The release of granule contents coincided with an omega shape visualized by IRM. (A) The fusion of a vesicle labeled with acridine orange (yellow circle) was visualized by TIRF (top) and IRM (bottom). Images were obtained sequentially at 3 Hz. Disappearance of the granule in the TIRF channel coincided in space and time with a local loss of destructive interference. Bar, 0.5 μm. (B) Time course of the change in the intensity of reflected light (black) and fluorescence (red) over an ROI (240 × 240 nm) located in the center of the yellow circle shown in A. Dashed lines indicate the baseline levels of fluorescence (red) and reflected light (black). (C) The fusion of a granule labeled with ANP-GFP (yellow circle) was visualized by sequential TIRF (top) and IRM (bottom). Disappearance of the granule in the TIRF channel coincided in space and time with a local loss of destructive interference. Bar, 0.5 μm. (D) Time course of the change in the intensity of reflected light (black) and fluorescence over an ROI located in the center of the yellow circle shown in C. Dashed lines indicate the baseline levels of fluorescence (red) and reflected light (black).