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Detection assay for HPV16 and HPV18 by loop-mediated isothermal amplification with lateral flow dipstick tests
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Document Title
Detection assay for HPV16 and HPV18 by loop-mediated isothermal amplification with lateral flow dipstick tests
Author
Kumvongpin R, Jearanaikoon P, Wilailuckana C, Sae-Ung N, Prasongdee P, Daduang S, Wongsena M, Boonsiri P, Kiatpathomchai W, Swangvaree SS, Sandee A, Daduang J
Name from Authors Collection
Affiliations
Khon Kaen University; Khon Kaen University; Khon Kaen University; National Science & Technology Development Agency - Thailand; National Center Genetic Engineering & Biotechnology (BIOTEC); National Cancer Institute - Thailand; Chulabhorn Research Institute; Khon Kaen University; Khon Kaen University
Type
Article
Source Title
MOLECULAR MEDICINE REPORTS
Year
2017
Volume
15
Issue
5
Page
3203-3209
Open Access
Bronze
Publisher
SPANDIDOS PUBL LTD
DOI
10.3892/mmr.2017.6370
Format
Abstract
Cervical cancer is the third highest cause of death in developing countries and most commonly results from high-risk human papillomavirus (HR-HPV) infection. Among HR-HPV genotypes, HPV16 and HPV18 are the most prevalent in cervical cancers. Therefore, the present study aimed to develop a detection assay for HPV16 and HPV18 infection using loop-mediated isothermal amplification (LAMP) with lateral flow dipstick (LFD) tests. This assay is a simplified, user-friendly method for the visual detection of HPV genotypes. DNA was extracted from clinical tissue samples, and HPV genotyping was performed using nested polymerase chain reaction (PCR). The clinical samples were demonstrated to include 44 HPV16-positive, 18 HPV18-positive and 80 HPV-negative samples. All DNA samples were also used as templates for a LAMP reaction (30 min at 65 degrees C), and subsequently, a fluorescein isothiocyanate-labelled probe was hybridized with the reaction product. Finally, the LFD test was performed. The sensitivity of the LAMP-LFD test was higher than LAMP-turbidity, exhibiting up to 100-fold higher sensitivity for HPV16 and 10-fold higher sensitivity for HPV18. All HPV16 and HPV18-positive samples generated positive results in both assays; however, 22 samples detected as HPV-negative by LAMP-turbidity exhibited positive results by LAMP-LFD test (22 of 80 samples). Therefore, these samples were further examined using quantitative (q) PCR. The results demonstrated that 20 out of the 22 samples designated positive by LAMP-LFD, but negative by LAMP turbidity, gave a positive result with qPCR, while the remaining 2 samples were negative by qPCR. The present results suggested that LAMP-LFD provided higher sensitivity than LAMP-turbidity and nested PCR. Thus, the LAMP-LFD test developed in the present study might be useful for the detection of HPV16 and HPV18 in local hospitals.
Funding Sponsor
Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences; HPV EBV; Carcinogenesis Research Group, Khon Kaen University, Thailand [KKU-2556]
License
Copyright
Publication Source
WOS