Dual-color total internal reflection fluorescence cross-correlation spectroscopy

Marcel Leutenegger; Michael Gösch; Rainer Andreas Leitgeb; Theo Lasser; Hans Blom; Jerker Widengren; Christian Eggeling

doi:10.1117/1.2221714

1 July 2006 Dual-color total internal reflection fluorescence cross-correlation spectroscopy

Marcel Leutenegger, Michael Gösch, Rainer Andreas Leitgeb, Theo Lasser, Hans Blom, Jerker Widengren, Christian Eggeling

Author Affiliations +

Journal of Biomedical Optics, Vol. 11, Issue 4, 040502 (July 2006). https://doi.org/10.1117/1.2221714

Abstract

We present the development and first application of a novel dual-color total internal reflection (TIR) fluorescence system for single-molecule coincidence analysis and fluorescence cross-correlation spectroscopy (FCCS). As a performance analysis, we measured a synthetic DNA-binding assay, demonstrating this dual-color TIR-FCCS approach to be a suitable method for measuring coincidence assays such as biochemical binding, fusion, or signal transduction at solid/liquid interfaces. Due to the very high numerical aperture of the epi-illumination configuration, our setup provides a very high fluorescence collection efficiency resulting in a two- to three-fold increase in molecular brightness compared to conventional confocal FCCS. Further improvements have been achieved through global analysis of the spectroscopic data.

©(2006) Society of Photo-Optical Instrumentation Engineers (SPIE)

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Marcel Leutenegger, Michael Gösch, Rainer Andreas Leitgeb, Theo Lasser, Hans Blom, Jerker Widengren, and Christian Eggeling "Dual-color total internal reflection fluorescence cross-correlation spectroscopy," Journal of Biomedical Optics 11(4), 040502 (1 July 2006). https://doi.org/10.1117/1.2221714

Published: 1 July 2006

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KEYWORDS

Luminescence

Fluorescence spectroscopy

Spectroscopy

Molecules

Reflectance spectroscopy

Interfaces

Confocal microscopy

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