title: Sensitivity of in situ detection with biotinylated probes of human papilloma virus type 16 DNA in frozen tissue sections of squamous cell carcinomas of the cervix.
abstract: The sensitivity of human papilloma virus type 16 (HPV-16) DNA detection by DNA in situ hybridization using biotinylated probes (bio-DISH) was estimated by performing this technique on snap-frozen tissue sections of 10 cervical squamous cell carcinomas containing increasing amounts of HPV-16 as determined by Southern blot hybridization. A protocol using serial sections for bio-DISH and DNA extraction was used. The number of positively stained cells and the detection limit were strongly dependent on the treatment of the sections with proteinase K prior to hybridization. At low proteinase K concentration (0.1 micrograms/ml), the detection limit appeared to be 30-40 HPV-16 DNA copies per carcinoma cell, whereas morphology was preserved. A high proteinase K concentration (1-5 micrograms/ml) often resulted in an increase in the number of positively stained cells but also in a poor morphology. The detection limit was improved to at least 20 HPV-16 DNA copies per carcinoma cell.
authors.affiliation: Department of Pathology, Free University Hospital, Amsterdam, The Netherlands.
authors.forename: J M
authors.initials: JM
authors.lastname: Walboomers
authors.forename: W J
authors.initials: WJ
authors.lastname: Melchers
authors.forename: H
authors.initials: H
authors.lastname: Mullink
authors.forename: C J
authors.initials: CJ
authors.lastname: Meijer
authors.forename: A
authors.initials: A
authors.lastname: Struyk
authors.forename: W G
authors.initials: WG
authors.lastname: Quint
authors.forename: J
authors.initials: J
authors.lastname: van der Noordaa
authors.forename: J
authors.initials: J
authors.lastname: ter Schegget
mesh_terms: D001710:Biotin; D002294:Carcinoma, Squamous Cell / Q000032:analysis*; D004279:DNA, Viral / Q000032:analysis*; D005260:Female; D005615:Freezing; D006652:Histological Techniques; D006801:Humans; D009693:Nucleic Acid Hybridization; D027383:Papillomaviridae / Q000235:genetics*; D002583:Uterine Cervical Neoplasms / Q000032:analysis*
chemical_list: D004279:DNA, Viral; D001710:Biotin