DETECTION OF HUMAN PAPILLOMAVIRUS DNA BY IN-SITU HYBRIDIZATION AND PCR

In situ hybridization with non radioactive probes is attractive for human papillomaviruses (HPV) detection. Its sensitivity has been greatly improved by using different hybridization conditions, techniques for revealing the DNA-DNA hybrids and method of observation and various cell lines derived from human uterine carcinomas (CaSki, SiHa and HeLa cells) which contain 500-600 copies of HPV DNA, 1-2 copies of HPV 16 and 20-50 copies of HPV DNA 18, respectively. In situ gene amplification increased the detection of HPV DNA since hybridization spots were visible in SiHa cells on slides; a specific signal was observed in HeLa cells in suspensions examined by flow cytometry. Confocal microscopy is an alternative method to in situ gene amplification since viral DNA is detectable in SiHa cells with or without gene amplification. Thus, the techniques used in this study are potentially useful for research of single cellular genes.