Monoclonal antibodies against bovine immunoglobulins and their use in isotype-specific ELISAs for rotavirus antibody

Van Zaane, Dick; Ijzerman, Johan

Monoclonal antibodies (MCA) against bovine immunoglobulin (BIg) isotypes were produced and characterized. MCAs were obtained which react specifically with IgG, IgG(1), IgG(2) or IgA while MCAs against IgM showed a partial cross-reaction with affinity purified IgA. MCAs with optimal characteristics for application in ELISA were selected and used as conjugates in an indirect double antibody sandwich assay (IDAS) and as the capturing antibody in an antibody capture assay (ACA) for the isotype-specific detection of antibodies against rotavirus. Based on theoretical grounds, experimental analysis of inter- and intra-isotype competition in IDAS and ACA, respectively, and direct comparison of both tests, the IDAS was selected for the detection of IgG(1) and IgG(2) anti-rotavirus antibodies. The ACA was the test of choice for the detection of IgM and IgA anti-rotavirus antibodies. The isotype specificity of these tests relies on the specificity of the MCAs and was confirmed for each test by the observation that samples containing rotavirus antibodies of only 1 particular isotypee reacted only in the homologous assay. The MCAs against bovine Ig isotypes and isotype-specific ELISAs were found to be very useful in the study of humoral mucosal immunity in calves infected with rotavirus.