关键词: 鮰爱德华菌;检测;间接ELISA;竞争ELISA

Abstract:

Two methods including indirect ELISA and competitive ELISA were developed to rapidly detect a pathogen of Edwardsiella ictaluri in the early stage. A monoclonal antibody 5D11 was used as the detection antibody. In indirect ELISA when antigen was coaed at 37℃ overnight, the plate had good adsorption effect on bacteria. The optimal dilution of mAb and AP-goat-anti-mouse IgG were 1∶80 and 1∶1 000, respectively. The lowest concentration of E.ictaluri that could be detected was 5×106 cells/mL. Cross-reactivity test proved that the method had strong specificity. In competitive ELISA, the optimum working combination of coating antigen-mAb was 5×108~1∶80. The proportion of mAb and competitive antigen was 3∶7. A standard curve with R2=0.990 1 was established. This method had strong specificity, and the detection limit was 106 cells/mL. This method can qualitatively and quantitatively detect E.ictaluri within limits, which makes up the deficiency of indirect ELISA.

Key words: Edwardsiella ictaluri, detection, indirect ELISA, competitive ELISA