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Fluorescence

Fluorescence. Energy diagram. Energy diagram. Quantum Yield - Q. Rate of depopulation. Red shift. Fluorimeter. Time resolved experiment . Spectrofluorimeter. Frequency domain (phase/modulation) . Excitation/ emmission spectrum :. Fluorophores. Intrinsic fluorophores. Extrinsic probes.

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Fluorescence

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  1. Fluorescence Biophysical Tools '04 - Fluorescence

  2. Energy diagram Energy diagram Quantum Yield - Q Rate of depopulation Red shift Biophysical Tools '04 - Fluorescence

  3. Fluorimeter Time resolved experiment Spectrofluorimeter Frequency domain (phase/modulation) Excitation/emmission spectrum: Biophysical Tools '04 - Fluorescence

  4. Fluorophores Intrinsic fluorophores Extrinsic probes Solvent Effects Biophysical Tools '04 - Fluorescence

  5. Iװ I┴ Anisotropy Time resolved anisotropy Anisotropy Motional depolarization Biophysical Tools '04 - Fluorescence

  6. Phosphorescence Phosphorescence Øtriplet probes: eosin, erythrosin S T Biophysical Tools '04 - Fluorescence

  7. Anisotropic motion Hindered rotation Perrin Plot A(t) A∞ Biophysical Tools '04 - Fluorescence

  8. Quenching the rate of quenching = k[Q] Stern-Volmer plots 2 Fluor (no Q) / fluor (Q=x) 1 Quencher conc. Biophysical Tools '04 - Fluorescence

  9. 5 donor 4 donor/aceptor donor (AEDANS) 3 Intensity 2 acceptor 1 acceptor (AF) 0 500 700 400 600 Wavelength (nm) FRET Efficiency of transfer: E = rate of transfer /total rate of relaxation = E = kFET/(k + kFET ) = 1-tAD/tD = 1- FAD/FD dipole-dipole interactions kFET  (R-3)2 ~ 1/R6 Ro distance @ E=0.5 i.e. when kFET = 1/t then: E = R-6/(Ro-6 + R-6) Orientation problem: k2 Biophysical Tools '04 - Fluorescence

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