FIG. 4.

Increase in fluorescence of cell-bound NBD-labeled PA G305C with time as recorded by confocal microscopy. (A) CHO-R1 ATR800 cells were plated overnight, placed on ice, and incubated with a 1:1 mixture of NBD-labeled PA G305C and Alexa 546-labeled PA K563C. After an hour, the cells were transferred to a humidified 37°C CO₂ incubator, and samples were removed at 10 min, 30 min, and 60 min and processed for imaging. Alexa 546-labeled PA was used as an internal reference, and DAPI was used to stain the nuclei of the cells tested. (B) Ratio of NBD fluorescence (green; environment dependent) to fluorescence of Alexa 546 (red; environment independent).