Evaluation of two commercialised in situ hybridisation assays for detecting HPV-DNA in formalin-fixed, paraffin-embedded tissue.
BACKGROUND
The role of human papilloma virus (HPV) in the pathogenesis of anogenital dysplasia is now conclusive. However, HPV detection in formalin-fixed and paraffin-embedded tissues remains controversial. Therefore, the aim of this study was to evaluate morphological changes directly in tissue specimens using a HPV-DNA detection system involving HPV in situ hybridisation.


METHODS
Samples from patients with cervical carcinoma were analysed using the GenPoint HPV DNA Probe Cocktail (Dako, Glostrup, Denmark) and the ZytoFast HPV Screening CISH-Kit (Zytomed, Berlin, Germany). Three cervical carcinoma cell lines with a well-defined HPV copy number per cell (SiHa, HeLa, and CaSki) served as positive controls for sensitivity testing, while two HPV-negative cell lines (AC-1M32, MCF-7) and brain tissue samples served as negative controls. Moreover, to assess the validity of the in situ hybridisation, the expression of HPV-16 DNA in cell lines was demonstrated by HPV-16 E6-specific PCR.


RESULTS
Both HPV-screening assays revealed strong signals of episomal and integrated HPV-DNA at a HPV copy number of more than 50 copies/cell. All cervical carcinoma samples were positive in the Dako assay, which identifies 13 high-risk HPV genotypes, whereas HPV-DNA could be detected in 9/10 cervical carcinoma samples using the Zytofast assay, identifying HPV 16, 18, 31, 33, and 35.


CONCLUSIONS
HPV in situ hybridisation is a convenient and powerful tool for detecting HPV-DNA in formalin-fixed and paraffin-embedded tissue samples. Therefore, this technique is suitable for analysis of a potential HPV infection using archival pathological slides.
First Department of Obstetrics and Gynaecology, Ludwig-Maximilians-University, Maistrasse 11, 80337 Munich, Germany.
Maresa
M
Montag
Thomas J F
TJ
Blankenstein
Naim
N
Shabani
Ansgar
A
Brüning
Ioannis
I
Mylonas
D045744:Cell Line, Tumor; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D052162:Human papillomavirus 16 / Q000235:genetics; D006801:Humans; D017403:In Situ Hybridization / Q000592:standards*; D027383:Papillomaviridae / Q000235:genetics*; D030361:Papillomavirus Infections / Q000473:pathology / Q000821:virology*; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D011237:Predictive Value of Tests; D012680:Sensitivity and Specificity; D002583:Uterine Cervical Neoplasms / Q000473:pathology / Q000821:virology*
D017931:DNA Primers; D004279:DNA, Viral