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Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus
Published on: March 8, 2012
Detection and typing of human papillomavirus by e6 nested multiplex PCR.
K Sotlar1, D Diemer, A Dethleffs
1Institute of Pathology, University of Tübingen, Liebermeisterstrasse 8, 72076 Tübingen, Germany. klsotlar@med.uni-tuebingen.de
Journal of Clinical Microbiology
|July 10, 2004
Summary
A new nested multiplex PCR (NMPCR) assay accurately detects and types 18 human papillomavirus (HPV) genotypes in cervical samples. This sensitive method is valuable for identifying multiple HPV infections and high-risk genotypes, aiding in cervical cancer screening.
Area of Science:
- Virology
- Molecular Diagnostics
- Oncology
Background:
- Human papillomavirus (HPV) is a major cause of cervical cancer.
- Accurate detection and genotyping of HPV are crucial for cervical cancer prevention and management.
- Existing PCR methods have limitations in sensitivity and comprehensive genotyping.
Purpose of the Study:
- To evaluate a novel nested multiplex PCR (NMPCR) assay for detecting and typing 18 HPV genotypes.
- To compare the performance of NMPCR with conventional and nested PCR methods.
- To assess HPV prevalence and genotype distribution in relation to cervical intraepithelial neoplasia (CIN) grades.
Main Methods:
- Developed and validated a nested multiplex PCR (NMPCR) assay using degenerate E6/E7 consensus and type-specific primers.
- Tested the NMPCR assay on HPV DNA plasmids and 1,525 cervical scrapes.
- Compared NMPCR performance against conventional PCR (MY09-MY11, GP5+-GP6+) and nested MY/GP PCR in 495 cervical scrapes.
Main Results:
- NMPCR demonstrated significantly higher sensitivity than MY09-MY11 and GP5+-GP6+ assays, comparable to the MY/GP assay.
- All 18 targeted HPV genotypes were detected in clinical samples.
- HPV prevalence increased with CIN grade (34.7% in CIN 0, 94.2% in CIN I/II, 97.8% in CIN III).
- The ratio of high-risk to low-risk HPV genotypes increased significantly from CIN 0 to CIN I-III.
- Multiple HPV infections were detected in 47.9% of patients.
Conclusions:
- The novel NMPCR method is a highly sensitive and effective tool for HPV DNA detection and genotyping.
- NMPCR is particularly useful for identifying multiple HPV infections and specific genotypes.
- This assay can significantly improve the accuracy of HPV detection in cervical cancer screening programs.
