A sensitive non-isotopic assay specific for HIV-1 associated reverse transcriptase
| Autor: | Katrin Meissner, Ralf W. Glaser, Sven-Holger Döpel, Tomas Porstmann, Günter Sydow |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
medicine.drug_class
Biology Monoclonal antibody Sensitivity and Specificity Virus Immunoenzyme Techniques chemistry.chemical_compound Virology medicine Thymidine triphosphate chemistry.chemical_classification Hydrolysis Antibodies Monoclonal RNA-Directed DNA Polymerase Templates Genetic Molecular biology Reverse transcriptase Enzyme chemistry Monoclonal biology.protein HIV-1 Primer (molecular biology) Antibody Deoxyuracil Nucleotides Thymidine |
| Zdroj: | Journal of virological methods. 31(2-3) |
| ISSN: | 0166-0934 |
| Popis: | A sensitive non-isotopic assay for specific detection of reverse transcriptase (RT) of the human immunodeficiency virus type 1 (HIV-1) is described using 5-bromo-2'-deoxyuridine triphosphate (BrdUTP) instead of tritiated thymidine triphosphate. After the RT reaction the template primer is degraded by alkaline hydrolysis. Single-stranded poly.(BrdU) is detected in an immunoenzymometric assay using monoclonal anti-BrdU antibodies. The specificity of the assay is demonstrated by the isolation of RT from virus lysate by an insolubilised monoclonal anti-HIV-1 RT antibody prior to the RT reaction. Immunological RT binding leads to a tenfold increase in analytical sensitivity since substances inhibiting the RT reaction can be removed. This non-isotopic assay is some 30 times more sensitive than the classical radioisotopic RT assay. In terms of RT determination, however, there is a good correlation between these tests (r = 0.96). Several filtrations are no longer necessary to remove non-incorporated nucleotides. The test can be adapted to microtitre plates and hence is easy to automate. |
| Databáze: | OpenAIRE |
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