2022/03/25 by Damir Sakhapov, Sakhapov, Damir, Ingo Gregor +5
Biochemistry, Genetics and Molecular Biology · Chemistry · #Advanced Fluorescence Microscopy Techniques #Chemical Physics (physics.chem-ph) #Electrochemical Analysis and Applications #FOS: Physical sciences #Photochemistry and Electron Transfer Studies
paper · pdf · doi:10.48550/arxiv.2203.13480
openalex publication_date 2022/03/25 · openalex created_date 2022/04/03 · openalex updated_date 2026/07/28
We present a new method that combines fluorescence correlation spectroscopy (FCS) on the microsecond time scale with fluorescence antibunching measurements on the nanosecond time scale for measuring photophysical rate constants of fluorescent molecules. The antibunching measurements allow us to quantify the average excitation rate of fluorescent molecules within the confocal detection volume of the FCS measurement setup. Knowledge of this value allows us then to quantify, in an absolute manner, the intersystem crossing rate and triplet state lifetime from the microsecond temporal decay of the FCS curves. We present a theoretical analysis of the method and estimate the maximum bias caused by the averaging of all quantities (excitation rate, photophysical rates) over the confocal detection volume, and we show that this bias is smaller than 5% in most cases. We apply the method for measuring the photophysical rate constants of the widely used dyes Rhodamine~110 and ATTO~655.