Direct microtitre plate enzyme immunoassay of testosterone in unextracted serum

A method for enzyme immunoassay of testosterone in serum has been developed which does not require extraction of the serum with organic solvents. The release of testosterone from the binding proteins is achieved by heating the serum at 70 degrees C for 30 min in an alkaline buffer. The results correlated well (r = 0.96) with those of a radioimmunoassay using 125I-labelled testosterone and with enzyme immunoassay with prior extraction of samples (r = 0.98). The detection limit of the assay is 1 pg per well and the turn around time for 36 samples is 3.5 h. The procedure is simple and well suited for routine analysis.