Abstract
Human papillomavirus (HPV) is an essential cofactor for cancer at many sites, including the genital tract, oral cavity, conjunctiva, and periungual region. The in situ detection of HPV allows us to determine the cellular targets of the virus. In situ-based coexpression analyses of HPV with putative target proteins provide tremendous insight into the molecular evolution of these viral-associated cancers. HPV DNA is present in high copy number in the precancerous lesions and is, thus, readily detected by in situ hybridization. However, viral integration, typical during oncogenesis, is associated with reduced copy number of the virus, necessitating in situ polymerase chain reaction amplification for accurate in situ detection of HPV. This chapter provides the protocols that can be used to detect HPV DNA in situ as well as to correlate viral DNA with coexpression of relevant protein targets.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsSpringer Nature is developing a new tool to find and evaluate Protocols. Learn more
References
Nuovo, G.J. (1992) PCR in situ hybridization. Protocols and applications. Raven Press, New York, NY.
Nuovo, G.J. (2002) Diagnosis of human papillomavirus using in situ hybridization and in situ polymerase chain reaction. Methods Mol Biol 179:113–136.
Bagasra, O., and Harris, T. (2006) Latest developments in in situ PCR. Methods Mol Biol 334:221–240.
Morrison, C., Catania, F., Wakely, P., and Nuovo, G.J. (2001) Highly keratinizing squamous cell cancer of the cervix: a rare, locally aggressive tumor not associated with human papillomavirus or squamous intraepithelial lesions. Am J Surg Pathol 25:1310–1315.
Nuovo, G.J., Gallery, F., MacConnell, P., Becker, J., and Bloch, W. (1991) An improved technique for the detection of DNA by in situ hybridization after PCR-amplification. Am J Pathol 139:1239–1244.
Nuovo, G.J., Nana-Sinkam, P., Elton, T., Croce, C., Volinia, S., and Schmittgen, T.S. (2009) A methodology for the combined in situ analyses of the precursor and mature forms of microRNAs and correlation with their putative targets. Nat Protoc 4:107–115.
Greer, C.E., Peterson, S.L., Kiviat, N.B., and Manos, M.M. (1991) PCR amplification from paraffin-embedded tissues: effects of fixative and fixative times. Am J Clin Pathol 95:117–124.
Nuovo, G.J., and Silverstein, S.J. (1988) Comparison of formalin, buffered formalin, and Bouin’s fixation on the detection of human papillomavirus DNA extracted from genital lesions. Lab Invest 59:720–724.
Zaravinos, A., Mammas, I.N., Sourvinos, G., and Spandidos, D.A. (2009) Molecular detection methods of human papillomavirus (HPV). Int J Biol Markers 24(4):215–222.
Nuovo, G.J. (2008) In situ detection of precursor and mature microRNAs in paraffin embedded, formalin fixed tissues and cell preparations. Methods 44(1):39–46.
Godlewski, J., Nowicki, M.O., Bronisz, A., Nuovo, G., Palatini, J., De Lay, M., Van Brocklyn, J., Ostrowski, M.C., Chiocca, E.A., and Lawler, S.E. (2010) MicroRNA-451 regulates LKB1/AMPK signaling and allows adaptation to metabolic stress in glioma cells. Mol Cell 37(5):620–632.
Nuovo, G.J., Hochman, H., Eliezri, Y.D., Comite, S., Lastarria, D., and Silvers, D. (1990) Human papillomavirus DNA in penile lesions histologically negative for condylomata: analysis by in situ hybridization and the polymerase chain reaction. Am J Surg Pathol 14:829–836.
Nuovo, G.J., Becker, J., MacConnell, P., Margiotta, M., Comite, S., and Hochman, H. (1992) Histological distribution of PCR-amplified HPV 6 and 11 DNA in penile lesions. Am J Surg Pathol 16:269–275.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2011 Humana Press
About this protocol
Cite this protocol
Nuovo, G.J. (2011). In Situ Detection of Human Papillomavirus DNA After PCR-Amplification. In: Theophilus, B., Rapley, R. (eds) PCR Mutation Detection Protocols. Methods in Molecular Biology, vol 688. Humana Press. https://doi.org/10.1007/978-1-60761-947-5_4
Download citation
DOI: https://doi.org/10.1007/978-1-60761-947-5_4
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-60761-946-8
Online ISBN: 978-1-60761-947-5
eBook Packages: Springer Protocols