DEVELOPMENT OF AN IMMUNOASSAY SPECIFIC FOR THE PSA-ACT COMPLEX WITHOUT THE PROBLEM OF HIGH BACKGROUND

We have developed an assay specific for the PSA-ACT (PSA-alpha 1-antic hymotrypsin) complex that effectively diminishes the problem of high a ssay background commonly reported by other investigators. The assay fo llows a two-site ELISA format. Polyclonal anti-PSA antibodies were coa ted on the microplate to capture the PSA complex from the serum, where as the biotinylated anti-ACT polyclonal antibodies and HRP-conjugated streptavidin were used for detection. The high background ordinarily a ssociated with this assay was greatly reduced when milk casein was add ed in addition to albumin for blocking and when the Super Block(TM) wa s also included in the diluents for sample dilution and dilution of en zyme conjugated detecting antibodies. The assay has a sensitivity of 0 .05 ng/mL. The within-run precision ranges from 4.2-7.2% and the betwe en-run precision falls between 5.8-8.5%. Cross reactions with ACT and free PSA (fPSA) are 0.0001% and 0.02%, respectively. The highest conce ntration of PSA-ACT complex in the maternal sera was < 0.4 ng/mL by th is assay, much less than reported in the literature. Using this improv ed assay, the sum of fPSA and PSA-ACT concentrations were less than th at of their corresponding total PSA (tPSA) most of the time. We believ e that this improved assay should be used to replace the current tPSA assay for screening, monitoring, and managing patients with prostate c ancer. (C) 1998 Wiley-Liss, Inc.