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

TitleSingle-molecule immunosorbent assay as a tool for human immunodeficiency virus-1 antigen detection
Publication TypeJournal Article
Year of Publication2009
AuthorsLi JW, Xie WJ, Fang N, Yeung ES
Journal TitleAnalytical and Bioanalytical Chemistry
Volume394
Pages489-497
Date Published05
ISBN Number1618-2642
Accession NumberISI:000265388400012
Keywordsantibody-affinity, diagnostics, DNA, early diagnosis, fluorescence detection, health-care workers, hiv, hiv-1, immunoassay, microarrays, nanopores, single-molecule immunosorbent assay (smisa)
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.
URL<Go to ISI>://000265388400012
DOI10.1007/S00216-009-2712-1