Fluorescence Correlation Spectroscopy

Fluorescence Correlation Spectroscopy

Fluorescence correlation spectroscopy (FCS) is a technology to measure the fluorescence fluctuation of molecular systems. It has been widely used in biomedicine, biophysics, chemistry and other fields. FCS can measure the fluctuation of fluorescence intensity caused by Brownian motion or chemical reaction in a small volume, and then quantitatively evaluate the concentration, diffusion coefficient and interaction of fluorescent molecules in a simple artificial model or living cells.

Principle, instrumentation, and data analysis of FCSFigure 1. Principle, instrumentation, and data analysis of FCS. (A) Schematic view of a confocal based FCS setup. (B) The intensity trace collected from the observation volume. (C) The autocorrelation curve calculated with the intensity trace in Figure 1B (Yu L S, et al. 2021).

Principles of Fluorescence Correlation Spectroscopy

In an equilibrium solution, the number of fluorescent molecules in a micro region is determined by the concentration of fluorescent molecules in the solution and the volume of the region. When fluorescent molecules diffuse due to Brownian motion or intermolecular chemical reaction, the fluorescence intensity in the micro region will change with time, and the phenomenon of fluorescence fluctuation will occur. FCS is a fluorescence spectroscopy technology that uses this phenomenon for analysis and detection.

FCS is usually performed in a confocal system, where fluorescence emitted by fluorescently labeled molecules within the observation volume is collected by the same objective and propagates in the opposite direction to the excitation/depletion light direction. After passing through the longpass dichroic mirror, the fluorescence is focused through the pinhole onto the avalanche photodiode. Fluorescence emitted in the out-of-focus region is rejected by the pinhole and it does not reach the detector. Therefore, the pinhole reduces the axial extension of the viewing volume. This restricted observation volume significantly improves the signal-to-noise ratio and also reduces the measurement time required to obtain a good correlation curve.

CD BioSciences is a biotechnology company, which has been committed to the development of fluorescent probes and imaging technology for many years. We can use fluorescence correlation spectroscopy to detect molecular concentrations, hydrodynamic radii, diffusion kinetics and interactions of different biomolecules. If you have any needs, please feel free to contact us.

Type of Fluorescence Correlation Spectroscopy

Dual-color FCCS
Pair correlation function (pCF) based FCS
Inverse-FCS
SPIM-FCS

Multi-focus FCS
Scanning FCS
Focus-reduced FCS
STED-FCS

Application of Fluorescence Correlation Spectroscopy

Nucleic acid breakage and nucleic acid hybridization
Molecular diffusion in the nucleus and cytoplasm
Binding equilibrium of drugs and other low molecular weight ligands
Lateral separation of lipids in double membranes
Protein-protein interactions
Aggregation of membrane-bound receptors
Various biological processes in cells

Advantages of Fluorescence Correlation Spectroscopy

  • High spatial and temporal resolution, short analysis times and high sensitivity
  • Measures the spatial and temporal correlation of individual molecules with themselves
  • Avoidance of artifacts caused by photobleaching and membrane motion
  • Experienced scientists provide experimental consultation
  • Reasonable price and short turnaround time

References

  1. Yu L, Lei Y, Ma Y, et al. A comprehensive review of fluorescence correlation spectroscopy[J]. Frontiers in Physics, 2021, 9.
  2. Elson E L. Fluorescence correlation spectroscopy: past, present, future[J]. Biophysical Journal, 2011, 101(12): 2855-2870.

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Please note: Our services can only be used for research purposes. Do not use in diagnostic or therapeutic procedures!

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