Analytical and Bioanalytical Chemistry

, Volume 394, Issue 2, pp 489–497 | Cite as

Single-molecule immunosorbent assay as a tool for human immunodeficiency virus-1 antigen detection

  • Jiangwei Li
  • Wenjun Xie
  • Ning Fang
  • Edward S. Yeung
Original Paper

Abstract

Ultrasensitive detection and quantification of viral antigen with a novel single-molecule immunosorbent assay (SMISA) was achieved. Antigen from human immunodeficiency virus type 1 (HIV-1), the major etiological agent of acquired immune deficiency syndrome, served as the screening target in this study. The target molecule was sandwiched between a polyclonal capture antibody and a monoclonal detector antibody. The capture antibody was covalently immobilized on (3-glycidoxypropyl) trimethoxy silane-modified glass slides. The detector antibody was conjugated with fluorescent Alexa Fluor 532 labeled secondary antibody prior to being used as a probe for the antigen. Imaging was performed with a total internal reflection fluorescence single-molecule detection system. This technique is demonstrated for detecting HIV-1 p24 antigen down to 0.1 pg/mL with a dynamic range of over four orders of magnitude. A Langmuir isotherm fits the molecule count dependence on the target concentration. The target antigen was further tested in 20% human serum, and the results showed that neither sensitivity nor dynamic range was affected by the biological matrix. SMISA is therefore a promising approach for the early diagnosis of viral induced diseases.

Keywords

Single-molecule immunosorbent assay (SMISA) Early diagnosis HIV-1 Fluorescence detection 

Notes

Acknowledgments

We thank Dr. Mary Jo Schmerr and Dr. Gufeng Wang for valuable discussions. E.S.Y. thanks the Robert Allen Wright Endowment for Excellence for support. The Ames Laboratory is operated for the U.S. Department of Energy by Iowa State University under contract no. DE-AC02-07CH11358. This work was supported by the Director of Science, Office of Basic Energy Science, Division of Chemical Sciences.

References

  1. 1.
    Howorka S, Cheley S, Bayley H (2001) Nat Biotechnol 19:636–639CrossRefGoogle Scholar
  2. 2.
    Ma YF, Shortreed MR, Li HL, Huang WH, Yeung ES (2001) Electrophoresis 22:421–426CrossRefGoogle Scholar
  3. 3.
    Wabuyele MB, Farquar H, Stryjewski W, Hammer RP, Soper SA, Cheng YW, Barany F (2003) J Am Chem Soc 125:6937–6945CrossRefGoogle Scholar
  4. 4.
    Storhoff JJ, Lucas AD, Garimella V, Bao YP, Muller UR (2004) Nat Biotechnol 22:883–887CrossRefGoogle Scholar
  5. 5.
    Li J, Lee JY, Yeung ES (2006) Anal Chem 78:6490–6496CrossRefGoogle Scholar
  6. 6.
    Lee JY, Li J, Yeung ES (2007) Anal Chem 79:8083–8089CrossRefGoogle Scholar
  7. 7.
    Jain KK (2007) Clin Chem 53:2002–2009CrossRefGoogle Scholar
  8. 8.
    Seidel M, Niessner R (2008) Anal Bioanal Chem 391:1521–1544CrossRefGoogle Scholar
  9. 9.
    Chan WCW, Nie SM (1998) Science 281:2016–2018CrossRefGoogle Scholar
  10. 10.
    Bieschke J, Giese A, Schulz-Schaeffer W, Zerr I, Poser S, Eigen M, Kretzschmar H (2000) Proc Natl Acad Sci USA 97:5468–5473CrossRefGoogle Scholar
  11. 11.
    Tetin SY, Swift KM, Matayoshi ED (2002) Anal Biochem 307:84–91CrossRefGoogle Scholar
  12. 12.
    Tetin SY, Stroupe SD (2004) Curr Pharm Biotechnol 5:9–16CrossRefGoogle Scholar
  13. 13.
    Nalefski EA, D'Antoni CM, Ferrell EP, Lloyd JA, Qiu HQ, Harris JL, Whitney DH (2006) Clin Chem 52:2172–2175CrossRefGoogle Scholar
  14. 14.
    Qiu H, Ferrell EP, Nolan N, Phelps BH, Tabibiazar R, Whitney DH, Nalefski EA (2007) Clin Chem 53:2010–2012CrossRefGoogle Scholar
  15. 15.
    Han A, Creus M, Schurmann G, Linder V, Ward TR, de Rooij NF, Staufer U (2008) Anal Chem 80:4651–4658CrossRefGoogle Scholar
  16. 16.
    Weiss RA (1993) Science 260:1273–1279CrossRefGoogle Scholar
  17. 17.
    Dybul M, Fauci AS, Bartlett JG, Kaplan JE, Pau AK (2002) Ann Intern Med 137:381–433Google Scholar
  18. 18.
    Saville RD, Constantine NT, Cleghorn FR, Jack N, Bartholomew C, Edwards J, Gomez P, Blattner WA (2001) J Clin Microbiol 39:2518–2524CrossRefGoogle Scholar
  19. 19.
    (1996) MMWR. Morb Mortal Wkly Rep 45:468-480Google Scholar
  20. 20.
    (1998) MMWR Recomm Rep 47:1-33Google Scholar
  21. 21.
    (2001) MMWR Recomm Rep 50:1-52Google Scholar
  22. 22.
    Cardo DM, Culver DH, Ciesielski CA, Srivastava PU, Marcus R, Abiteboul D, Heptonstall J, Ippolito G, Lot F, McKibben P, Bell DM (1997) N Engl J Med 337:1485–1490CrossRefGoogle Scholar
  23. 23.
    Wang SA, Panlilio AL, Doi PA, White AD, Stek M, Saah A, Grp HPR (2000) Infect Control Hosp Epidemiol 21:780–785CrossRefGoogle Scholar
  24. 24.
    Panlilio AL, Cardo DM, Grohskopf LA, Heneine W, Ross CS (2005) MMWR Recomm Rep 54:1–17Google Scholar
  25. 25.
    Reeves JD, Doms RW (2002) J Gen Virol 83:1253–1265Google Scholar
  26. 26.
    Kusnezow W, Jacob A, Walijew A, Diehl F, Hoheisel JD (2003) Proteomics 3:254–264CrossRefGoogle Scholar
  27. 27.
    He Y, Li HW, Yeung ES (2005) J Phys Chem B 109:8820–8832CrossRefGoogle Scholar
  28. 28.
    Fang N, Zhang H, Li J, Li HW, Yeung ES (2007) Anal Chem 79:6047–6054CrossRefGoogle Scholar
  29. 29.
    Bhatia SK, Shriverlake LC, Prior KJ, Georger JH, Calvert JM, Bredehorst R, Ligler FS (1989) Anal Biochem 178:408–413CrossRefGoogle Scholar
  30. 30.
    Temirov JP, Bradbury ARM, Werner JH (2008) Anal Chem 80:8642–8648CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Jiangwei Li
    • 1
  • Wenjun Xie
    • 1
  • Ning Fang
    • 1
  • Edward S. Yeung
    • 1
  1. 1.Ames Laboratory-USDOE and Department of ChemistryIowa State UniversityAmesUSA

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