Real-Time G-Protein-Coupled Receptor Imaging to Understand and Quantify Receptor Dynamics
Figure 3
Photobleaching correction. D2LR-YFP-transfected HEK-293 cells were recorded every second along 6 min. One plane from the stack was selected for analysis and fluorescence intensity on the plasma membrane was measured at each time point. (a) The effect of fluorescence intensity fading on the plasma membrane was clearly visible due to repetitive laser exposition. (b) Photobleaching effect was corrected in each frame for each pixel using a biexponential equation. (c) The temporal evolution of fluorescence intensity of a single pixel (blue) followed the biexponential decay (red). The indicated biexponential equation (see Section 2.7.1) was used to correct the intensity value of this pixel along time (green). (d) Graphical results of the analysis showing images prior (upper row) and after correction (lower row); note that the fading is only visible in the upper row.
