Development and evaluation of an indirect ELISA based on recombinant structural protein VP2 to detect antibodies
Yalan Lai1, Ling He1, Anchun Cheng1
1Institute of Preventive Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China; Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Wenjiang, Chengdu City, Sichuan 611130, PR China.
Abstract:
An indirect enzyme-linked immunosorbent assay (I-ELISA) based on the VP2 protein of duck hepatitis A virus type 3 (DHAV-3) was established in this study. The optimal dilutions of antigen, serum and goat anti-duck IgG conjugate were 1:1600 (2.23 μg/mL), 1:160 and 1:2000, respectively. The optimal blocking buffer was 1% skim milk. The cut-off value for the method was 0.25, and the analytical sensitivity of the method was 1:5120. The results of specific evaluation showed that except for DHAV-1, DHAV-3 antisera did not cross-react with any other common duck-sensitive pathogens, indicating that this method can be used to detect DHAV-3 and DHAV-1 antibodies. The coefficients of variation (CVs) were lower than 10 %. The coincidence rate between the VP2-DHAV-3-ELISA and the neutralization test was 93.3 %. In summary, the I-ELISA method based on VP2 protein has high sensitivity, specificity, and coincidence rate compared with the neutralization test and has advantages in serum monitoring. The I-ELISA method based on VP2 protein provides a simple and rapid method for the detection of anti-DHAV antibodies and the epidemiological monitoring of DHAV.
