Microball Double Antibody Sandwich Elisa for the Detection of Newcastle Disease Virus Antigen in Chickens

A mieroball double antibody sandwich ELISA (MDS-ELISA) was developed to detect Newcastle disease virus (NDV) antigen in the tissues of chickens infected with NDV. In the assay, a monoclonal antibody (specific for NDV.IgM isotype), and its conjugate of horse radish peroxidase (FN-HRP) were served as the capture antibody and the indicator antibody, respectively. When FN was at 400g/ml, the detectable antigen amount was as low as 3g NDV protein per millilitre.The results of detection of NDV antigen from NDV-infected chickens by the MDS-ELISA and the direct immunof luorescent-antibody test (DFA) showed that the sensitivity of the MDS-ELISA was higher than of the DFA. All samples that were positive inthe DFA were also positive in the MDS-ELlSA. However, the MDS-ELISA indicated that a small number of samples were NDV antigen positive which were negative by the DFA. The concordance rate between tese two methods was 92%.The MDS-ELISA was evaluated on detection of the samples from chickens inoculated with various NDV strains. The experiment demonstrated that: 1) the positive rates in the various organs from the chickens infected with F48 strain of NDV were 100% in a mixture of lung, kidney and ileocecal tonsil, and in kidney. 90% in lung, 80% in ileo-cecal tonsil, liver and spleen, respectively; 2) only some positive samples were found in Mukteswar-inoculated chickens in 6 days post-injection; 3) no positive samples had been found in Bl strain infected chickens.The MDS-ELISA was proved to be a most sensitive, specific, rapid and convenient method. It would be most useful for diagnosis of ND in poultry farms.