Csu x1 m1 confocal head

Imaging was performed by mounting larvae on a 5% agarose pad in 1 mM levamisole solution in M9 buffer to induce paralysis. Images were taken with a Nikon Ti-U microscope driven by MetaMorph Microscopy Automation and Image Analysis Software (Molecular Devices) and equipped with a Yokogawa CSU-X1-M1 confocal head and an Andor iXon DU-885 camera, using a 60× 1.4 NA objective and with 0.25 μm z-step intervals. Exposure settings were customized for each fluorescently tagged protein, due to wide variations in expression levels and signal intensities. To activate cluster formation with blue light from starting dark conditions, as in Figure 2, z-stacks were taken of the sample with the blue laser with 300ms exposure, 50% laser power, and 50-80 z-stacks (depending on the sample depth). For Figures 3456, animals were mounted with white light therefore clusters were pre-activated before imaging. Animals were then directly imaged, and z-stacks were obtained with both green and blue lasers. Images were analyzed and processed with ImageJ/FIJI (Schindelin et al., 2012) . Photoshop was used to non-destructively prepare images and Adobe Illustrator was used for figure preparation.