IMMUNOFLUOROMETRIC ASSAY FOR SENSITIVE AND SPECIFIC MEASUREMENT OF HUMAN PROSTATIC GLANDULAR KALLIKREIN (HK2) IN SERUM

Prostate-specific antigen (PSA) and human prostatic glandular kallikre in (hK2) have 79% identity with the primary structure, When we used re combinant hK2 protein, only 7 of 23 monoclonal anti-PSA IgGs (monoclon al antibodies, MAbs) cross-reacted with hK2, which enabled us to desig n a novel immunofluorometric MAb-MAb assay for the specific detection of hK2, In the first incubation, an excess of MAb 2H11, which does not cross-react with hK2, is added to prevent both free and complexed PSA from reacting in subsequent immunoreactions. In the second incubation , biotinylated MAb H50, which cross-reacts with hK2 by an epitope over lapping with MAb 2H11, served to bind only hK2 to the microtitration w ells coated with streptavidin, In the third step, Eu-labeled MAb H117, which cross-reacts with hK2, detected the immobilized hK2, The hK2 as say was calibrated with recombinant hK2, The detection limit of the as say was 0.1 mu g/L, and the cross-reactivity with recombinant PSA was less than or equal to 0.7%. The concentration of hK2 was measured in s erum samples from 334 males with total PSA concentrations ranging from 1 to 3400 mu g/L, Most of the samples (57%) had hK2 concentrations be low the detection limit, The proportions of hK2 relative to total PSA were 0-2% in 79%, 2-5% in 14%, 5-10% in 4%, and >10% in 3% of the samp les, Gel filtration of 10 serum samples with increased hK2 concentrati ons showed a single peak of hK2 immunoreactivity with an apparent mole cular size of similar to 30 kDa, corresponding to that of recombinant hK2 and free PSA.