HHMI Janelia Research Campus 3 articles published in JoVE Biochemistry Expression and Purification of the Human Lipid-sensitive Cation Channel TRPC3 for Structural Determination by Single-particle Cryo-electron Microscopy Emery Haley*1, Wooyoung Choi*1, Chen Fan1, Weinan Sun2,3, Juan Du1, Wei Lü1 1Van Andel Institute, 2Vollum Institute, 3Janelia Research Campus This protocol describes techniques used to determine ion channel structures by cryo-electron microscopy, including a baculovirus system used to efficiently express genes in mammalian cells with minimum effort and toxicity, protein extraction, purification, and quality checking, sample grid preparation and screening, as well as data collection and processing. Immunology and Infection Fabricating Optical-quality Glass Surfaces to Study Macrophage Fusion James J. Faust1,2, Wayne Christenson3,4,5, Kyle Doudrick6, John Heddleston7, Teng-Leong Chew7, Marko Lampe8, Arnat Balabiyev1,2, Robert Ros3,4,5, Tatiana P. Ugarova1,2 1Center for Metabolic and Vascular Biology, Mayo Clinic, 2Molecular and Cellular Biosciences, School of Life Sciences, Arizona State University, 3Department of Physics, Arizona State University, 4Center for Biological Physics, Arizona State University, 5Biodesign Institute, Arizona State University, 6Department of Civil and Environmental Engineering and Earth Sciences, University of Notre Dame, 7Advanced Imaging Center, HHMI Janelia Research Campus, 8Advanced Light Microscopy Facility, European Molecular Biology Laboratory This protocol describes the fabrication of optical-quality glass surfaces adsorbed with compounds containing long-chain hydrocarbons that can be used to monitor macrophage fusion of living specimens and enables super-resolution microscopy of fixed specimens. Developmental Biology Using Light Sheet Fluorescence Microscopy to Image Zebrafish Eye Development Jaroslav Icha*1, Christopher Schmied*1, Jaydeep Sidhaye1, Pavel Tomancak1, Stephan Preibisch1,2,3, Caren Norden1 1Max Planck Institute of Molecular Cell Biology and Genetics, 2HHMI Janelia Research Campus, 3Berlin Institute of Medical Systems Biology of the Max Delbrück Center Light sheet fluorescence microscopy is an excellent tool for imaging embryonic development. It allows recording of long time-lapse movies of live embryos in near physiological conditions. We demonstrate its application for imaging zebrafish eye development across wide spatio-temporal scales and present a pipeline for fusion and deconvolution of multiview datasets.
