ELISA r-p24: optimization and validation of an enzyme immunoassay for the diagnosis of Feline Immunodeficiency Virus infections

Abstract

The Feline Immunodeficiency Virus (FIV) is a lentivirus that affects cats worldwide. The detection of the virus’ 24 Kda capsid protein (p24) antibody is the most common way to diagnose FIV infections.  In Brazil, the most used commercial assay for FIV diagnosis is a rapid ELISA test based on the lateral flow principle, with high sensitivity and specificity (SNAP^®test). However, the high cost of this imported assay undermines not only the diagnosis of this infection but also its epidemiology. The objective of this study was to optimize and validate an indirect ELISA using the protein p24 recombinant antigen (r-p24) with national technology and maintaining sensitivity and specificity comparable to those of available commercial rapid ELISA tests. Twenty-six reference cats’ sera samples (13 negative and 13 positives for FIV), previously analyzed by PCR and the SNAP^® test, were used as a reference to optimize the ELISA. Serum samples of 226 cats from private owners and from shelters of the metropolitan area of Rio de Janeiro were used to validate the assay. The sensitivity and specificity of the r-p24 ELISA were 95.4% and 99.4%, respectively. When compared to the SNAP^® Combo Test, the accuracy of r-p24 ELISA was 99%, with a Kappa index of 0.96. Our results indicate that the national technology-based ELISA r-p24 has sensibility and specificity values that are comparable to rapid ELISA kits. This test is therefore recommendable, using national technology for FIV routine diagnosis, research, and epidemiological studies.