Application of real-time reverse transcriptase-polymerase chain reaction in urological oncology.
OBJECTIVE
During the last decade numerous different reverse transcriptase-polymerase chain reaction (RT-PCR) techniques have been described. However, the lack of highly sensitive, quantitative and reliable methodology has been responsible for its limited use in modern urology. Early semiquantitative RT-PCR techniques often proved not to produce consistent results and have a high failure rate due to complicated working models. In this article we provide a comprehensive and intelligible description of real-time PCR technology, which is a novel quantitative methodology to analyze gene expression. In addition, we report the first preclinical and clinical applications in molecular urology.


METHODS
The current literature was reviewed in regard to different current real-time RT-PCR protocols and their use in modern urological oncology.


RESULTS
Real-time RT-PCR is a reliable, rapid and relatively inexpensive technique that can be easily adapted for standardized preclinical and clinical applications at different centers. Its sensitivity equals at least that of conventional RT-PCR and the option of exact quantification of gene expressions allows proper differentiation among high, low and illegitimate RNA transcription. It eliminates post-PCR processing of PCR products, thereby, increasing throughput and decreasing the chance of carryover contamination.


CONCLUSIONS
Although the application of real-time RT-PCR has gained wide acceptance in urological research, its routine clinical use is still in its infancy. However, due to its high sensitivity and exact quantitation real-time RT-PCR may be the method of choice for modern preclinical and clinical studies in the future.
Schrader
Andres Jan
AJ
Department of Urology, Philipps-University of Marburg, Marburg, Germany.
Lauber
Joerg
J
Lechner
Oskar
O
Heidenreich
Axel
A
Hofmann
Rainer
R
Buer
Jan
J
D006801:Humans; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013997:Time Factors; D014571:Urologic Neoplasms / Q000235:genetics*