Detection of HPV transcripts by nested RT-PCR

Human papillomaviruses (HPVs) are etiologic for the development of cervical cancer and its precursor lesions, cervical intraepithelial neoplasia (CIN). Nearly all cervical carcinomas (CaCx) harbor HPV DNA, but the presence of HPV alone is not indicative of the future development of neoplasia. While both normal and abnormal smears may harbor HPV DNA, the detection of HPV mRNA is associated, although not exclusively, with abnormal cytology. Recent observations suggest women with a high HPV viral load are at a significantly greater risk for CIN development--particularly those infected with high-risk (HR) HPV types, such as HPV type 16 (HPV 16). Thus, assays capable of detecting HPV transcripts may have useful prognostic value and could be utilized to identify biological markers for progression to high-grade cervical disease. This chapter describes the nested reverse transcriptase (nRT)-polymerase chain reaction (PCR) methods developed in our laboratory for the detection of the majority of all early region HPV-16 transcripts.