Figure 3
Two-photon time-lapse imaging of fusion and fission of vesicles containing BChol. CHO cells were pulse-labeled with BChol followed by a chase to obtain the steady-state distribution. The cells were placed on a temperature-controlled stage of a home-built 2P-microscope maintained at 35 ± 1°C, and images were acquired without pause at a frame rate of 1.05 s. We observed vesicle fusion (A’): one vesicle (white arrow) moved with a directed motion toward another vesicle (black arrow) over a period of 18 s. Further observation of the fused vesicle over 100 s did not show subsequent fission of the vesicle. We also observed vesicle fission (B, B’): over a period of ~1 min the vesicle was heavily stretched and deformed before it finally separated into two vesicles with lower fluorescence intensity.