Fast solid support detection of human papillomavirus in in vitro amplified DNA using a DNP-anti DNP monoclonal antibody couple
| Autor: | P. Bernard, B. Alpha-Bazin, Jean-Marie Seigneurin, S. Sauvaigo, R. Téoule, M. Baccard-Longère, C. Chypre |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Male
Genital Neoplasms Female medicine.drug_class Molecular Sequence Data Biotin Biology Monoclonal antibody Polymerase Chain Reaction Virus law.invention Immunoenzyme Techniques chemistry.chemical_compound Bacterial Proteins law Virology medicine Humans DNA Probes HPV Prospective Studies Papillomaviridae Penile Neoplasms Immunosorbent Techniques Polymerase chain reaction Southern blot Base Sequence Papillomavirus Infections Antibodies Monoclonal virus diseases Anus Neoplasms Uterine Cervical Dysplasia Blotting Southern Tumor Virus Infections chemistry Condylomata Acuminata Biotinylation DNA Viral Female Streptavidin Oligomer restriction Nested polymerase chain reaction Dinitrophenols DNA |
| Zdroj: | Journal of Virological Methods. 46:29-38 |
| ISSN: | 0166-0934 |
| DOI: | 10.1016/0166-0934(94)90014-0 |
| Popis: | In some anogenital lesions the detection of certain types of human papilloma virus, especially oncogenic types, is of interest. In a first step during a prospective study, we compared two methods for the detection of human papillomavirus (HPV) DNA in clinical samples: Southern blotting followed by hybridization with a cloned radioactive genomic probe and a classical polymerase chain reaction (PCR) followed by hybridization with a 32 P-labelled oligonucleotide probe. 118 biopsies and swabs were examined for HPV 6/11, 16, 18 and 33, 67 positive reactions were found by both methods, 5 positives only by PCR and 2 positives only by Southern blot for unidentified HPV. Patients with anogenital condylomas, dysplasias and carcinomas or asymptomatic patients were studied. Most high grade (II and III) dysplasias were associated with HPV 16 and HPV 18. Condylomata lesions and low grade dysplasia (grade I) were associated mostly with HPV 6/11, mixed type of HPV, less frequently with HPV 16 or HPV 18. As a second step a nested PCR coupled to solid support detection method was used as described by Sauvaigo et al. (1990) Nucleic Acids Res. 18, 3175–3183) to study a panel of 30 previously qualified different HPV DNA extracts. In this procedure the second round of PCR amplification involves biotinylated and dinitrophenylated labelled primers allowing the capture of PCR amplified HPV DNA sequences on streptavidin coated tubes and its revelation. We describe an improvement of HPV DNA detection by means of single-step immunoenzymatic revelation involving anti-DNP monoclonal antibodies conjugated to horseradish peroxidase enzyme. A perfect correlation with the previous results was obtained. This solid support method allows a faster and easier HPV typing compared to methods using membrane transfer. |
| Databáze: | OpenAIRE |
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