Protocol

Integrin and Cell Adhesion Molecules

Volume 757 of the series Methods in Molecular Biology pp 191-204

Date:

Live Imaging of LFA-1-Dependent T-Cell Motility and Stop Signals

  • Andrew J. WiemerAffiliated withDepartment of Medical Microbiology and Immunology, University of Wisconsin
  • , Sarah WernimontAffiliated withProgram in Cellular and Molecular Biology, University of Wisconsin
  • , Anna HuttenlocherAffiliated withDepartment of Medical Microbiology and Immunology, University of WisconsinDepartment of Pediatrics, University of Wisconsin Email author 

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Abstract

T-cell motility is critical for leukocyte trafficking both in normal host defense and in pathologic conditions including chronic inflammatory disease. Despite progress in understanding the mechanisms of T-cell polarity and motility, we have limited understanding of the mechanisms that contribute to antigen-induced T cell arrest. Here, we describe methods to analyze leukocyte function antigen-1-mediated T-cell motility and T-cell receptor-induced stop signals using in vitro assays on two-dimensional surfaces. Specifically, methods for live time-lapse imaging of T cell random migration and arrest on ICAM-1-coated surfaces are described. Additionally, we detail methods for live imaging of T-cell motility within 3D substrates to analyze T cell–antigen-presenting cell (APC) interactions and APC-mediated stop signals.

Key words

Integrin Leukocyte function-associated antigen-1 (LFA-1) Leukocyte, lymphocyte T cell Stop signal Immune synapse D10 T cell Conjugation Arrest Microscopy