Fluorescence correlation spectroscopy is a valuable tool in many scientific disciplines. In particular, such a spectroscopic technique has received a great deal of attention because of its remarkable potential for single-molecule detection. It is understood, however, that quantitative measurements can be considered reliable as long as molecular photophysics has been well characterized. To that end, molecular saturation and probe volume effects, which can worsen experimental accuracy, are treated here. These phenomena are adequately incorporated into the well-known three-dimensional Gaussian approximation by a novel method applied to interpret saturated fluorescence signals [Opt. Lett. 28, 2016 (2003)]. Comparisons with literature data are given to show the improvements of the suggested method compared with other approaches.

© 2004 Optical Society of America

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