title: Retrieval of HPV oncogenes E6 and E7 mRNA from cervical specimens using a manual open technology protocol.
abstract: BACKGROUND
  HPV oncogenes mRNA detection gains momentum as an adjuvant for HPV-related cervical abnormalities diagnosis, but is based on costly detection assays not allowing viral type targeting.
  
  
  OBJECTIVE
  To assess detection rate of HPV oncogenes E6/E7 mRNA from cervical specimens using a manual, open technology, fully customizable protocol and determine whether HPV-related epidemiological features influence mRNA retrieval. We reviewed literature and compared our retrieval rate with automated technologies.
  
  
  METHODS
  We used 60 samples positive for HPV DNA types 16, 18, 31 and/or 45. We extracted mRNA with a TRizol-based protocol, and tested mRNA purity and concentration using light absorbance. We reverse-transcribed mRNA into cDNA for E6/7 detection.
  
  
  RESULTS
  HPV oncogenes E6/E7 mRNA was retrieved from 36 (60%) out of 60 specimens. No HPV load-related clinical or epidemiological feature was significantly associated with mRNA retrieval. Presence of HPV-DNA 16/18 was associated with mRNA retrieval (OR = 9.08; 95% CI 1.26 to 65.32 for HPV 16; and 18.2; IC95% 1.86 to 391.44 for HPV 18).
  
  
  CONCLUSIONS
  The open-technology protocol yielded an mRNA detection rate similar to that of automated technologies. Advantages are lower costs and target HPV type customization.
authors.affiliation: Gynecologic Oncology Department, Brasilia Mothers & Babies Hospital, SGAS 608 Mód A, Brasília, DF 70203-900 Brazil.
authors.forename: Leonardo Martins
authors.initials: LM
authors.lastname: Campbell
authors.forename: Denise Rocha
authors.initials: DR
authors.lastname: Pitta
authors.forename: Angela Maria
authors.initials: AM
authors.lastname: De Assis
authors.forename: Sophie Francoise Mauricette
authors.initials: SF
authors.lastname: Derchain
authors.forename: Elisabete Aparecida
authors.initials: EA
authors.lastname: Campos
authors.forename: Luis Otavio Zanatta
authors.initials: LO
authors.lastname: Sarian