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A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens.

Abstract

Early diagnosis and genotyping of high-risk human papillomavirus (HR-HPV) in cervical tissue specimens is significant for cervical cancer prevention. A sensitive microplate fluorometric hybridization assay (MFHA) was designed for the detection of HPV DNA 16 and 18 in cervical tissue. Following optimization and validation of the method, 60 formalin-fixed and paraffin-embedded cervical samples representing different cervical intraepithelial neoplasia grades of HPV-associated lesions were tested to determine the sensitivity and specificity of the assay. Using consensus GP5+/6+ biotin-labeled primers to amplify a conserved region within the L1 gene, the amplicons were added to the microplate wells coated with specific probes for the hybridization of HPV 16 and 18 individually. Final detection was performed with streptavidin-AlexaFluor488 conjugated. The results were then compared with type-specific nested polymerase chain reaction (PCR) and colorimetric microplate assay. While the agreement between the results obtained by the type-specific nested PCR and fluorometric assay for the detection of both HR-HPV types was 100%, this agreement for the detection of HPV type 16 and 18 using microplate colorimetric assay was 94.2% and 85% respectively. Overall, the results of the fluorometric and colorimetric assays are promising for detecting both HR-HPV types in a large number of cervical tissue samples with the higher MFHA assay sensitivity.

Authors+Show Affiliations

Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran. Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.Organ Transplant Research Centre, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran.Diagnostic Laboratory Sciences and Technology Research Centre, School of Paramedical Sciences, Shiraz University of Medical Sciences, Shiraz, Iran. behzadba@sums.ac.ir.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31707717

Citation

Nikouyan, Negin, et al. "A Fluorometric Hybridization Assay for Detecting and Genotyping High-risk Human Papillomavirus 16 and 18 in Archival Tissues of Cervical Specimens." Brazilian Journal of Microbiology : [publication of the Brazilian Society for Microbiology], 2019.
Nikouyan N, Farhadi A, Gorzin AA, et al. A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens. Braz J Microbiol. 2019.
Nikouyan, N., Farhadi, A., Gorzin, A. A., Geramizadeh, B., Okhovat, M. A., Seyyedi, N., ... Behzad-Behbahani, A. (2019). A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens. Brazilian Journal of Microbiology : [publication of the Brazilian Society for Microbiology], doi:10.1007/s42770-019-00137-8.
Nikouyan N, et al. A Fluorometric Hybridization Assay for Detecting and Genotyping High-risk Human Papillomavirus 16 and 18 in Archival Tissues of Cervical Specimens. Braz J Microbiol. 2019 Nov 9; PubMed PMID: 31707717.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens. AU - Nikouyan,Negin, AU - Farhadi,Ali, AU - Gorzin,Ali Akbar, AU - Geramizadeh,Bita, AU - Okhovat,Mohammad Ali, AU - Seyyedi,Noorossadat, AU - Dehbidi,Gholamreza Rafiei, AU - Ranjbaran,Reza, AU - Behzad-Behbahani,Abbas, Y1 - 2019/11/09/ PY - 2018/07/19/received PY - 2019/02/22/accepted PY - 2019/11/11/entrez PY - 2019/11/11/pubmed PY - 2019/11/11/medline KW - Cervical cancer KW - FFPE cervical samples KW - Fluorometric assay KW - High-risk human papillomavirus (HR-HPV) KW - Microplate hybridization JF - Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology] JO - Braz. J. Microbiol. N2 - Early diagnosis and genotyping of high-risk human papillomavirus (HR-HPV) in cervical tissue specimens is significant for cervical cancer prevention. A sensitive microplate fluorometric hybridization assay (MFHA) was designed for the detection of HPV DNA 16 and 18 in cervical tissue. Following optimization and validation of the method, 60 formalin-fixed and paraffin-embedded cervical samples representing different cervical intraepithelial neoplasia grades of HPV-associated lesions were tested to determine the sensitivity and specificity of the assay. Using consensus GP5+/6+ biotin-labeled primers to amplify a conserved region within the L1 gene, the amplicons were added to the microplate wells coated with specific probes for the hybridization of HPV 16 and 18 individually. Final detection was performed with streptavidin-AlexaFluor488 conjugated. The results were then compared with type-specific nested polymerase chain reaction (PCR) and colorimetric microplate assay. While the agreement between the results obtained by the type-specific nested PCR and fluorometric assay for the detection of both HR-HPV types was 100%, this agreement for the detection of HPV type 16 and 18 using microplate colorimetric assay was 94.2% and 85% respectively. Overall, the results of the fluorometric and colorimetric assays are promising for detecting both HR-HPV types in a large number of cervical tissue samples with the higher MFHA assay sensitivity. SN - 1678-4405 UR - https://www.unboundmedicine.com/medline/citation/31707717/A_fluorometric_hybridization_assay_for_detecting_and_genotyping_high-risk_human_papillomavirus_16_and_18_in_archival_tissues_of_cervical_specimens L2 - https://dx.doi.org/10.1007/s42770-019-00137-8 DB - PRIME DP - Unbound Medicine ER -