A comparison of clinically utilized human papillomavirus detection methods in head and neck cancer

Mod Pathol. 2011 Oct;24(10):1295-305. doi: 10.1038/modpathol.2011.91. Epub 2011 May 13.


Detection of human papillomavirus (HPV) in head and neck cancer has therapeutic implications. In situ hybridization and immunohistochemistry for p16 are used by surgical pathologists. We compared the sensitivity and specificity of three popular commercial tests for HPV detection in head and neck squamous cell carcinomas with a 'gold standard' HPV PCR assay. A total of 110 prospectively collected, formalin-fixed tumor specimens were compiled onto tissue microarrays and tested for HPV DNA by in situ hybridization with two probe sets, a biotinylated probe for high-risk (HR) HPV types 16/18 (Dako, CA, USA) and a probe cocktail for 16/18, plus 10 additional HR types (Ventana, AZ, USA). The p16(INK4) expression was also assessed using a Pharmingen immunohistochemistry antibody (BD Biosciences, CA, USA). Tissue microarrays were stained and scored at expert laboratories. HPV DNA was detected by MY09/11-PCR, using Gold AmpliTaq and dot-blot hybridization on matched-fresh frozen specimens in a research laboratory. HPV 16 E6 and E7-RNA expression was also measured using RT-PCR. Test performance was assessed by a receiver operating characteristic analysis. HR-HPV DNA types 16, 18 and 35 were detected by MY-PCR in 28% of tumors, with the majority (97%) testing positive for type 16. Compared with MY-PCR, the sensitivity and specificity for HR-HPV DNA detection with Dako in situ hybridization was 21% (95% confidence interval (CI): 7-42) and 100% (95% CI: 93-100), respectively. Corresponding test results by Ventana in situ hybridization were 59% (95% CI: 39-78) and 58% (95% CI: 45-71), respectively. The p16 immunohistochemistry performed better overall than Dako (P=0.042) and Ventana (P=0.055), with a sensitivity of 52% (95% CI: 32-71) and specificity of 93% (95% CI: 84-98). Compared with a gold standard HPV-PCR assay, HPV detection by in situ hybridization was less accurate for head and neck squamous cell carcinoma on tissue microarrays than p16 immunohistochemistry. Further testing is warranted before these assays should be recommended for clinical HPV detection.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • Biomarkers, Tumor / analysis*
  • Biopsy
  • Carcinoma, Squamous Cell / chemistry
  • Carcinoma, Squamous Cell / diagnosis*
  • Carcinoma, Squamous Cell / virology
  • Cyclin-Dependent Kinase Inhibitor p16 / analysis*
  • DNA, Viral / analysis*
  • Female
  • Head and Neck Neoplasms / chemistry
  • Head and Neck Neoplasms / diagnosis*
  • Head and Neck Neoplasms / virology
  • Humans
  • Immunohistochemistry*
  • In Situ Hybridization*
  • Male
  • Middle Aged
  • Oncogene Proteins, Viral / genetics
  • Papillomaviridae / genetics*
  • Papillomavirus E7 Proteins / genetics
  • Papillomavirus Infections / complications
  • Papillomavirus Infections / diagnosis*
  • Papillomavirus Infections / virology
  • Polymerase Chain Reaction*
  • Predictive Value of Tests
  • Prognosis
  • Prospective Studies
  • ROC Curve
  • Repressor Proteins / genetics
  • Sensitivity and Specificity
  • Tissue Array Analysis


  • Biomarkers, Tumor
  • Cyclin-Dependent Kinase Inhibitor p16
  • DNA, Viral
  • E6 protein, Human papillomavirus type 16
  • Oncogene Proteins, Viral
  • Papillomavirus E7 Proteins
  • Repressor Proteins
  • oncogene protein E7, Human papillomavirus type 16