Spinning disk confocal upright microscope

For imaging, animals were mounted by placing them in a drop of cold, liquid 36% Pluronic F-127 with 1 mM tetramisole solution and pressed between two coverslips. The slides were brought to room temperature, solidifying the Pluronic F-127 gel and immobilizing the animals. Co-localization images were made using iVision software. Images were taken using a Zeiss Imager D1m upright compound microscope with a 40x dry objective. For confocal imaging, animals were immobilized by using 7.5% M9 agarose pads with 2.5 μl PolySciences 0.05 μm polystyrene microspheres. A Zeiss spinning disk confocal upright microscope with 100x oil immersion objective was used for select images to show additional details, including lysosomal imaging and connection imaging.