Detection and Typing of Papillomavirus DNA in Formaldehyde-Fixed Paraffin-Embedded Tissue

Abstract

Clinical specimens from nine patients with papillomatosis of the vocal cords and three patients with vocal cord polyps were evaluated for the presence of human papillomavirus (HPV) DNA using two complementary molecular hybridization techniques. In one method, involving polymerase chain reaction (PCR) amplification, HPV DNA sequences were replicated in vitro from tissue DNA extracted from paraffin sections prior to hybridization. Polymerase chain reaction amplification was compared with the standard method of Southern blot hybridization. Results of the two techniques for all nine laryngeal papillomas agreed completely: five patients harbored HPV type 6 and four HPV type 11. Both PCR amplification and Southern blot hybridization found two of the three polyps to be free of HPV infection, while PCR detected HPV type 18 in one polyp specimen that was reported negative by Southern blot hybridization, suggesting a greater sensitivity of PCR. Our results demonstrate that PCR amplification is as reliable and at least as sensitive as Southern blot hybridization. Moreover the PCR technique opens the way to the undertaking of a whole variety of retrospective studies using formaldehyde-fixed paraffin-embedded tissues.

Rising Incidence of Oropharyngeal Cancer and the Role of Oncogenic Human Papilloma Virus

“…In l990, Brandsma et al 33 determined that the PCR method of HPV detection was reliable when archived paraffin‐embedded tissue was used as a study specimen. The PCR method was also recognized to be more sensitive and less tedious than the Southern blot hybridization technique of viral genome identification.…”

“…By 2000, investigators determined that the oropharynx and dominantly the palatine tonsils harbored the HPV genome in approximately 50% of all identified SCCas at that site, making this the dominant site in the HN in the United States. 5,7 In l990, Brandsma et al 33 determined that the PCR method of HPV detection was reliable when archived paraffin-embedded tissue was used as a study specimen. The PCR method was also recognized to be more sensitive and less tedious than the Southern blot hybridization technique of viral genome identification.…”

Papilomatose laríngea recorrente: experiência de 10 anos

“…Quanto à etiologia, experimentos clínicos e estudos com microscopia óptica e eletrônica têm sugerido relação da papilomatose laríngea com infecções virais 5,6 . Somente com o desenvolvimento da biologia molecular e de diferentes técnicas de hibridização do DNA viral foi possível a demonstração desta relação, tendo sido confirmada pela técni-ca de PCR (polymerase chain reaction) 7,8,9 . Recentemente, foi possível definir que o papilloma virus humano (HPV), um DNA-vírus pertencente ao grupo de vírus lentos da família Papovaviridae é o agente causal da papilomatose laríngea.…”

“…Na forma adulta, demonstrou-se que a papilomatose laríngea é causada pelos tipos 6 e/ou 11 dentre os mais de 80 tipos já identificados 7,10 . Nas lesões localizadas, apenas um tipo é encontrado, diferente das lesões múltiplas, onde ambos os tipos podem ser detectados.…”

Molecular biology: The polymerase chain reaction

“…The practical limit of template length is approximately 2 kilobases (kb), although fragments up to 10 kb have been synthesized. Genetically engineered mutants of Taq polymerase may soon allow amplification of longer ~t r a n d s .~ With the power of automated PCR available, amplification of an undesired piece of DNA is an unfavorable possibility.…”