NONRADIOISOTOPIC DETECTION AND TYPING OF HUMAN PAPILLOMAVIRUSES BY USE OF POLYMERASE CHAIN-REACTION AND SINGLE-STRAND CONFORMATION POLYMORPHISM

The polymerase chain reaction (PCR), used to detect human papillomavir us (HPV), is finding increasing applications in clinical laboratories. The standard method of analysis to detect amplified PCR products is e thidium bromide eel electrophoresis combined with labor-intensive blot hybridization. In this study, we describe single-strand conformation polymorphism (SSCP) to detect and genotype simultaneously general prim er GP5+/GP6+ amplified HPV DNA using semiautomated electrophoresis on polyacrylamide gels (PAGE) combined with sensitive silver staining. To establish a standard for the band patterns of the various HPV types, we used HPV plasmid DNA, which allowed us to distinguish HPV 6, 11, 16 , 18, 31, 33, 35, 45, 51, 52, 56, and 58, covering the most frequently recognized types. All the types tested are separated from each other, demonstrating diverse band patterns, HPV 16 being the most distinct. We also investigated PCR-SSCP for HPV detection and typing of 86 cervi cal biopsies diagnosed as cervical intraepithelial neoplasia (GIN) I-I II and known to be HPV positive by PCR-slot blot hybridization and in situ hybridization. The correlation with SSCP was 91% for in situ hybr idization and 98% for PCR-slot blot hybridization. SSCP is reproducibl e and specific, Its sensitivity is comparable to slot-blot hybridizati on, The interval to SSCP is approximately 2 h after PCR compared with several days' work when using conventional blot hybridization. We conc luded that SSCP may be more advantageous than other PCR-based typing t echnologies.