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Fluorescence Correlation Spectroscopy (FCS)

Fluorescence Correlation Spectroscopy (FCS). Olga Pekar Helin Räägel EMBO Practical Course 2009 Heidelberg, DE. Why do FCS?. Study the kinetics of molecules in relation to: - their surrounding environment (temperature, viscosity, etc.) molecule size modified forms of the molecule.

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Fluorescence Correlation Spectroscopy (FCS)

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  1. Fluorescence Correlation Spectroscopy (FCS) Olga Pekar Helin Räägel EMBO Practical Course 2009 Heidelberg, DE

  2. Why do FCS? Study the kinetics of molecules in relation to: - their surrounding environment (temperature, viscosity, etc.) • molecule size • modified forms of the molecule.

  3. Theory behind FCS

  4. Dye dilution in water 1/N = 0,2 concentration of dye number of molecules = 5 4.15 x 10-16 L focal point volume a=1.5 µm b=0.25 µm a b

  5. FCS in live cells before after *theoretically diffusion time should increase by a factor of 41/3 = 1.58

  6. Advantages Measure the diffusion of molecules Transfected cells can be used Very small sample volumes Single molecule sensitivity Provides direct measure of stoichiometry Disadvantages Only cells with low expression of the fluorescently tagged protein can be used movement of cells during measurement Need for special equipment FCStechnique

  7. Thank You: Nicolas Jörg Oleg

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