Computational protocol: Identification of Late Larval Stage Developmental Checkpoints in Caenorhabditis elegans Regulated by Insulin/IGF and Steroid Hormone Signaling Pathways

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Protocol publication

[…] Images were acquired using either a Zeiss AxioImager A1 microscope with a 10×, 20×, or 100× plan-apochromat objective and a Zeiss AxioCam MR charge-coupled device camera, controlled by Zeiss Axiovision software (Zeiss Microimaging, Inc., Thornwood, NJ), or with a Yokogawa spinning disk confocal microscope mounted on a Zeiss AxioImager A1 microscope using iVision software (Biovision Technologies, Exton, PA). Images were processed in ImageJ (NIH Image) and Photoshop CS6 (Adobe Systems Inc., San Jose, CA). Z-stack projections were generated using IMARIS 6.0 (Bitplane, Inc., Saint Paul, MN).Quantification of fluorescence intensity was performed on images acquired at identical exposure settings using ImageJ. For quantifying GFP::DAF-16 in the hypodermis, the fluorescence intensity in four nuclei (excluding nucleoli) were averaged per animal. All measurement of nuclear GFP::DAF-16 were taken within 5 min of removal from food to minimize relocalization of DAF-16 into the nucleus. For quantifying DAF-9::GFP, a contiguous area of the hypodermal syncytium that excluded nuclei was measured in a region below the pharynx. […]

Pipeline specifications

Software tools ImageJ, Imaris
Applications Laser scanning microscopy, Microscopic phenotype analysis
Organisms Caenorhabditis elegans
Diseases Heart Arrest