Abstract
Murine monoclonal antibodies (MAbs) of the immunoglobulin classes IgM and IgA, from two different hybridoma cell lines, and rabbit (polyclonal) antiserum raised against mycelial antigens from an AG4 isolate of R. solani were used to develop magnetic microsphere-enzyme immunoassays (MM-EIAs) for the detection and quantification of the pathogen in soil. The assay can also be used to differentiate R. solani anastomosis group (AG) 2-2 isolates from other AG isolates grown in vitro. Mycelial antigens were solubilised in saline buffer containing detergent and following high speed centrifugation soluble antigen was delineated by the addition either of a mixture of specific murine MAbs of different immunoglobulin classes or a mixture of murine MAb supernatant and rabbit antiserum. Due to the low concentration of reactants a soluble immune complex was formed. The soluble complex was isolated by the addition of magnetic beads coated with antibodies that specifically recognised the murine IgM MAbs. The bound antibody-antigen complex was then detected using a commercial antibody-enzyme conjugate and chromogen.
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