Detection of human papillomavirus type-16 DNA utilising microtitre-plate based amplification reactions and a solid-phase enzyme-immunoassay detection system

被引:13
作者
Cavuslu, S
Starkey, WG
Kaye, JN
Biswas, C
Mant, C
Kell, B
Rice, P
Best, JM
Cason, J
机构
[1] UNITED MED & DENT SCH, ST THOMAS HOSP, DEPT VIROL, RAYNE INST, LONDON SE1 7EH, ENGLAND
[2] HAYDARPASA TEACHING HOSP, DEPT INFECT DIS, GULHANE MIL MED ACAD, HAYDARPASA 81327, ISTANBUL, TURKEY
基金
英国惠康基金;
关键词
human papillomavirus type 16; nested PCR; enzyme-immunoassay;
D O I
10.1016/0166-0934(95)01988-X
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The development of a nested polymerase chain reaction (PCR) assay to detect low concentrations of human papillomavirus type-16 (HPV-16) DNA for epidemiological studies is described. The PCR utilises primers located in the E5 open reading frame, has an analytical sensitivity of 4 HPV-16 genomes and does not produce amplicons from other common genital HPVs (types-6, -11, -18, -31 and 33). This assay was carried out in 96-well plates utilising internal primers labelled with dinitrophenol (DNP) and biotin so that amplicons can be captured onto streptavidin-coated plates and detected using an alkaline phosphatase-labelled monoclonal antibody to DNP. The assay was effective for detecting HPV-16 DNA in plasmids, cell-lines and, both freshly collected or archival (formalin-fixed/paraffin embedded) clinical specimens. This system is therefore suitable for epidemiological studies to identify individuals infected with HPV-16 DNA in episomal form who may be at increased risk of developing anogenital carcinomas.
引用
收藏
页码:59 / 69
页数:11
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