title: Comparison of five protocols to extract DNA from paraffin-embedded tissues for the detection of human papillomavirus.
abstract: Formalin-fixed paraffin-embedded (FFPE) tissues are a valuable source of DNA with which to perform large retrospective studies on the epidemiology of HPV infection. Five different DNA extraction protocols were carried out to evaluate the DNA obtained from FFPE samples with polymerase chain reaction (PCR) using two primer sets to amplify a constitutive human gene, β-globin, and two primer sets to detect the L1 and E6 HPV genes. From the five DNA extraction protocols evaluated, the best results were obtained with protocol A, corresponding to a crude extract from the sample. With the procedures described herein, we were able to amplify DNA extracted from archival paraffin blocks stored for six years. However, the amplification products were more efficiently obtained with primers that amplified shorter fragments. This result indicates that a major factor limiting the extraction process in these samples is DNA fragmentation, a factor that will naturally vary between the different specimens evaluated. Also, depending upon the extraction method, PCR amplification of a human gene does not necessarily guarantee the successful extraction of viral DNA. In conclusion, different DNA and HPV detection methods can significantly influence the results. Therefore, the DNA extraction methods and primers used for DNA amplification in fixed tissues need to be chosen carefully, depending on the specific requirements of the study being carried out.
authors.affiliation: Facultad de Ciencias de la Salud, Departamento de Ciencias Básicas, Grupo Infección e Inmunidad, Universidad Tecnológica de Pereira, Pereira, Colombia. Electronic address: adalucyalvarez@utp.edu.co.
authors.forename: Adalucy
authors.initials: A
authors.lastname: Alvarez-Aldana
authors.affiliation: Facultad de Ciencias de la Salud, Departamento de Medicina Comunitaria, Grupo Epidemiología, Salud y Violencia, Universidad Tecnológica de Pereira, Pereira, Colombia. Electronic address: jose01william@utp.edu.co.
authors.forename: José William
authors.initials: JW
authors.lastname: Martínez
authors.affiliation: Facultad de Ciencias de la Salud, Departamento de Ciencias Básicas, Grupo Infección e Inmunidad, Universidad Tecnológica de Pereira, Pereira, Colombia. Electronic address: jcsepulv@utp.edu.co.
authors.forename: Juan C
authors.initials: JC
authors.lastname: Sepúlveda-Arias
mesh_terms: D052685:Alphapapillomavirus / Q000235:genetics / Q000302:isolation & purification*; D002294:Carcinoma, Squamous Cell / Q000473:pathology / Q000821:virology*; D004279:DNA, Viral / Q000032:analysis / Q000302:isolation & purification*; D005260:Female; D005819:Genetic Markers; D006801:Humans; D016612:Paraffin Embedding; D012189:Retrospective Studies; D002583:Uterine Cervical Neoplasms / Q000473:pathology / Q000821:virology*; D014764:Viral Proteins / Q000235:genetics; D002578:Uterine Cervical Dysplasia / Q000473:pathology / Q000821:virology*
chemical_list: D004279:DNA, Viral; D005819:Genetic Markers; D014764:Viral Proteins
keywords: Cervical cancer; DNA extraction; Deoxyribonucleic acid; Formalin fixed paraffin-embedded tissues; Human papillomavirus