A comparison of biotin and isotope-labeled ribonucleic acid probes for in situ detection of HPV-16 ribonucleic acid in genital precancers

Genital precancers were analyzed by in situ hybridization for the presence of HPV RNA using single-stranded RNA probes. To compare the sensitivity of biotin- and 35S-labeled probes, serial sections from three biopsies containing variable amounts of HPV 16 RNA were analyzed with probes containing each label. The probe template was a cloned fragment of the HPV 16 genome spanning portions of the E2, E5 and L2 orf's. When a focus contained a strong hybridization signal with 35S-labeled probes (i.e., target/background ratio greater than 40) serial sections of the same area usually demonstrated a strongly positive signal with biotin-labeled probes. When the signal with 35S in a given focus was weak (target/background ratio less than 20), serial sections contained either a very weak or no detectable signal with the biotinylated probe. From this study it appears than 35S-labeled RNA probes exceed the sensitivity of biotin-labeled probes by nearly an order of magnitude with short exposures (4 days), and with longer exposures (1 to 2 weeks), approach the sensitivity of that reported for tritiated probes and exposures of 1 month or longer.