Development of a highly sensitive direct microtiter plate ELISA for human serum testosterone.

A simple, highly sensitive, direct, competitive ELISA for human serum testosterone has been indigenously developed. Specific antisera against testosterone were raised in rabbits using testosterone-3carboxymethyl oxime (CMO)-bovine serum albumin (BSA) as the antigen. For the enzyme conjugate, testoterone-3CMO was coupled with horse raddish peroxidase by the active ester method. The standard curve covered a wide range from 3.9 pg/ml to 500 pg/ml. The inter and intra-assay variation were found to be low and within the acceptable limits. Specificity and accuracy for the assay was established by having negligible crossreactivity with the related steroids and an excellent parallelism between the sample and standard dilution curve. Samples measured by RIA and ELISA showed very high degree of correlation (r=0.991).