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. 1991 Oct;139(4):847–854.

Detection of human papillomavirus DNA in formalin-fixed tissues by in situ hybridization after amplification by polymerase chain reaction.

G J Nuovo 1, P MacConnell 1, A Forde 1, P Delvenne 1
PMCID: PMC1886300  PMID: 1656763

Abstract

The authors describe the detection of human papillomavirus (HPV) 16 DNA in paraffin-embedded, formalin-fixed tissues of cervical squamous intraepithelial lesions (SILs) by in situ hybridization after amplification by the polymerase chain reaction (PCR). Using conventional in situ hybridization and a biotin-labeled probe, variable numbers of superficial cells and none of the basal cells in the SILs showed detectable HPV 16 DNA. When the in situ assay was done after amplification, increased numbers of superficial cells had detectable HPV DNA, and the hybridization signal was much more intense. HPV DNA was also detected in basal and parabasal cells at the site of the lesion whereas not detectable in directly adjacent, normal squamous epithelium. Amplified HPV DNA was demonstrated in formalin-fixed SiHa cells using a biotin-labeled probe, demonstrating the ability to detect one copy of HPV 16 DNA. This technique should allow for direct visualization in cells of other DNA sequences of low copy number from achival specimens otherwise undetectable by conventional in situ hybridization analysis.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Crum C. P., Nuovo G., Friedman D., Silverstein S. J. A comparison of biotin and isotope-labeled ribonucleic acid probes for in situ detection of HPV-16 ribonucleic acid in genital precancers. Lab Invest. 1988 Mar;58(3):354–359. [PubMed] [Google Scholar]
  2. Crum C. P., Nuovo G., Friedman D., Silverstein S. J. Accumulation of RNA homologous to human papillomavirus type 16 open reading frames in genital precancers. J Virol. 1988 Jan;62(1):84–90. doi: 10.1128/jvi.62.1.84-90.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Crum C. P., Roche J. K. Molecular pathology of the lower female genital tract. The papillomavirus model. Am J Surg Pathol. 1990;14 (Suppl 1):26–33. [PubMed] [Google Scholar]
  4. Haase A. T., Retzel E. F., Staskus K. A. Amplification and detection of lentiviral DNA inside cells. Proc Natl Acad Sci U S A. 1990 Jul;87(13):4971–4975. doi: 10.1073/pnas.87.13.4971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Hurlin P. J., Kaur P., Smith P. P., Perez-Reyes N., Blanton R. A., McDougall J. K. Progression of human papillomavirus type 18-immortalized human keratinocytes to a malignant phenotype. Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):570–574. doi: 10.1073/pnas.88.2.570. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Lorincz A. T., Temple G. F., Kurman R. J., Jenson A. B., Lancaster W. D. Oncogenic association of specific human papillomavirus types with cervical neoplasia. J Natl Cancer Inst. 1987 Oct;79(4):671–677. [PubMed] [Google Scholar]
  7. McCance D. J., Kopan R., Fuchs E., Laimins L. A. Human papillomavirus type 16 alters human epithelial cell differentiation in vitro. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7169–7173. doi: 10.1073/pnas.85.19.7169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Nuovo G. J., Darfler M. M., Impraim C. C., Bromley S. E. Occurrence of multiple types of human papillomavirus in genital tract lesions. Analysis by in situ hybridization and the polymerase chain reaction. Am J Pathol. 1991 Jan;138(1):53–58. [PMC free article] [PubMed] [Google Scholar]
  9. Nuovo G. J., Friedman D., Richart R. M. In situ hybridization analysis of human papillomavirus DNA segregation patterns in lesions of the female genital tract. Gynecol Oncol. 1990 Feb;36(2):256–262. doi: 10.1016/0090-8258(90)90184-m. [DOI] [PubMed] [Google Scholar]
  10. Nuovo G. J., Richart R. M. A comparison of biotin- and 35S-based in situ hybridization methodologies for detection of human papillomavirus DNA. Lab Invest. 1989 Oct;61(4):471–476. [PubMed] [Google Scholar]
  11. Nuovo G. J., Richart R. M. A comparison of slot blot, southern blot, and in situ hybridization analyses for human papillomavirus DNA in genital tract lesions. Obstet Gynecol. 1989 Oct;74(4):673–678. [PubMed] [Google Scholar]
  12. Nuovo G. J., Richart R. M. Buffered formalin is the superior fixative for the detection of HPV DNA by in situ hybridization analysis. Am J Pathol. 1989 Apr;134(4):837–842. [PMC free article] [PubMed] [Google Scholar]
  13. Pecoraro G., Lee M., Morgan D., Defendi V. Evolution of in vitro transformation and tumorigenesis of HPV16 and HPV18 immortalized primary cervical epithelial cells. Am J Pathol. 1991 Jan;138(1):1–8. [PMC free article] [PubMed] [Google Scholar]
  14. Reid R., Greenberg M., Jenson A. B., Husain M., Willett J., Daoud Y., Temple G., Stanhope C. R., Sherman A. I., Phibbs G. D. Sexually transmitted papillomaviral infections. I. The anatomic distribution and pathologic grade of neoplastic lesions associated with different viral types. Am J Obstet Gynecol. 1987 Jan;156(1):212–222. doi: 10.1016/0002-9378(87)90241-9. [DOI] [PubMed] [Google Scholar]
  15. Rozenblatt S., Eizenberg O., Ben-Levy R., Lavie V., Bellini W. J. Sequence homology within the morbilliviruses. J Virol. 1985 Feb;53(2):684–690. doi: 10.1128/jvi.53.2.684-690.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Shibata D. K., Arnheim N., Martin W. J. Detection of human papilloma virus in paraffin-embedded tissue using the polymerase chain reaction. J Exp Med. 1988 Jan 1;167(1):225–230. doi: 10.1084/jem.167.1.225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Walboomers J. M., Melchers W. J., Mullink H., Meijer C. J., Struyk A., Quint W. G., van der Noordaa J., ter Schegget J. 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. Am J Pathol. 1988 Jun;131(3):587–594. [PMC free article] [PubMed] [Google Scholar]

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