|Budget Amount *help
¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1999 : ¥1,300,000 (Direct Cost : ¥1,300,000)
Fiscal Year 1998 : ¥1,800,000 (Direct Cost : ¥1,800,000)
Objectives. To develop a rapid, sensitive, reverse transcriptase-polymerase chain reaction (RT-PCR) prostate-specific antigen (PSA) messenger ribonucleic acid (mRNA) detection method by applying colorimetric enzyme-linked immunosorbent assay (ELISA).
Methods. Total RNA was extracted form 16 urogenital cancer cells (including PSA-producing LNCaP cells) from pelvic and inguinal lymph node aspiration biopsy samples from patients with prostate, bladder, and penile cancer, as well as from blood samples of 500 patients with urogenital cancer. We used rTth polymerase for RT and PCR. The RNA target was amplified by RT-PCR with dinitrophenyl-labeled primer. The PCR product was denatured and hybridized on a PSA-specific probe-coated microwell plate.
Results. In 16 cancer cell lines, only LNCaP cells expressed especially high PSA mRNA values, with an optical density (OD) greater than 3. In other cell lines, two testicular cells had relatively high ODs, 1.909 and 0.987, respectively. A high PSA mRNA value was obtained by fine needle aspiration from pelvic lymph node specimens of cytologically positive lymph nodes from patients with prostate cancer but not from patients with cytologically proved bladder or penile cancer. Sensitivity and specificity of fine needle aspiration samples were 70% and 100%, respectively. Blood tests obtained from patients with prostate cancer demonstrated high PSA mRNA values.
Conclusions. The PSA mRNA RT-PCR ELISA method provides a sensitive photometric enzyme immunoassay for the detection of PSA mRNA, using nonradioactive techniques. UROLOGY 53 : 228-235, 1999. (c) 1999, Elsevier Science Inc. All rights reserved.