Measuring Membrane Dipole Potential With Di-8-ANEPPS

Results Summary

Effect of acute cholesterol depletion on membrane dipole potential. (a) Representative confocal micrographs of CHO-K1 cells labeled with di-8-ANEPPS (λex = 458 nm (upper panel), and λex = 514 nm (lower panel), the emission bandpass being 650–710 nm in both cases) with increasing concentrations of MβCD. The fluorescence intensity of images was corrected for laser power at two different excitation wavelengths (458 and 514 nm). (b) The corresponding fluorescence intensity ratio (R) map (color-coded in a scale of 0.8–3) under these conditions. R is defined as the ratio of fluorescence intensities at an excitation wavelength of 458 nm to that at 514 nm (emission band pass at 650–710 nm in both cases) and was calculated using ImageJ (NIH, Bethesda, MD). (c) Effect of increasing concentration of MβCD on the R-value (means ± S.E.), averaged over at least fifteen independent measurements (*** corresponds to significant (p < 0.001) difference in R). The inset shows the chemical structure of di-8-ANEPPS. The scale bar indicates 10 μm.