Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1998 Sep 1;26(17):3915–3924. doi: 10.1093/nar/26.17.3915

Staining of cell surface human CD4 with 2'-F-pyrimidine-containing RNA aptamers for flow cytometry.

K A Davis 1, Y Lin 1, B Abrams 1, S D Jayasena 1
PMCID: PMC147797  PMID: 9705498

Abstract

We have used recombinant human CD4 presented on beads as an affinity matrix to screen a 2'-F-pyrimidine-containing RNA library with a complexity of approximately 10(14) molecules. Affinity-selected aptamers bind recombinant CD4 with low nanomolar equilibrium dissociation constants. These high-affinity aptamers conjugated to different fluorophores such as fluorescein and phycoerythrin were used to stain cells, expressing human CD4 on cell surface, for analysis by flow cytometry. Aptamers, conjugated to fluorophores, stained mouse T cells that express human CD4 on the surface, but not the control mouse T cells lacking human CD4. The control cells, however, do express mouse CD4 whose extracellular domain has 55% sequence identity to the human form. These human CD4-specific aptamers selectively stained CD4(+) T cells in a preparation of human peripheral blood mononuclear cells. These results and others suggest that aptamers are emerging as a versatile class of molecules that can be used for various diagnostic applications performed under different formats or platforms.

Full Text

The Full Text of this article is available as a PDF (579.6 KB).

Selected References

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

  1. Burgin A. B., Pace N. R. Mapping the active site of ribonuclease P RNA using a substrate containing a photoaffinity agent. EMBO J. 1990 Dec;9(12):4111–4118. doi: 10.1002/j.1460-2075.1990.tb07633.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Dalgleish A. G., Beverley P. C., Clapham P. R., Crawford D. H., Greaves M. F., Weiss R. A. The CD4 (T4) antigen is an essential component of the receptor for the AIDS retrovirus. Nature. 1984 Dec 20;312(5996):763–767. doi: 10.1038/312763a0. [DOI] [PubMed] [Google Scholar]
  3. Dang C., Jayasena S. D. Oligonucleotide inhibitors of Taq DNA polymerase facilitate detection of low copy number targets by PCR. J Mol Biol. 1996 Nov 29;264(2):268–278. doi: 10.1006/jmbi.1996.0640. [DOI] [PubMed] [Google Scholar]
  4. Davis K. A., Abrams B., Lin Y., Jayasena S. D. Use of a high affinity DNA ligand in flow cytometry. Nucleic Acids Res. 1996 Feb 15;24(4):702–706. doi: 10.1093/nar/24.4.702. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. De Wolf F., Roos M., Lange J. M., Houweling J. T., Coutinho R. A., van der Noordaa J., Schellekens P. T., Goudsmit J. Decline in CD4+ cell numbers reflects increase in HIV-1 replication. AIDS Res Hum Retroviruses. 1988 Dec;4(6):433–440. doi: 10.1089/aid.1988.4.433. [DOI] [PubMed] [Google Scholar]
  6. Doyle C., Strominger J. L. Interaction between CD4 and class II MHC molecules mediates cell adhesion. Nature. 1987 Nov 19;330(6145):256–259. doi: 10.1038/330256a0. [DOI] [PubMed] [Google Scholar]
  7. Drolet D. W., Moon-McDermott L., Romig T. S. An enzyme-linked oligonucleotide assay. Nat Biotechnol. 1996 Aug;14(8):1021–1025. doi: 10.1038/nbt0896-1021. [DOI] [PubMed] [Google Scholar]
  8. Eaton B. E., Gold L., Zichi D. A. Let's get specific: the relationship between specificity and affinity. Chem Biol. 1995 Oct;2(10):633–638. doi: 10.1016/1074-5521(95)90023-3. [DOI] [PubMed] [Google Scholar]
  9. Gold L. Oligonucleotides as research, diagnostic, and therapeutic agents. J Biol Chem. 1995 Jun 9;270(23):13581–13584. doi: 10.1074/jbc.270.23.13581. [DOI] [PubMed] [Google Scholar]
  10. Hicke B. J., Watson S. R., Koenig A., Lynott C. K., Bargatze R. F., Chang Y. F., Ringquist S., Moon-McDermott L., Jennings S., Fitzwater T. DNA aptamers block L-selectin function in vivo. Inhibition of human lymphocyte trafficking in SCID mice. J Clin Invest. 1996 Dec 15;98(12):2688–2692. doi: 10.1172/JCI119092. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Jellinek D., Green L. S., Bell C., Lynott C. K., Gill N., Vargeese C., Kirschenheuter G., McGee D. P., Abesinghe P., Pieken W. A. Potent 2'-amino-2'-deoxypyrimidine RNA inhibitors of basic fibroblast growth factor. Biochemistry. 1995 Sep 12;34(36):11363–11372. doi: 10.1021/bi00036a009. [DOI] [PubMed] [Google Scholar]
  12. Kwong P. D., Ryu S. E., Hendrickson W. A., Axel R., Sweet R. M., Folena-Wasserman G., Hensley P., Sweet R. W. Molecular characteristics of recombinant human CD4 as deduced from polymorphic crystals. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6423–6427. doi: 10.1073/pnas.87.16.6423. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lin Y., Jayasena S. D. Inhibition of multiple thermostable DNA polymerases by a heterodimeric aptamer. J Mol Biol. 1997 Aug 8;271(1):100–111. doi: 10.1006/jmbi.1997.1165. [DOI] [PubMed] [Google Scholar]
  14. Lin Y., Nieuwlandt D., Magallanez A., Feistner B., Jayasena S. D. High-affinity and specific recognition of human thyroid stimulating hormone (hTSH) by in vitro-selected 2'-amino-modified RNA. Nucleic Acids Res. 1996 Sep 1;24(17):3407–3414. doi: 10.1093/nar/24.17.3407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lin Y., Qiu Q., Gill S. C., Jayasena S. D. Modified RNA sequence pools for in vitro selection. Nucleic Acids Res. 1994 Dec 11;22(24):5229–5234. doi: 10.1093/nar/22.24.5229. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Littman D. R. The structure of the CD4 and CD8 genes. Annu Rev Immunol. 1987;5:561–584. doi: 10.1146/annurev.iy.05.040187.003021. [DOI] [PubMed] [Google Scholar]
  17. Maddon P. J., Dalgleish A. G., McDougal J. S., Clapham P. R., Weiss R. A., Axel R. The T4 gene encodes the AIDS virus receptor and is expressed in the immune system and the brain. Cell. 1986 Nov 7;47(3):333–348. doi: 10.1016/0092-8674(86)90590-8. [DOI] [PubMed] [Google Scholar]
  18. Maddon P. J., Littman D. R., Godfrey M., Maddon D. E., Chess L., Axel R. The isolation and nucleotide sequence of a cDNA encoding the T cell surface protein T4: a new member of the immunoglobulin gene family. Cell. 1985 Aug;42(1):93–104. doi: 10.1016/s0092-8674(85)80105-7. [DOI] [PubMed] [Google Scholar]
  19. Maddon P. J., Molineaux S. M., Maddon D. E., Zimmerman K. A., Godfrey M., Alt F. W., Chess L., Axel R. Structure and expression of the human and mouse T4 genes. Proc Natl Acad Sci U S A. 1987 Dec;84(24):9155–9159. doi: 10.1073/pnas.84.24.9155. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. O'Connell D., Koenig A., Jennings S., Hicke B., Han H. L., Fitzwater T., Chang Y. F., Varki N., Parma D., Varki A. Calcium-dependent oligonucleotide antagonists specific for L-selectin. Proc Natl Acad Sci U S A. 1996 Jun 11;93(12):5883–5887. doi: 10.1073/pnas.93.12.5883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Pieken W. A., Olsen D. B., Benseler F., Aurup H., Eckstein F. Kinetic characterization of ribonuclease-resistant 2'-modified hammerhead ribozymes. Science. 1991 Jul 19;253(5017):314–317. doi: 10.1126/science.1857967. [DOI] [PubMed] [Google Scholar]
  22. Uphoff K. W., Bell S. D., Ellington A. D. In vitro selection of aptamers: the dearth of pure reason. Curr Opin Struct Biol. 1996 Jun;6(3):281–288. doi: 10.1016/s0959-440x(96)80045-5. [DOI] [PubMed] [Google Scholar]
  23. Wang J. H., Yan Y. W., Garrett T. P., Liu J. H., Rodgers D. W., Garlick R. L., Tarr G. E., Husain Y., Reinherz E. L., Harrison S. C. Atomic structure of a fragment of human CD4 containing two immunoglobulin-like domains. Nature. 1990 Nov 29;348(6300):411–418. doi: 10.1038/348411a0. [DOI] [PubMed] [Google Scholar]
  24. Wood G. S., Warner N. L., Warnke R. A. Anti-Leu-3/T4 antibodies react with cells of monocyte/macrophage and Langerhans lineage. J Immunol. 1983 Jul;131(1):212–216. [PubMed] [Google Scholar]
  25. Wright A., Shin S. U., Morrison S. L. Genetically engineered antibodies: progress and prospects. Crit Rev Immunol. 1992;12(3-4):125–168. [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES