Skip to main content
NCBI home page
Journal of Clinical Microbiology logoLink to Journal of Clinical Microbiology
. 1987 Sep;25(9):1717–1721. doi: 10.1128/jcm.25.9.1717-1721.1987

Sensitive reverse transcriptase assay to detect and quantitate human immunodeficiency virus.

M H Lee 1, K Sano 1, F E Morales 1, D T Imagawa 1
PMCID: PMC269314  PMID: 2443532

Abstract

A sensitive biochemical assay of viral reverse transcriptase (RT) was developed that is useful for both the detection and quantitation of human immunodeficiency virus (HIV), the agent responsible for acquired immune deficiency syndrome in humans. This assay gave a 20- to 40-fold increase in enzyme activity over the current method used for RT detection of HIV. The test is based on a previous biochemical study showing the unusual stability of avian oncornavirus RNA-dependent DNA polymerases at 30 degrees C for at least 2 days. Our study shows that the HIV polymerase is stable at 30 to 37 degrees C for up to 3 days. By using this sensitive RT assay, as few as 250 HIV virions can be quantitated directly in tissue culture medium. This assay should prove useful in studies in which the detection of HIV or the quantitation of the number of virions is required.

Full text

PDF
1717

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barré-Sinoussi F., Chermann J. C., Rey F., Nugeyre M. T., Chamaret S., Gruest J., Dauguet C., Axler-Blin C., Vézinet-Brun F., Rouzioux C. Isolation of a T-lymphotropic retrovirus from a patient at risk for acquired immune deficiency syndrome (AIDS). Science. 1983 May 20;220(4599):868–871. doi: 10.1126/science.6189183. [DOI] [PubMed] [Google Scholar]
  2. Biberfeld G., Bredberg-Råden U., Böttiger B., Biberfeld P., Morfeldt-Månsson L., Suni J., Vaheri A., Saxinger C., Gallo R. Antibodies to human T lymphotropic virus type III demonstrated by a dot immunobinding assay. Scand J Immunol. 1985 Mar;21(3):289–292. doi: 10.1111/j.1365-3083.1985.tb01432.x. [DOI] [PubMed] [Google Scholar]
  3. Brun-Vezinet F., Rouzioux C., Barre-Sinoussi F., Klatzmann D., Saimot A. G., Rozenbaum W., Christol D., Gluckmann J. C., Montagnier L., Chermann J. C. Detection of IgG antibodies to lymphadenopathy-associated virus in patients with AIDS or lymphadenopathy syndrome. Lancet. 1984 Jun 9;1(8389):1253–1256. doi: 10.1016/s0140-6736(84)92444-9. [DOI] [PubMed] [Google Scholar]
  4. Essex M., McLane M. F., Lee T. H., Falk L., Howe C. W., Mullins J. I., Cabradilla C., Francis D. P. Antibodies to cell membrane antigens associated with human T-cell leukemia virus in patients with AIDS. Science. 1983 May 20;220(4599):859–862. doi: 10.1126/science.6342136. [DOI] [PubMed] [Google Scholar]
  5. FOLEY G. E., LAZARUS H., FARBER S., UZMAN B. G., BOONE B. A., MCCARTHY R. E. CONTINUOUS CULTURE OF HUMAN LYMPHOBLASTS FROM PERIPHERAL BLOOD OF A CHILD WITH ACUTE LEUKEMIA. Cancer. 1965 Apr;18:522–529. doi: 10.1002/1097-0142(196504)18:4<522::aid-cncr2820180418>3.0.co;2-j. [DOI] [PubMed] [Google Scholar]
  6. Fine D. L., Arthur L. O., PLOWMAN J. K., Hillman E. A., Klein F. In vitro system for production of mouse mammary tumor virus. Appl Microbiol. 1974 Dec;28(6):1040–1046. doi: 10.1128/am.28.6.1040-1046.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gallo R. C., Salahuddin S. Z., Popovic M., Shearer G. M., Kaplan M., Haynes B. F., Palker T. J., Redfield R., Oleske J., Safai B. Frequent detection and isolation of cytopathic retroviruses (HTLV-III) from patients with AIDS and at risk for AIDS. Science. 1984 May 4;224(4648):500–503. doi: 10.1126/science.6200936. [DOI] [PubMed] [Google Scholar]
  8. Harada S., Koyanagi Y., Yamamoto N. Infection of HTLV-III/LAV in HTLV-I-carrying cells MT-2 and MT-4 and application in a plaque assay. Science. 1985 Aug 9;229(4713):563–566. doi: 10.1126/science.2992081. [DOI] [PubMed] [Google Scholar]
  9. Hoffman A. D., Banapour B., Levy J. A. Characterization of the AIDS-associated retrovirus reverse transcriptase and optimal conditions for its detection in virions. Virology. 1985 Dec;147(2):326–335. doi: 10.1016/0042-6822(85)90135-7. [DOI] [PubMed] [Google Scholar]
  10. Levy J. A., Hoffman A. D., Kramer S. M., Landis J. A., Shimabukuro J. M., Oshiro L. S. Isolation of lymphocytopathic retroviruses from San Francisco patients with AIDS. Science. 1984 Aug 24;225(4664):840–842. doi: 10.1126/science.6206563. [DOI] [PubMed] [Google Scholar]
  11. Liebes L. F., Rich M. A., McCormick J. J., Salmeen I., Rimai L. Reverse transcriptase activity per virion for avian myeloblastosis virus and Rauscher murine leukemia virus. J Virol. 1976 Apr;18(1):42–47. doi: 10.1128/jvi.18.1.42-47.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. McDougal J. S., Cort S. P., Kennedy M. S., Cabridilla C. D., Feorino P. M., Francis D. P., Hicks D., Kalyanaraman V. S., Martin L. S. Immunoassay for the detection and quantitation of infectious human retrovirus, lymphadenopathy-associated virus (LAV). J Immunol Methods. 1985 Jan 21;76(1):171–183. doi: 10.1016/0022-1759(85)90489-2. [DOI] [PubMed] [Google Scholar]
  13. Mitsuya H., Broder S. Inhibition of the in vitro infectivity and cytopathic effect of human T-lymphotrophic virus type III/lymphadenopathy-associated virus (HTLV-III/LAV) by 2',3'-dideoxynucleosides. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1911–1915. doi: 10.1073/pnas.83.6.1911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Popovic M., Sarngadharan M. G., Read E., Gallo R. C. Detection, isolation, and continuous production of cytopathic retroviruses (HTLV-III) from patients with AIDS and pre-AIDS. Science. 1984 May 4;224(4648):497–500. doi: 10.1126/science.6200935. [DOI] [PubMed] [Google Scholar]
  15. Resnick L., Veren K., Salahuddin S. Z., Tondreau S., Markham P. D. Stability and inactivation of HTLV-III/LAV under clinical and laboratory environments. JAMA. 1986 Apr 11;255(14):1887–1891. [PubMed] [Google Scholar]
  16. Rey M. A., Spire B., Dormont D., Barre-Sinoussi F., Montagnier L., Chermann J. C. Characterization of the RNA dependent DNA polymerase of a new human T-lymphotropic retrovirus (lymphadenopathy associated virus). Biochem Biophys Res Commun. 1984 May 31;121(1):126–133. doi: 10.1016/0006-291x(84)90696-x. [DOI] [PubMed] [Google Scholar]
  17. Rho H. M., Poiesz B., Ruscetti F. W., Gallo R. C. Characterization of the reverse transcriptase from a new retrovirus (HTLV) produced by a human cutaneous T-cell lymphoma cell line. Virology. 1981 Jul 15;112(1):355–360. doi: 10.1016/0042-6822(81)90642-5. [DOI] [PubMed] [Google Scholar]
  18. Sano K. Electron microscopic comparison of adult T-cell leukemia associated virus (ATLV) and murine leukemia virus (MuLV). Bull Osaka Med Sch. 1984 Jul;30(1):1–13. [PubMed] [Google Scholar]
  19. Schüpbach J., Popovic M., Gilden R. V., Gonda M. A., Sarngadharan M. G., Gallo R. C. Serological analysis of a subgroup of human T-lymphotropic retroviruses (HTLV-III) associated with AIDS. Science. 1984 May 4;224(4648):503–505. doi: 10.1126/science.6200937. [DOI] [PubMed] [Google Scholar]
  20. Shaw G. M., Harper M. E., Hahn B. H., Epstein L. G., Gajdusek D. C., Price R. W., Navia B. A., Petito C. K., O'Hara C. J., Groopman J. E. HTLV-III infection in brains of children and adults with AIDS encephalopathy. Science. 1985 Jan 11;227(4683):177–182. doi: 10.1126/science.2981429. [DOI] [PubMed] [Google Scholar]
  21. Spire B., Dormont D., Barré-Sinoussi F., Montagnier L., Chermann J. C. Inactivation of lymphadenopathy-associated virus by heat, gamma rays, and ultraviolet light. Lancet. 1985 Jan 26;1(8422):188–189. doi: 10.1016/s0140-6736(85)92026-4. [DOI] [PubMed] [Google Scholar]
  22. Tereba A., Murti K. G. A very sensitive biochemical assay for detecting and quantitating avian oncornaviruses. Virology. 1977 Jul 1;80(1):166–176. doi: 10.1016/0042-6822(77)90389-0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES