Fig. 3

Generating colour transmitted light images in ImageJ. Balanced colour images can be generated from individual images or image stacks using a simple procedure in ImageJ, even if the images recorded for the 3 different wavelengths were of different intensities. A confocal hyperstack showing onion epidermal cells (3 transmitted light channels imaged over time) was opened in ImageJ. These were separated into image stacks representing the 633 nm red, 561 nm green and 488 nm blue transmitted light series using the command ‘reduce dimensionality’ (in the ‘hyperstacks’ option in the ‘image’ menu). For each of the stacks, the histogram was adjusted using the ‘brightness/contrast’ command (in the ‘adjust’ option in the ‘image’ menu). This involved adjusting the minimum and maximum values on the histogram to match the edges of the histogram, as indicated by the red arrows. A combined colour image was then generated using the ‘merge channels’ command (in the ‘color’ option in the ‘image’ menu). Scale bar 50 µm