Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1995 Jun;69(6):3807–3815. doi: 10.1128/jvi.69.6.3807-3815.1995

Serotyping of primary human immunodeficiency virus type 1 isolates from diverse geographic locations by flow cytometry.

S Zolla-Pazner 1, J O'Leary 1, S Burda 1, M K Gorny 1, M Kim 1, J Mascola 1, F McCutchan 1
PMCID: PMC189098  PMID: 7745728

Abstract

The immunologic relatedness of the various human immunodeficiency virus type 1 (HIV-1) clades was determined with 13 human anti-HIV-1 monoclonal antibodies (MAbs) to six immunogenic regions of the HIV-1 structural proteins. The immunoreactivity of the native, oligomeric viral envelope glycoproteins expressed on the surfaces of human peripheral blood mononuclear cells infected in vitro with primary isolates from clades A through E was determined by flow cytometry. Some epitopes in the immunodominant region of gp41 and the C terminus of gp120 appear to be HIV-1 group specific in that they are expressed on the surfaces of cells in cultures infected with the majority of viruses tested from clades A to E. Epitopes within the V3 region appear to be clade restricted. Surprisingly, one MAb to an epitope in the C terminus of gp120 was entirely clade B specific. Staining with anti-V2 and anti-CD4 binding domain (CD4bd) reagents was infrequently detected. Anti-CD4bd MAbs stained only CD4-negative T cells because the CD4bd of gp120 appeared to be complexed with membrane CD4. When present, the epitopes of V2 and the CD4bd appeared to be expressed on cells infected with various clades. Thus, the results suggest that MAbs to gp41, the C terminus, and the V3 loop of gp120 are most useful in serotyping primary isolates of HIV-1, providing group-specific, clade-restricted, and clade-specific reagents. The use of the immunofluorescent method with the reagents described herein distinguishes infection with clade B from that with all other HIV-1 clades. With additional MAbs, this technique will allow a broadly applicable, reproducible, and practical method for serotyping HIV-1.

Full Text

The Full Text of this article is available as a PDF (520.8 KB).

Selected References

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

  1. Akerblom L., Hinkula J., Broliden P. A., Mäkitalo B., Fridberger T., Rosen J., Villacres-Eriksson M., Morein B., Wahren B. Neutralizing cross-reactive and non-neutralizing monoclonal antibodies to HIV-1 gp120. AIDS. 1990 Oct;4(10):953–960. doi: 10.1097/00002030-199010000-00002. [DOI] [PubMed] [Google Scholar]
  2. Arendrup M., Sönnerborg A., Svennerholm B., Akerblom L., Nielsen C., Clausen H., Olofsson S., Nielsen J. O., Hansen J. E. Neutralizing antibody response during human immunodeficiency virus type 1 infection: type and group specificity and viral escape. J Gen Virol. 1993 May;74(Pt 5):855–863. doi: 10.1099/0022-1317-74-5-855. [DOI] [PubMed] [Google Scholar]
  3. Barin F., M'Boup S., Denis F., Kanki P., Allan J. S., Lee T. H., Essex M. Serological evidence for virus related to simian T-lymphotropic retrovirus III in residents of west Africa. Lancet. 1985 Dec 21;2(8469-70):1387–1389. doi: 10.1016/s0140-6736(85)92556-5. [DOI] [PubMed] [Google Scholar]
  4. Blomberg J., Lawoko A., Pipkorn R., Moyo S., Malmvall B. E., Shao J., Dash R., Tswana S. A survey of synthetic HIV-1 peptides with natural and chimeric sequences for differential reactivity with Zimbabwean, Tanzanian and Swedish HIV-1-positive sera. AIDS. 1993 Jun;7(6):759–767. doi: 10.1097/00002030-199306000-00002. [DOI] [PubMed] [Google Scholar]
  5. Bou-Habib D. C., Roderiquez G., Oravecz T., Berman P. W., Lusso P., Norcross M. A. Cryptic nature of envelope V3 region epitopes protects primary monocytotropic human immunodeficiency virus type 1 from antibody neutralization. J Virol. 1994 Sep;68(9):6006–6013. doi: 10.1128/jvi.68.9.6006-6013.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bugge T. H., Lindhardt B. O., Hansen L. L., Kusk P., Hulgaard E., Holmbäck K., Klasse P. J., Zeuthen J., Ulrich K. Analysis of a highly immunodominant epitope in the human immunodeficiency virus type 1 transmembrane glycoprotein, gp41, defined by a human monoclonal antibody. J Virol. 1990 Sep;64(9):4123–4129. doi: 10.1128/jvi.64.9.4123-4129.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Burke D. S., Fowler A. K., Redfield R. R., Dilworth S., Oster C. N. Isolation of HIV-1 from the blood of seropositive adults: patient stage of illness and sample inoculum size are major determinants of a positive culture. The Walter Reed Retroviral Research Group. J Acquir Immune Defic Syndr. 1990;3(12):1159–1167. [PubMed] [Google Scholar]
  8. Clavel F., Guétard D., Brun-Vézinet F., Chamaret S., Rey M. A., Santos-Ferreira M. O., Laurent A. G., Dauguet C., Katlama C., Rouzioux C. Isolation of a new human retrovirus from West African patients with AIDS. Science. 1986 Jul 18;233(4761):343–346. doi: 10.1126/science.2425430. [DOI] [PubMed] [Google Scholar]
  9. Conley A. J., Gorny M. K., Kessler J. A., 2nd, Boots L. J., Ossorio-Castro M., Koenig S., Lineberger D. W., Emini E. A., Williams C., Zolla-Pazner S. Neutralization of primary human immunodeficiency virus type 1 isolates by the broadly reactive anti-V3 monoclonal antibody, 447-52D. J Virol. 1994 Nov;68(11):6994–7000. doi: 10.1128/jvi.68.11.6994-7000.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Conley A. J., Kessler J. A., 2nd, Boots L. J., Tung J. S., Arnold B. A., Keller P. M., Shaw A. R., Emini E. A. Neutralization of divergent human immunodeficiency virus type 1 variants and primary isolates by IAM-41-2F5, an anti-gp41 human monoclonal antibody. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3348–3352. doi: 10.1073/pnas.91.8.3348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dumitrescu O., Kalish M. L., Kliks S. C., Bandea C. I., Levy J. A. Characterization of human immunodeficiency virus type 1 isolates from children in Romania: identification of a new envelope subtype. J Infect Dis. 1994 Feb;169(2):281–288. doi: 10.1093/infdis/169.2.281. [DOI] [PubMed] [Google Scholar]
  12. Ebenbichler C., Westervelt P., Carrillo A., Henkel T., Johnson D., Ratner L. Structure-function relationships of the HIV-1 envelope V3 loop tropism determinant: evidence for two distinct conformations. AIDS. 1993 May;7(5):639–646. [PubMed] [Google Scholar]
  13. Freed E. O., Martin M. A. Evidence for a functional interaction between the V1/V2 and C4 domains of human immunodeficiency virus type 1 envelope glycoprotein gp120. J Virol. 1994 Apr;68(4):2503–2512. doi: 10.1128/jvi.68.4.2503-2512.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Geleziunas R., Bour S., Wainberg M. A. Cell surface down-modulation of CD4 after infection by HIV-1. FASEB J. 1994 Jun;8(9):593–600. doi: 10.1096/fasebj.8.9.8005387. [DOI] [PubMed] [Google Scholar]
  15. Gorny M. K., Conley A. J., Karwowska S., Buchbinder A., Xu J. Y., Emini E. A., Koenig S., Zolla-Pazner S. Neutralization of diverse human immunodeficiency virus type 1 variants by an anti-V3 human monoclonal antibody. J Virol. 1992 Dec;66(12):7538–7542. doi: 10.1128/jvi.66.12.7538-7542.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gorny M. K., Gianakakos V., Sharpe S., Zolla-Pazner S. Generation of human monoclonal antibodies to human immunodeficiency virus. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1624–1628. doi: 10.1073/pnas.86.5.1624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gorny M. K., Moore J. P., Conley A. J., Karwowska S., Sodroski J., Williams C., Burda S., Boots L. J., Zolla-Pazner S. Human anti-V2 monoclonal antibody that neutralizes primary but not laboratory isolates of human immunodeficiency virus type 1. J Virol. 1994 Dec;68(12):8312–8320. doi: 10.1128/jvi.68.12.8312-8320.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gorny M. K., Xu J. Y., Gianakakos V., Karwowska S., Williams C., Sheppard H. W., Hanson C. V., Zolla-Pazner S. Production of site-selected neutralizing human monoclonal antibodies against the third variable domain of the human immunodeficiency virus type 1 envelope glycoprotein. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3238–3242. doi: 10.1073/pnas.88.8.3238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gorny M. K., Xu J. Y., Karwowska S., Buchbinder A., Zolla-Pazner S. Repertoire of neutralizing human monoclonal antibodies specific for the V3 domain of HIV-1 gp120. J Immunol. 1993 Jan 15;150(2):635–643. [PubMed] [Google Scholar]
  20. Guyader M., Emerman M., Sonigo P., Clavel F., Montagnier L., Alizon M. Genome organization and transactivation of the human immunodeficiency virus type 2. Nature. 1987 Apr 16;326(6114):662–669. doi: 10.1038/326662a0. [DOI] [PubMed] [Google Scholar]
  21. Heynen C. A., Holzer T. J. Evaluation of a flow cytometric model for monitoring HIV antigen expression in vitro. J Immunol Methods. 1992 Jul 31;152(1):25–33. doi: 10.1016/0022-1759(92)90085-8. [DOI] [PubMed] [Google Scholar]
  22. Ho D. D., McKeating J. A., Li X. L., Moudgil T., Daar E. S., Sun N. C., Robinson J. E. Conformational epitope on gp120 important in CD4 binding and human immunodeficiency virus type 1 neutralization identified by a human monoclonal antibody. J Virol. 1991 Jan;65(1):489–493. doi: 10.1128/jvi.65.1.489-493.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jabbar M. A., Nayak D. P. Intracellular interaction of human immunodeficiency virus type 1 (ARV-2) envelope glycoprotein gp160 with CD4 blocks the movement and maturation of CD4 to the plasma membrane. J Virol. 1990 Dec;64(12):6297–6304. doi: 10.1128/jvi.64.12.6297-6304.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kang C. Y., Nara P., Chamat S., Caralli V., Ryskamp T., Haigwood N., Newman R., Köhler H. Evidence for non-V3-specific neutralizing antibodies that interfere with gp120/CD4 binding in human immunodeficiency virus 1-infected humans. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6171–6175. doi: 10.1073/pnas.88.14.6171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Karwowska S., Gorny M. K., Buchbinder A., Gianakakos V., Williams C., Fuerst T., Zolla-Pazner S. Production of human monoclonal antibodies specific for conformational and linear non-V3 epitopes of gp120. AIDS Res Hum Retroviruses. 1992 Jun;8(6):1099–1106. doi: 10.1089/aid.1992.8.1099. [DOI] [PubMed] [Google Scholar]
  26. Kim P. K., He S., HO J. L. Rapid culture and quantitation of human immunodeficiency virus type 1 from patient cells without the use of mitogen-stimulated donor cells. Clin Diagn Lab Immunol. 1994 Nov;1(6):660–666. doi: 10.1128/cdli.1.6.660-666.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Koot M., Vos A. H., Keet R. P., de Goede R. E., Dercksen M. W., Terpstra F. G., Coutinho R. A., Miedema F., Tersmette M. HIV-1 biological phenotype in long-term infected individuals evaluated with an MT-2 cocultivation assay. AIDS. 1992 Jan;6(1):49–54. doi: 10.1097/00002030-199201000-00006. [DOI] [PubMed] [Google Scholar]
  28. Louwagie J., Delwart E. L., Mullins J. I., McCutchan F. E., Eddy G., Burke D. S. Genetic analysis of HIV-1 isolates from Brazil reveals presence of two distinct genetic subtypes. AIDS Res Hum Retroviruses. 1994 May;10(5):561–567. doi: 10.1089/aid.1994.10.561. [DOI] [PubMed] [Google Scholar]
  29. Mascola J. R., Louwagie J., McCutchan F. E., Fischer C. L., Hegerich P. A., Wagner K. F., Fowler A. K., McNeil J. G., Burke D. S. Two antigenically distinct subtypes of human immunodeficiency virus type 1: viral genotype predicts neutralization serotype. J Infect Dis. 1994 Jan;169(1):48–54. doi: 10.1093/infdis/169.1.48. [DOI] [PubMed] [Google Scholar]
  30. McKeating J. A., Bennett J., Zolla-Pazner S., Schutten M., Ashelford S., Brown A. L., Balfe P. Resistance of a human serum-selected human immunodeficiency virus type 1 escape mutant to neutralization by CD4 binding site monoclonal antibodies is conferred by a single amino acid change in gp120. J Virol. 1993 Sep;67(9):5216–5225. doi: 10.1128/jvi.67.9.5216-5225.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Moore J. P., Ho D. D. Antibodies to discontinuous or conformationally sensitive epitopes on the gp120 glycoprotein of human immunodeficiency virus type 1 are highly prevalent in sera of infected humans. J Virol. 1993 Feb;67(2):863–875. doi: 10.1128/jvi.67.2.863-875.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Moore J. P., McCutchan F. E., Poon S. W., Mascola J., Liu J., Cao Y., Ho D. D. Exploration of antigenic variation in gp120 from clades A through F of human immunodeficiency virus type 1 by using monoclonal antibodies. J Virol. 1994 Dec;68(12):8350–8364. doi: 10.1128/jvi.68.12.8350-8364.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Moore J. P., Sattentau Q. J., Yoshiyama H., Thali M., Charles M., Sullivan N., Poon S. W., Fung M. S., Traincard F., Pinkus M. Probing the structure of the V2 domain of human immunodeficiency virus type 1 surface glycoprotein gp120 with a panel of eight monoclonal antibodies: human immune response to the V1 and V2 domains. J Virol. 1993 Oct;67(10):6136–6151. doi: 10.1128/jvi.67.10.6136-6151.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Muster T., Steindl F., Purtscher M., Trkola A., Klima A., Himmler G., Rüker F., Katinger H. A conserved neutralizing epitope on gp41 of human immunodeficiency virus type 1. J Virol. 1993 Nov;67(11):6642–6647. doi: 10.1128/jvi.67.11.6642-6647.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Nakamura G. R., Byrn R., Wilkes D. M., Fox J. A., Hobbs M. R., Hastings R., Wessling H. C., Norcross M. A., Fendly B. M., Berman P. W. Strain specificity and binding affinity requirements of neutralizing monoclonal antibodies to the C4 domain of gp120 from human immunodeficiency virus type 1. J Virol. 1993 Oct;67(10):6179–6191. doi: 10.1128/jvi.67.10.6179-6191.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ohlsson-Wilhelm B. M., Cory J. M., Kessler H. A., Eyster M. E., Rapp F., Landay A. Circulating human immunodeficiency virus (HIV) p24 antigen-positive lymphocytes: a flow cytometric measure of HIV infection. J Infect Dis. 1990 Nov;162(5):1018–1024. doi: 10.1093/infdis/162.5.1018. [DOI] [PubMed] [Google Scholar]
  37. Ohno T., Terada M., Yoneda Y., Shea K. W., Chambers R. F., Stroka D. M., Nakamura M., Kufe D. W. A broadly neutralizing monoclonal antibody that recognizes the V3 region of human immunodeficiency virus type 1 glycoprotein gp120. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10726–10729. doi: 10.1073/pnas.88.23.10726. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Osterhaus A. D., van Wezel A. L., van Steenis B., Drost G. A., Hazendonk T. G. Monoclonal antibodies to polioviruses. Production of specific monoclonal antibodies to the Sabin vaccine strains. Intervirology. 1981;16(4):218–224. doi: 10.1159/000149270. [DOI] [PubMed] [Google Scholar]
  39. Palker T. J., Matthews T. J., Clark M. E., Cianciolo G. J., Randall R. R., Langlois A. J., White G. C., Safai B., Snyderman R., Bolognesi D. P. A conserved region at the COOH terminus of human immunodeficiency virus gp120 envelope protein contains an immunodominant epitope. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2479–2483. doi: 10.1073/pnas.84.8.2479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Posner M. R., Cavacini L. A., Emes C. L., Power J., Byrn R. Neutralization of HIV-1 by F105, a human monoclonal antibody to the CD4 binding site of gp120. J Acquir Immune Defic Syndr. 1993 Jan;6(1):7–14. [PubMed] [Google Scholar]
  41. Potts K. E., Kalish M. L., Lott T., Orloff G., Luo C. C., Bernard M. A., Alves C. B., Badaro R., Suleiman J., Ferreira O. Genetic heterogeneity of the V3 region of the HIV-1 envelope glycoprotein in Brazil. Brazilian Collaborative AIDS Research Group. AIDS. 1993 Sep;7(9):1191–1197. doi: 10.1097/00002030-199309000-00007. [DOI] [PubMed] [Google Scholar]
  42. Roben P., Moore J. P., Thali M., Sodroski J., Barbas C. F., 3rd, Burton D. R. Recognition properties of a panel of human recombinant Fab fragments to the CD4 binding site of gp120 that show differing abilities to neutralize human immunodeficiency virus type 1. J Virol. 1994 Aug;68(8):4821–4828. doi: 10.1128/jvi.68.8.4821-4828.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Rübsamen-Waigmann H., von Briesen H., Holmes H., Björndal A., Korber B., Esser R., Ranjbar S., Tomlinson P., Galvao-Castro B., Karita E. Standard conditions of virus isolation reveal biological variability of HIV type 1 in different regions of the world. WHO Network for HIV Isolation and Characterization. AIDS Res Hum Retroviruses. 1994 Nov;10(11):1401–1408. doi: 10.1089/aid.1994.10.1401. [DOI] [PubMed] [Google Scholar]
  44. Sattentau Q. J., Moore J. P. Conformational changes induced in the human immunodeficiency virus envelope glycoprotein by soluble CD4 binding. J Exp Med. 1991 Aug 1;174(2):407–415. doi: 10.1084/jem.174.2.407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Sattentau Q. J., Zolla-Pazner S., Poignard P. Epitope exposure on functional, oligomeric HIV-1 gp41 molecules. Virology. 1995 Jan 10;206(1):713–717. doi: 10.1016/s0042-6822(95)80094-8. [DOI] [PubMed] [Google Scholar]
  46. Spear G. T., Takefman D. M., Sullivan B. L., Landay A. L., Zolla-Pazner S. Complement activation by human monoclonal antibodies to human immunodeficiency virus. J Virol. 1993 Jan;67(1):53–59. doi: 10.1128/jvi.67.1.53-59.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Wrin T., Loh T. P., Vennari J. C., Schuitemaker H., Nunberg J. H. Adaptation to persistent growth in the H9 cell line renders a primary isolate of human immunodeficiency virus type 1 sensitive to neutralization by vaccine sera. J Virol. 1995 Jan;69(1):39–48. doi: 10.1128/jvi.69.1.39-48.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Xu J. Y., Gorny M. K., Palker T., Karwowska S., Zolla-Pazner S. Epitope mapping of two immunodominant domains of gp41, the transmembrane protein of human immunodeficiency virus type 1, using ten human monoclonal antibodies. J Virol. 1991 Sep;65(9):4832–4838. doi: 10.1128/jvi.65.9.4832-4838.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. van Wezel A. L., Hazendonk A. G. Intratypic serodifferentiation of poliomyelitis virus strains by strain-specific antisera. Intervirology. 1979;11(1):2–8. doi: 10.1159/000149005. [DOI] [PubMed] [Google Scholar]

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

RESOURCES