4.5. Confocal Microscopy
A Zeiss LSM 780 confocal microscope was used to capture high-resolution Z-stacks of images from the fluorescence-stained slides described above. An oil-immersion 60× Plan-Apo objective was used (numerical aperture 1.4). Spatial calibrations were xy = 67 nm/pixel and z = 200 nm/pixel. Fluorescence emission and excitation filters were matched with the green and red fluorochromes. Stacks of images (2048 × 2048 pixels) were captured and stored as uncompressed .tif image files. A fluorescence microscopy co-localization algorithm was used to highlight the areas (pixels) with statistically significant co-localization (ImageJ/FIJI, [