Molecular detection of human papillomavirus (HPV) in highly fragmented DNA from cervical cancer biopsies using double-nested PCR

Abstract

Graphical abstract

Comparison of HPV detection rate in formalin‐fixed paraffin‐embedded tissues of head and neck carcinoma using two DNA extraction kits and three amplification methods

“…The abovementioned findings by us and others indicate the need to use nested PCR, as it is a more sensitive method compared to single PCR, for detection of HPV DNA in FFPE samples. Recently, TAWE et al (39) proposed a double-nested PCR method with significant improvement of HPV positivity from cervical cancer biopsy tissues (one-step PCR -3.0%, nested PCR -55.2%, and double-nested PCR -96.3% HPV positivity rate). To decrease the risk of contamination, the authors of that study recommended the use of three separate PCR work stations for the sample preparation stage, the PCR setup stage, and the post-PCR stage.…”

“…The HPV detection using Real-time PCR method was done using the commercial HPV Screening kit (Sansure Biotech, Changsha, China) according to the manufacturer's instructions. The test is designed for HPV DNA detection of 15 high-risk types (HPV 16,18,31,33,35,39,45,51,52,53,56,58,59, 66, and 68) with the detection limit of 1,000 copies ml À1 , but without the possibility of genotyping. This assay includes internal control for the detection of human beta-globin.…”

Promoter Hypermethylation Analysis of Host Genes in Cervical Cancer Patients With and Without Human Immunodeficiency Virus in Botswana

“…DNA was extracted from FFPE cervical specimens using an established protocol (38). DNA concentration was measured and quality assessed.…”

“…Touchdown PCR was designed and used due to its ability to amplify degraded DNA associated with formalin fixation and long-term storage in paraffin. PCR products were run on an agarose gel, and the results are reported as methylated, unmethylated, or a mixture of both at the target DNA sequence (34)(35)(36)(37)(38)(39)(40)(41)(42)(43). Methylation-specific PCR was performed twice on all specimens with a third repeat performed if discrepant results were obtained from the first two runs.…”

Research Mentoring Plan Involving North-South Collaborations

“…The study has now been published. 8 The second phase was to identify the prevalence of HPV genotypes among the study population. A manuscript is under review.…”