Production and characterization of novel anti-prostate-specific antigen (PSA) monoclonal antibodies that do not detect internally cleaved Lys145-Lys146 inactive PSA

Background: The nature of free, uncomplexed prostate-specific antigen (PSA) in the circulation is still unknown. In this study, we developed novel ant i-PSA antibodies using PSA produced by a metastasized cancer cell line, LNC aP, as an immunogen, Methods: Hybridoma cell lines were screened with different methods that aim ed at finding antibodies specific for the forms of free PSA produced by LNC aP cell line. Obtained antibodies were further studied for their characteri stics related to previously characterized monoclonal antibodies. Results: Numerous anti-PSA antibodies were obtained, of which four represen ted unique epitopes previously unrecognized by us. One free-PSA-specific an tibody was hound to PSA on two distinct epitopes, and one antibody was boun d to the carboxyl-terminal peptide of PSA. Two antibodies were found to bin d to the peptide sequence adjacent to the internal cleavage site Lys145-Lys 146. These antibodies failed to recognize internally cleaved PSA at Lys145- Lys146. We could not find antiproPSA antibodies despite the fact that LNCaP PSA contained more than one-half of the zymogen form of PSA. Conclusions: We report; for the first time, novel anti-PSA antibodies that do not recognize internally cleaved PSA at Lys145-Lys146 and thus are speci fic for intact, unclipped PSA. (C) 2000 American Association for Clinical C hemistry.