RESUMO
Fat accumulation is a complex phenotype affected by factors such as neuroendocrine signaling, feeding, activity, and reproductive output. Accordingly, the most informative screens for genes and compounds affecting fat accumulation would be those carried out in whole living animals. Caenorhabditis elegans is a well-established and effective model organism, especially for biological processes that involve organ systems and multicellular interactions, such as metabolism. Every cell in the transparent body of C. elegans is visible under a light microscope. Consequently, an accessible and reliable method to visualize worm lipid-droplet fat depots would make C. elegans the only metazoan in which genes affecting not only fat mass but also body fat distribution could be assessed at a genome-wide scale. Here we present a radical improvement in oil red O worm staining together with high-throughput image-based phenotyping. The three-step sample preparation method is robust, formaldehyde-free, and inexpensive, and requires only 15min of hands-on time to process a 96-well plate. Together with our free and user-friendly automated image analysis package, this method enables C. elegans sample preparation and phenotype scoring at a scale that is compatible with genome-wide screens. Thus we present a feasible approach to small-scale phenotyping and large-scale screening for genetic and/or chemical perturbations that lead to alterations in fat quantity and distribution in whole animals.
