Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments

Yahiatene I, Doose S, Huser T, Sauer M (2012)
Analytical Chemistry 84(6): 2729-2736.

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Abstract
We introduce a robust and relatively easy-to-use method to evaluate the quality of two-color (or more) fluorescence coincidence measurements based on close investigation of the coincidence correlation-matrix. This matrix contains temporal correlations between the number of detected bursts in individual channels and their coincidences. We show that the Euclidian norm of a vector Gamma derived from elements of the correlation matrix takes a value between 0 and 2 depending on the relative coincidence frequency. We characterized the Gamma-norm and its dependence on various experimental conditions by computer simulations and fluorescence microscopy experiments. Single-molecule experiments with two differently colored dye molecules diffusing freely in aqueous solution, a sample that generates purely random coincidence events, return a Gamma-norm less than one, depending on the concentration of the fluorescent dyes. As perfect coincidence sample we monitored broad autofluorescence of 2.8 mu m beads and determined the F-norm to be maximal and close to two. As in realistic diagnostic applications, we show that two-color coincidence detection of single-stranded DNA molecules, using differently labeled Molecular Beacons hybridizing to the same target, reveal a value between one and two representing a mixture of an optimal coincidence sample and a sample generating random coincidences. The Gamma-norm introduced for data analysis provides a quantifiable measure for quickly judging the outcome of single-molecule coincidence experiments and estimating the quality of detected coincidences.
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Yahiatene I, Doose S, Huser T, Sauer M. Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments. Analytical Chemistry. 2012;84(6):2729-2736.
Yahiatene, I., Doose, S., Huser, T., & Sauer, M. (2012). Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments. Analytical Chemistry, 84(6), 2729-2736.
Yahiatene, I., Doose, S., Huser, T., and Sauer, M. (2012). Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments. Analytical Chemistry 84, 2729-2736.
Yahiatene, I., et al., 2012. Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments. Analytical Chemistry, 84(6), p 2729-2736.
I. Yahiatene, et al., “Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments”, Analytical Chemistry, vol. 84, 2012, pp. 2729-2736.
Yahiatene, I., Doose, S., Huser, T., Sauer, M.: Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments. Analytical Chemistry. 84, 2729-2736 (2012).
Yahiatene, Idir, Doose, Soeren, Huser, Thomas, and Sauer, Markus. “Correlation-Matrix Analysis of Two-Color Coincidence Events in Single-Molecule Fluorescence Experiments”. Analytical Chemistry 84.6 (2012): 2729-2736.
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