Assessment of CD4+ and CD8+ IFN-gamma producing cells by ELISPOT in naïve and FIV-infected cats

IFN-γ is critical for the development of antiviral cell-mediated immunity in HIV infected humans and FIV infected cats. The ELISPOT has proven to be a technically straightforward assay to quantify the number of IFN-γ producing cells and offers a reasonable alternative for the quantitative measurement of T-cell function in cats. We used a feline-specific ELISPOT to identify constitutive as well as Con A stimulated IFN-γ production in T-cell subsets and determine if there were differences between purified (positively sorted) and negatively depleted populations from naïve and FIV infected cats. We found no difference in the total number of PBMC constitutively producing IFN-γ in naïve and FIV+ cats. Con A exposure was associated with increased numbers of IFN-γ producing PBMC in naïve, but not FIV+, cats. Equivalent numbers of CD4+ and CD8+ T cells constitutively expressed IFN-γ in naïve cats. However, in FIV+ cats, the number of IFN-γ producing CD8+ T-cells was approximately two-fold over that seen for CD4+ T-cells. We found minimal differences between purified (e.g. CD4+ or CD8+) and corresponding depleted (e.g. CD8− or CD4−) populations in samples from FIV+ cats. In contrast, depleted populations from naïve cats showed greater response to Con A than did purified populations. Thus, while determination of the number of IFN-γ producing cells by feline-specific ELISPOT is a useful tool for the evaluation of the feline immune response, determination of the initial sample population and T-cell subset is critical for optimal interpretation of the IFN-γ ELISPOT.