Α 2-stage, nested-like nucleic acid amplification method (IsoPCR) for the highly sensitive detection of HPV16 and HPV18 DNA

M Daskou; D Tsakogiannis; T G Dimitriou; M Manali; C Apti; G D Amoutzias; D Mossialos; C Kottaridi; P Markoulatos

doi:10.1016/j.mcp.2019.03.003

Α 2-stage, nested-like nucleic acid amplification method (IsoPCR) for the highly sensitive detection of HPV16 and HPV18 DNA

Mol Cell Probes. 2019 Jun:45:1-7. doi: 10.1016/j.mcp.2019.03.003. Epub 2019 Mar 19.

Authors

M Daskou¹, D Tsakogiannis², T G Dimitriou¹, M Manali¹, C Apti¹, G D Amoutzias³, D Mossialos¹, C Kottaridi⁴, P Markoulatos¹

Affiliations

¹ University of Thessaly, School of Health Sciences, Department of Biochemistry & Biotechnology, Microbiology-Virology Laboratory, Biopolis, 41500, Larissa, Greece.
² University of Thessaly, School of Health Sciences, Department of Biochemistry & Biotechnology, Microbiology-Virology Laboratory, Biopolis, 41500, Larissa, Greece. Electronic address: ditsakog@uth.gr.
³ Bioinformatics Laboratory, University of Thessaly, School of Health Sciences, Department of Biochemistry & Biotechnology, Biopolis, Larissa, Greece.
⁴ Department of Cytopathology, National and Kapodistrian University of Athens, Medical School, "ATTIKON" University Hospital, 1 Rimini, Haidari, 12462, Athens, Greece.

PMID: 30902662
DOI: 10.1016/j.mcp.2019.03.003

Abstract

Molecular detection of HPV DNA is considered as the gold standard for the diagnosis of cervical disease. Although the molecular assays for the identification of HPV16 and HPV18 have helped identify cervical cancer incidents, they are restricted to specialized laboratories. Thus, we developed a novel 2-stage, nested-like nucleic acid amplification method, named IsoPCR, to amplify the E6 gene of HPV16 and HPV18 with high analytical sensitivity and specificity. The performance of IsoPCR was compared to that of conventional PCR and LAMP. The analytical sensitivity of IsoPCR (1 copy/test) was 10-fold higher than conventional PCR and 25-fold higher than conventional LAMP. IsoPCR displayed significant amplification specificity (100%) and efficiency, as well. In conclusion, IsoPCR is a highly sensitive and specific diagnostic tool and it is suitable for the detection of low copy number of viral DNA in clinical specimens, providing critical information to healthcare providers.

Keywords: Cervical dysplasia; HPV genotyping; HPV16; HPV18; IsoPCR; LAMP.

Publication types

Comparative Study
Research Support, Non-U.S. Gov't

MeSH terms

DNA, Viral / genetics
DNA-Binding Proteins / genetics*
Female
Human papillomavirus 16 / genetics
Human papillomavirus 16 / isolation & purification
Human papillomavirus 18 / genetics
Human papillomavirus 18 / isolation & purification
Humans
Molecular Diagnostic Techniques / methods
Nucleic Acid Amplification Techniques / methods*
Oncogene Proteins, Viral / genetics*
Papillomavirus Infections / diagnosis*
Polymerase Chain Reaction / methods
Repressor Proteins / genetics*
Sensitivity and Specificity
Uterine Cervical Neoplasms / virology*

Substances

DNA, Viral
DNA-Binding Proteins
E6 protein, Human papillomavirus type 16
E6 protein, Human papillomavirus type 18
Oncogene Proteins, Viral
Repressor Proteins

Supplementary concepts

LAMP assay