Absolute quantitative real-time polymerase chain reaction for the measurement of human papillomavirus E7 mRNA in cervical cytobrush specimens.
BACKGROUND
Few reports of the utilization of an accurate, cost-effective means for measuring HPV oncogene transcripts have been published. Several papers have reported the use of relative quantitation or more expensive Taqman methods. Here, we report a method of absolute quantitative real-time PCR utilizing SYBR-green fluorescence for the measurement of HPV E7 expression in cervical cytobrush specimens.


RESULTS
The construction of a standard curve based on the serial dilution of an E7-containing plasmid was the key for being able to accurately compare measurements between cervical samples. The assay was highly reproducible with an overall coefficient of variation of 10.4%.


CONCLUSIONS
The use of highly reproducible and accurate SYBR-based real-time polymerase chain reaction (PCR) assays instead of performing Taqman-type assays allows low-cost, high-throughput analysis of viral mRNA expression. The development of such assays will help in refining the current screening programs for HPV-related carcinomas.
Scheurer
Michael E
ME
The University of Texas M. D. Anderson Cancer Center, Department of Epidemiology, Unit 1340, PO Box 301439, Houston, TX 77230-1439, USA. mescheur@mdanderson.org
Dillon
Laura M
LM
Chen
Zhuo
Z
Follen
Michele
M
Adler-Storthz
Karen
K