title: Isolation of RNA from residual BD SurePath liquid-based cytology specimens and detection of HPV E6/E7 mRNA using the PreTectt HPV-Proofer assay.
abstract: Infection with high-risk human papillomavirus (HPV) is known to be associated directly with the development of cervical cancer. Recent data suggests that the detection of E6/E7 mRNA from high-risk HPV types may serve as a better diagnostic method for detecting the presence of cervical pre-cancer than HPV DNA testing. This report details a commercially available nucleic acid isolation protocol which can be used to isolate reproducibly RNA from residual BD SurePath liquid-based cytology specimens stored for up to 28 days, and have demonstrated the quality and quantity of mRNA is sufficient for detection with the NorChip PreTect HPV-Proofer assay. Of the 242 specimens tested in this study, 236 (97.5%) tested positive for U1A internal control gene expression. HPV type 16, 18, 31, 33 or 45 mRNA was detected in 16/20 (80%) of the analyzed high-grade squamous intraepithelial lesion (HSIL) specimens, with a low frequency of HPV mRNA detected in the normal lesions (3%). The presence of HPV E6 expression in a subset of HPV positive specimens was also detected by real-time RT-PCR. These findings confirm that RNA of sufficient quality can be isolated from residual BD SurePath cervical cytology specimens for use in downstream NASBA and RT-PCR-based assays.
authors.affiliation: BD Diagnostics - TriPath, 4025 Stirrup Creek Dr., Suite 400, Durham, NC, USA. Eric_Dixon@BD.com
authors.forename: Eric P
authors.initials: EP
authors.lastname: Dixon
authors.forename: Lorraine M
authors.initials: LM
authors.lastname: King
authors.forename: Melissa D
authors.initials: MD
authors.lastname: Adams
authors.forename: Petter
authors.initials: P
authors.lastname: Grønn
authors.forename: Patricia G
authors.initials: PG
authors.lastname: Murphy
authors.forename: Charlotte A
authors.initials: CA
authors.lastname: Brown
authors.forename: George H
authors.initials: GH
authors.lastname: Brough
authors.forename: Hanne
authors.initials: H
authors.lastname: Skomedal
authors.forename: Douglas P
authors.initials: DP
authors.lastname: Malinowski
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Fischer