Indirect ELISA for Detecting Specific IgA antibody to Porcine Epidemic Diarrhea Virus
Abstract
The object of this study is to establish an indirect ELISA method for detecting IgA antibody against porcine epidemic diarrhea virus (PEDV) and provide a technological means to PEDV infection detection and immune effect evaluation. Antigen coating concentration, blocking and dilution buffer, optimal dilutions of test serum and enzyme-labeled antibody were determined and the indirect ELISA method for detecting PEDV-IgA antibody was developed. Furthermore, the specificity, intra- and inter-batch repeatability tests of the ELISA were examined, the coincidence rate were measured between the ELISA and the existing ELISA kit, and the correlation between the specific IgA and the neutralizing antibodies in serum was analyzed. The results showed that the optimal antigen coating concentration of the ELISA was 1μg/mL, the blocking and dilution buffers were PBS containing 5% calf serum and 0.05% Tween-20, and the working concentrations of tested serum and enzyme-labeled antibody were 1:40 and 1:1500, respectively. The method could be used to detect the PEDV specific antibodies without cross reactions with those antibodies against porcine reproductive and respiratory syndrome virus, classical swine fever virus, pseudorabies virus, and porcine circovirus type 2 using the ELISA.The coefficients of variation (CV) of intra- and inter-batch repetitive tests of the ELISA were less than 10%. The coincidence rate was 94.8% when the same serum samples were detected using the ELISA and existing ELISA kit for detecting PEDV antibody. The level of specific IgA antibody was positively correlated with that of the neutralizing antibody in serum (r=0.69, p<0.001).
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DOI: https://doi.org/10.18686/pas.v2i1.1314
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