A RAPID AND SIMPLE METHOD FOR THE DETECTION OF PROSTATE-SPECIFIC ANTIGEN MESSENGER-RNA IN ARCHIVAL TISSUE SPECIMENS USING A REVERSE TRANSCRIPTION-POLYMERASE CHAIN-REACTION ASSAY

Objectives. Polymerase chain reaction (PCR) amplification of DNA from archival, fixed tissue sources is now performed routinely. In this rep ort, we describe a reproducible technique for mRNA amplification from archival tissues. Methods. Archival, fixed tissue was treated with a m odification of a rapid, acid guanidinium technique. The RNA yielded wa s reverse transcribed and subjected to 40 cycles of two-step PCR, usin g a panel of primers designed to encompass relatively short (less than 250 bp) cDNA fragments. PCR products were analyzed by agarose gel ele ctrophoresis followed by ethidium bromide staining. Results. Using fix ed, paraffin-embedded specimens of human prostate, we have demonstrate d a reproducible pattern of reverse transcription (RT)-PCR products us ing several different primer sets. Our data suggest that RNA degradati on proceeds to fragments approximately 250 bp or shorter in length but that many of these fragments survive intact over extended periods of time and are of suitable quality to serve as a template for RT and sub sequent PCR amplification. Conclusions. We describe a technique that w ill allow the retrospective analysis of archival tissues for gene expr ession. These methods are generally applicable to a wide variety of sy stems. Such a method will make it possible to perform retrospective st udies of gene expression in the archival tissues of patients whose eve ntual clinical course is already known, greatly shortening the time ne eded for genetic outcome studies in slowly growing tumors, such as pro state cancer.