2010 AACC MEETING, LOS ANGELES, CA
D.C. Duffy1, D. M. Rissin1, C. W. Kan1, T. G. Campbell1, S. C. Howes1, D. R. Fournier1, L. Song1, T. Piech1, P. P. Patel1, L. Chang1, A. J. Rivnak1, E. P. Ferrell1, J. D. Randall1, G. K. Provuncher1, D. R. Walt2. 1Quanterix Corporation, Cambridge, MA, 2Tufts University, Medford, MA.
Objective:
The aim of this study was to detect prostate specific antigen (PSA) in the serum of patients who had undergone radical prostatectomy (RP). To achieve this objective, an ultra–sensitive ELISA for PSA based on single molecule detection was developed and evaluated.
Methodology:
We have developed a method for detecting single immunocomplexes formed in the enzyme–linked immunosorbent assay (ELISA) using single molecule arrays (SiMoA). This digital ELISA method is based on isolating single immunocomplexes labeled with an enzyme in arrays of femtoliter wells, sealing the arrays in the presence of the enzyme substrate, and fluorescently imaging the array. Fluorescent product molecules of the enzyme–substrate reaction are confined in the femtoliter volume, giving rise to a local high concentration that can be easily detected using a standard fluorescent microscope. By using high density arrays of femtoliter wells, hundreds to thousands of single immunocomplexes can be detected simultaneously. Isolation of single immunocomplexes using SiMoA gives rise to a dramatic increase in sensitivity over bulk, ensemble detection methods. An ultra–sensitive digital ELISA for detecting PSA was developed that has a limit of detection (LOD) of 6 fg/mL (200 aM) in serum. This assay was used to measure PSA in the sera of thirty RP patients.
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