Analyzing Receptor Assemblies in the Cell Membrane Using Fluorescence Anisotropy Imaging with TIRF Microscopy
Figure 5
Red-edge excitation suppresses homo-FRET.
(a) Excitation and emission spectra of mTFP1 with the applied excitation changed to the 488 nm laser line and with a narrow-range emission filter. In this situation, the red-edge detection suppresses homo-FRET when the spectral overlap between the excitation and emission spectra of mTFP1 is diminished. (b) Red-edge anisotropy measurement in the TIRF mode shows no loss of anisotropy upon agonist addition, demonstrating that the change of rotational diffusion is negligible, and the anisotropy change is the result of increased homo-FRET upon dimerization.