Analytical and Bioanalytical Chemistry

, Volume 394, Issue 2, pp 489–497

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

DOI: 10.1007/s00216-009-2712-1

Cite this article as:
Li, J., Xie, W., Fang, N. et al. Anal Bioanal Chem (2009) 394: 489. doi:10.1007/s00216-009-2712-1
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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 diagnosisHIV-1Fluorescence detection

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