3 articles published in JoVE
Modified Roller Tube Method for Precisely Localized and Repetitive Intermittent Imaging During Long-term Culture of Brain Slices in an Enclosed System Benjamin B. Fixman*1, Isaac W. Babcock*1, Laurie S. Minamide*1, Alisa E. Shaw1, Marina I. Oliveira da Silva1,2, Avery M. Runyan1, Michael T. Maloney1,3, Jeffrey J. Field1, James R. Bamburg1 1Department of Biochemistry and Molecular Biology and Molecular, Cellular and Integrated Neuroscience Program, Colorado State University, 2IBMC-Instituto de Biologia Molecular e Celular, i3S-Instituto de Investigaçãoe Inovação em Saúde, ICBAS, Universidade do Porto, 3Denali Therapeutics Presented here is a modified roller tube method for culturing and intermittent high-resolution imaging of rodent brain slices over many weeks with precise repositioning on photoetched coverslips. Neuronal viability and slice morphology are well maintained. Applications of this fully enclosed system using viruses for cell-type specific expression are provided.
A Standardized Approach for Multispecies Purification of Mammalian Male Germ Cells by Mechanical Tissue Dissociation and Flow Cytometry Ana C. Lima*1,2,3,4, Min Jung*1, Jannette Rusch1, Abul Usmani1, Alexandra M. Lopes3,4, Donald F. Conrad1 1Department of Genetics, Washington University School of Medicine, 2Graduate Program in Areas of Basic and Applied Biology (GABBA), Abel Salazar Institute of Biomedical Sciences, University of Porto, 3Instituto de Investigação e Inovação em Saúde, University of Porto, 4IPATIMUP - Instituto de Patologia e Imunologia Molecular da Universidade do Porto, 5Department of Pathology & Immunology, Washington University School of Medicine This work describes the standardization of a method to obtain purified germ cell populations from testicular tissue of different mammalian species. It is a straightforward protocol that combines mechanical testis dissociation, staining with Hoechst-33342 and propidium iodide, and FACS sorting, with wide applications in comparative studies of male reproductive biology.
Analysis of Cell Suspensions Isolated from Solid Tissues by Spectral Flow Cytometry Sandrine Schmutz1, Mariana Valente2,3,4,5, Ana Cumano2, Sophie Novault1 1Flow Cytometry Core Facility, Center for Translational Research-Technical Core, Institut Pasteur, 2Unit for Lymphopoiesis, Immunology Department, INSERM U1223, University Paris Diderot, Sorbonne Paris Cité, Cellule Pasteur, Institut Pasteur, 3Stem-Cell Microenvironments in Repair/Regeneration Team, Instituto de Investigação e Inovação em Saúde (i3s), INEB - Instituto de Engenharia Biomédica, 4ICBAS - Instituto de Ciências Biomédicas Abel Salazar, Universidade do Porto, 5Stem Cells and Regenerative Medicine Team, UMRS 1166, ICAN - Institute of Cardiometabolism And Nutrition, UPMC - Université Pierre et Marie Curie - Paris 6, INSERM This article describes spectral cytometry, a new approach in flow cytometry that uses the shapes of emission spectra to distinguish fluorochromes. An algorithm replaces compensations and can treat auto-fluorescence as an independent parameter. This new approach allows for the proper analysis of cells isolated from solid organs.