A Biotin-Streptavidin Enzyme Immunoassay for Detection of Antibodies to Porcine Granulosa Cell Antigens

Abstract A colorimetric solid-phase enzyme immunoassay has been developed which quantifies antibodies to porcine granulosa cell membrane antigens in rabbits immunized with porcine granulosa cells. A cell-free, particulate membrane preparation of porcine granulosa cells was used as coating antigen. A biotinylated second antibody in conjunction with a streptavidin-β-galactosidase conjugate was utilized to amplify reactivity. The enzyme β-galactosidase was used due to high background obtained using peroxidase, presumably due to endogenous peroxidase activity of the tissue. Sigmoidal serum dilution curves were obtained with immune rabbit sera indicating that absorbance was related to the concentration of antibodies. Assay activity was reduced by preincubation of immune serum with granulosa cell membranes. Sera from ovariectomized or pre-immune rabbits did not yield any specific binding in the assay. This assay has potential applicability for quantifying antiovarian and anti-granulosa cell antibodies in women ...