Enzyme-antigen immunoassay for human placental alkaline phosphatase in serum and tissue extracts, and its application as a tumor marker

Abstract

In this enzyme-antigen immunoassay for human placental alkaline phosphatase (hPLAP; EC 3.1.3.1.) in serum and tissue extracts, polyclonal rabbit antiserum to mouse IgG2(b) is absorbed to the wells of a microtiter plate, its excess binding sites are blocked, then it is incubated with murine monoclonal anti-hPLAP and mixed with serially diluted standard or sample antigen. The amount of antigen bound is determined by measuring its enzymic activity. The standard curve is linear for hPLAP concentrations of 0.2 to 1 U/L. The mean within-assay CV was 3.8% (SD 0.9%) for a serum sample and 6.1% (SD 3.0%) for a tissue extract. The respective mean between-assay CVs were: 6.7% (SD 2.0%), and 7.0% (SD 2.0%). Serum hPLAP concentrations, determined in four different dilutions, had a CV of 5.5%. We evaluated the method by standard additions and by comparing dilution curves for purified hPLAP, hPLAP in serum, and hPLAP in tissue extracts. The upper limit of activity in normal subjects was 0.1 U/L for serum samples, and 1.0 mU/g wet weight of tissue for tissue extracts. hPLAP activity was increased in 9.8% of all cancer patients, and in 40% of ovarian cancer patients. Almost half of the tumor biopsies were positive for hPLAP activity, and 94% of the biopsies from ovarian neoplasia had an increased activity of this isoenzyme. Of the nonmalignant tissues examined, normal lung tissue had the highest hPLAP activity.