Thermally reversible hydrogel based fluoroimmunoassay of methyltestosterone

A novel immunoassay of methyltestosterone (MT) in serum is developed. We sy nthesized the antigen of MT (Ag-MT) by covalently bonding it to BSA and rai sed its antibody (Ab(MT)) from rabbit. Ab(MT) was conjugated to poly-Nisopr opylacrylamide (p-NIPAAm) to form Ab(MT) bound thermally reversible hydroge l (p-NIPAAm-Ab(MT)). A competitive immunoassay method based on the competit ion of fluorescein isothiocyanate (FITC) labeled MT antigen (FITC-Ag-MT) an d MT with Limited amounts of p-NIPAAm-Ab(MT) was established. The separatio n in the assay process was achieved by precipitation of the immunocomplex a bove its critical solution temperature. The detection limit of MT is 50 ng/ ml. The recoveries of MT in human serum are above 93%.