27 antigen (PetChek FeLV 15™), we refer to studies published elsewhere. 7,8,16,21,[40][41][42] FeLV PCR testing is not without its limitations, with false-positives caused by DNA contamination during PCR set up and falsenegatives caused by mutations in the targeted region of the virus being possible. However, one important advantage of PCR testing over all other currently available diagnostic methodologies is its ability to detect regressive FeLV infections.…”

Section: Felv Diagnosis Using Whole Blood and Poc Antigen Test Kitsmentioning

confidence: 99%

Diagnosing feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) infection: an update for clinicians

2019

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With the commercial release in Australia in 2004 of a vaccine against feline immunodeficiency virus (FIV; Fel‐O‐Vax FIV®), the landscape for FIV diagnostics shifted substantially. Point‐of‐care (PoC) antibody detection kits, which had been the mainstay for diagnosing FIV infection since the early 1990s, were no longer considered accurate to use in FIV‐vaccinated cats, because of the production of vaccine‐induced antibodies that were considered indistinguishable from those produced in natural FIV infections. Consequently, attention shifted to alternative diagnostic methods such as nucleic acid detection. However, over the past 5 years we have published a series of studies emphasising that FIV PoC test kits vary in their methodology, resulting in differing accuracy in FIV‐vaccinated cats. Importantly, we demonstrated that two commercially available FIV antibody test kits (Witness™ and Anigen Rapid™) were able to accurately distinguish between FIV‐vaccinated and FIV‐infected cats, concluding that testing with either kit offers an alternative to PCR testing. This review summarises pertinent findings from our work published in a variety of peer‐reviewed research journals to inform veterinarians (particularly veterinarians in Australia, New Zealand and Japan, where the FIV vaccine is currently commercially available) about how the approach to the diagnosis of FIV infection has shifted. Included in this review is our work investigating the performance of three commercially available FIV PoC test kits in FIV‐vaccinated cats and our recommendations for the diagnosis of FIV infection; the effect of primary FIV vaccination (three FIV vaccines, 4 weeks apart) on PoC test kit performance; our recommendations regarding annual testing of FIV‐vaccinated cats to detect ‘vaccine breakthroughs’; and the potential off‐label use of saliva for the diagnosis of FIV infection using some FIV PoC test kits. We also investigated the accuracy of the same three brands of test kits for feline leukaemia virus (FeLV) diagnosis, using both blood and saliva as diagnostic specimens. Based on these results, we discuss our recommendations for confirmatory testing when veterinarians are presented with a positive FeLV PoC test kit result. Finally, we conclude with our results from the largest and most recent FIV and FeLV seroprevalence study conducted in Australia to date.

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“…The core protein of FeLV, p27, is a soluble antigen that is produced in great excess which makes it ideal for detection by immunoassays. Several of the immunoassays that detect p27 antigen in cats employ a pair of high affinity, monoclonal antibodies that, when optimized in an enzyme-linked immunosorbent assay (ELISA), can be very sensitive and specific [22][23][24]. Studies in both experimentally infected and naturally infected cats have demonstrated a strong association between the copy numbers of proviral DNA and p27 antigen level in the circulation and clinical course of infection [5,25].…”

Section: Introductionmentioning

confidence: 99%

Feline Leukemia Virus p27 Antigen Concentration and Proviral DNA Load Are Associated with Survival in Naturally Infected Cats

Beall

Buch

Clark

et al. 2021

Viruses

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Longitudinal studies of cats naturally infected with feline leukemia virus (FeLV) are important for understanding disease outcomes. Levels of p27 antigen and copy numbers of proviral DNA have been associated with FeLV-infection courses. The purpose of this prospective study was to establish cutoff values for p27 antigen concentration and proviral DNA load that distinguished high positive from low positive groups of cats and to evaluate an association with survival. At enrollment, 254 cats were tested by point-of-care and microtiter plate enzyme-linked immunosorbent assays (ELISAs) for p27 antigen and real-time polymerase chain reaction (PCR) for proviral DNA. The 127 positive cats were retested monthly for six months and monitored for survival over the four-year study. A receiver operating characteristic-based analysis of samples with concordant or discordant qualitative results for p27 antigen and proviral DNA was used to establish cutoff values, and when applied to test results at enrollment for classifying cats as high positive or low positive, a significant difference in survival was observed. High positive cats had a median survival of 1.37 years (95% CI 0.83–2.02) from time of enrollment, while most low positive cats were still alive (93.1% survival). Quantitative results for p27 antigen concentration and proviral DNA load were highly correlated with survival times in FeLV-infected cats.

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Analytical validation of a reference laboratory ELISA for the detection of feline leukemia virus p27 antigen

Cited by 10 publications

References 12 publications

Performance of 4 Point‐of‐Care Screening Tests for Feline Leukemia Virus and Feline Immunodeficiency Virus

Performance of 4 Point‐of‐Care Screening Tests for Feline Leukemia Virus and Feline Immunodeficiency Virus

Diagnosing feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) infection: an update for clinicians

Feline Leukemia Virus p27 Antigen Concentration and Proviral DNA Load Are Associated with Survival in Naturally Infected Cats

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