title: High-resolution fluorescence microscopy to study transendothelial migration.
abstract: Immune system functions rely heavily on the ability of immune cells (i.e., blood leukocyte) to traffic throughout the body as they conduct immune surveillance and respond to pathogens. A monolayer of vascular endothelial cells (i.e., the "endothelium") provides a critical, selectively permeable barrier between two principal compartments of the body: the blood circulation and the tissue. Thus, knowledge of the basic mechanisms by which leukocytes migrate across the endothelium (i.e., undergo "transendothelial migration"; TEM) is critical for understanding immune system function. Cultured endothelial cell monolayers, used in combination with isolated blood leukocytes, provide a basis for highly useful in vitro models for study of TEM. When used in conjunction with high spatial and temporal resolution imaging approaches, such models have begun to reveal complex and dynamic cell behaviors in leukocytes and endothelial cells that ultimately determine TEM efficiency. In this chapter, we provide protocols for setting up a basic in vitro TEM system and for conducting high-resolution dynamic live-cell and three-dimensional fixed-cell imaging of TEM.
authors: Center for Vascular Biology Research, Division of Molecular and Vascular Medicine, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA. ccarman@bidmc.harvard.edu
authors: Christopher V
authors: CV
authors: Carman
mesh_terms: D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism; D006801:Humans; D007962:Leukocytes / Q000378:metabolism; D008856:Microscopy, Fluorescence; D013194:Staining and Labeling; D059008:Time-Lapse Imaging / Q000379:methods; D057705:Transendothelial and Transepithelial Migration / Q000502:physiology*