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J Biomol Screen. 2003 Feb;8(1):19-33.

Single-molecule detection technologies in miniaturized high-throughput screening: fluorescence intensity distribution analysis.

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  • 1GlaxoSmithKline, Direvo, Cologne, Germany.


Single-molecule detection technologies are becoming a powerful readout format to support ultra-high-throughput screening. These methods are based on the analysis of fluorescence intensity fluctuations detected from a small confocal volume element. The fluctuating signal contains information about the mass and brightness of the different species in a mixture. The authors demonstrate a number of applications of fluorescence intensity distribution analysis (FIDA), which discriminates molecules by their specific brightness. Examples for assays based on brightness changes induced by quenching/dequenching of fluorescence, fluorescence energy transfer, and multiple-binding stoichiometry are given for important drug targets such as kinases and proteases. FIDA also provides a powerful method to extract correct biological data in the presence of compound fluorescence.

[PubMed - indexed for MEDLINE]
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