Nucleic acid sandwich hybridization in detection of HPV 16 DNA: technique and its clinical application

Nucleic acid sandwich hybridization technique was used for detection of HPV 16 specific DNA in cervical scrapes. Alternating HPV 16-PstI fragments were cloned into plasmid pBR322 and phage M13mp10. pBR322-clones were used as 32P-labelled probe reagents and the M13mp10 clones as catching reagents in the assay. The detection limit of the test proved to be 1-5 X 10(5) HPV 16 molecules per test. A weak cross reaction was seen with HPV 31 DNA but not with the other types tested, e.g. HPV 6, 11, 18 and 33. Cervical scrapings obtained from 163 consecutive patients included in a prospective follow up study were analyzed for HPV 16 DNA with the sandwich hybridization method, dot-blot hybridization being used as a reference method. Sandwich hybridization assay detected 25 positive cases out of 163 specimens (15.3%). Of these 6 and 3 additional specimens were positive in dot-blot hybridization assay. HPV 16 DNA was related to higher PAP grades, and HPV 16 appeared more frequently in HPV CIN than HPV NCIN lesions. None of the infections caused by HPV 16 regressed, and 24% progressed during the follow up.