Quantifying Intestinal Lipid Droplets and Secretion

Black and white images of oil Red O stained animals and fluorescent images were captured using a × 10 objective on a Zeiss Axio Imager microscope. In all cases, lipid droplet staining in the first four pairs of intestinal cells was quantified using NIH Image J software, by measuring pixel intensity over the first four pairs of intestinal cells, which fully encapsulate the variation seen in the tested conditions1 (link). For all atgl-1::GFP images, fluorescence intensity in the second and third pairs of intestinal cells was quantified. Fluorescent images of the reporters for FLP-7 secretion were captured using a × 20 objective on a Zeiss Axio Imager microscope. For all ‘secretion line' animals, the first pair of coelomocytes was imaged. mCherry fluorescence intensity in one of the two imaged coelomocytes was quantified and normalized to the surface area of the coelomocyte. Within each experiment, approximately 15–20 animals from each condition were quantified. All images were quantified using ImageJ (NIH).