Analysis of NP-40 responses of HEK293 cells by scanning and transmission elecronmicroscopy (SEM and TEM). (A) Demonstration of NP-40–induced MEND in a cell population. Cells were incubated with FM 1-43FX during NP-40 application for 30 s, and then washed and fixed as described in Materials and methods without FM dye. A bright rim of fluorescence was generated in cells treated with detergent, thereby verifying the operation of MEND in this protocol. (B) Scanning electron micrographs of HEK293 cells before (left) and after (right) treatment with NP-40. Control cells show an extensive mesh of membrane ruffles, whereas NP-40 cells appear nearly smooth in comparison. Bar, 10 µm. (C) TEM image of an NP-40–treated HEK293 cell. Detergent-treated cells show large DAB-stained vacuoles (>20 observations). Bar, 5 µm. Stained vesicles are indicated with arrows. (D) DAB-stained vacuoles are significantly increased in HEK293 cells after treatment with NP-40 for 60 s (n ≥ 26; P < 0.001).