Isolation and Characterization of Dendritic Cells and Macrophages from the Mouse Intestine

• Type of Material: Article
• Authors: Duke Geem, Emory University; Oscar Medina-Contreras, Emory University; Wooki Kim, Emory University; Clifton S. Huang, Emory University; Timothy Denning, Emory University
• Language: English
• Date: 2012-05-01
• Publisher: Journal of Visualized Experiments (JoVE)
• Publication Version: Final Published Version
• Copyright Statement: Copyright © 2012, Journal of Visualized Experiments
• Final Published Version (URL): http://dx.doi.org/10.3791/4040
• Title of Journal or Parent Work: Journal of Visualized Experiments
• ISSN: 1940-087X
• Issue: 63
• Start Page: e4040
• End Page: e4040
• Grant/Funding Information: This work was supported by NIH grant AA01787001; a Career Development Award from the Crohn's and Colitis Foundation of America; and an Emory-Egleston Children's Research Center seed grant to T.L.D.
• Keywords: Multidisciplinary Sciences; APCs; immunology; cell culture; intestine; macrophages; dendritic cells; Issue 63; Immunology; Science & Technology; Science & Technology - Other Topics
• Research Categories: Health Sciences, Pathology; Health Sciences, Medicine and Surgery

Abstract

Within the intestine reside unique populations of innate and adaptive immune cells that are involved in promoting tolerance towards commensal flora and food antigens while concomitantly remaining poised to mount inflammatory responses toward invasive pathogens. Antigen presenting cells, particularly DCs and macrophages, play critical roles in maintaining intestinal immune homeostasis via their ability to sense and appropriately respond to the microbiota. Efficient isolation of intestinal DCs and macrophages is a critical step in characterizing the phenotype and function of these cells. While many effective methods of isolating intestinal immune cells, including DCs and macrophages, have been described, many rely upon long digestions times that may negatively influence cell surface antigen expression, cell viability, and/or cell yield. Here, we detail a methodology for the rapid isolation of large numbers of viable, intestinal DCs and macrophages. Phenotypic characterization of intestinal DCs and macrophages is carried out by directly staining isolated intestinal cells with specific fluorescence-labeled monoclonal antibodies for multi-color flow cytometric analysis. Furthermore, highly pure DC and macrophage populations are isolated for functional studies utilizing CD11c and CD11b magnetic-activated cell sorting beads followed by cell sorting.