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Fluorescence correlation spectroscopy example: shift of autocorrelation curve.

Masataka Kinjo, Hiroshi Sakata, Shintaro Mikuni

Cold Spring Harbor Protocols

|October 5, 2011

Summary

Fluorescence correlation spectroscopy (FCS) analyzes molecular interactions by monitoring fluorescence fluctuations. This method provides precise physical parameters, including diffusion times and molecular counts, in aqueous conditions.

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Area of Science:

Biochemistry
Biophysics
Physical Chemistry

Background:

Fluorescence correlation spectroscopy (FCS) is a sensitive technique for studying molecular dynamics.
It measures fluorescence fluctuations in a small observation volume to extract physical parameters.
FCS is valuable for investigating molecular interactions in solution.

Purpose of the Study:

To outline a Fluorescence Correlation Spectroscopy (FCS) protocol.
To demonstrate an example of FCS data analysis involving a shift in the autocorrelation curve.
To highlight the utility of FCS in determining molecular properties and interactions.

Main Methods:

Monitoring fluorescence emission intensity fluctuations over time.
Calculating the autocorrelation function from fluorescence signals.
Fitting the autocorrelation curves to physical models to extract parameters.

Main Results:

FCS directly yields parameters like average molecular number and diffusion time.
The technique provides insights into molecular brightness.
Analysis of autocorrelation curve shifts can reveal specific molecular behaviors or interactions.

Conclusions:

FCS is a powerful tool for quantitative analysis of molecular systems.
The method allows for precise determination of molecular interactions in aqueous environments.
Protocol details for FCS experiments, including autocorrelation curve analysis, are essential for accurate interpretation.