Computational protocol: RAB-5- and DYNAMIN-1-Mediated Endocytosis of EFF-1 Fusogen Controls Cell-Cell Fusion

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Protocol publication

[…] Images of fixed samples were taken on Zeiss LSM 700 confocal microscope with X63/1.4 PlanApo objective. Superresolution images were obtained using Elyra S.1 structured illumination system (SIM) with X63/1.4 PlanApo objective and EMCCD iXon camera (Andor). Confocal or SIM z stacks were analyzed and processed by Zen software (Zeiss).For the imaging of live embryos, gravid adult hermaphrodites were dissected in 50 μl of 0.7× egg salts and embryos were moved to 2% agar pad with a mouth pipette. The specimen was covered with a coverslip (1.5H) and sealed with scotch tape to prevent drying. Images were acquired using spinning disc Revolution XD confocal system (Andor) based on Nikon Eclipse Ti microscope equipped with 100×/1.3 PlanFluor objective, perfect focus system; CSU-X1 spinning disc (Yokogawa) and iXon3 EMCCD camera (Andor) operated by IQ2 software (Andor). Image sequences were further processed and analyzed with ImageJ (NIH). Figures were prepared using Adobe Photoshop CS5 and Adobe Illustrator CS11. […]

Pipeline specifications

Software tools ImageJ, Adobe Illustrator
Applications Miscellaneous, Laser scanning microscopy, Microscopic phenotype analysis
Organisms Caenorhabditis elegans