High-resolution imaging of the immunological synapse and T-cell receptor microclustering through microfabricated substrates

T-cell activation via antigen presentation is associated with the formation of a macromolecular membrane assembly termed the immunological synapse (IS). The genesis of the IS and the onset of juxtacrine signalling is characterized by the formation of cell membrane microclusters and the organization...

Full description

Saved in:
Bibliographic Details
Published in:Journal of the Royal Society interface 2011-10, Vol.8 (63), p.1462-1471
Main Authors: Biggs, M. J. P., Milone, M. C., Santos, L. C., Gondarenko, A., Wind, S. J.
Format: Article
Language:English
Subjects:
Antigen presentation
Antigen-presenting cells
Calcium - metabolism
CD3
Cell adhesion
Cell Culture Techniques - instrumentation
Cell Culture Techniques - methods
Cell membranes
Cell migration
Cluster Analysis
Coculture Techniques - instrumentation
Coculture Techniques - methods
Feeder Cells
Genetic Engineering
Histocompatibility
Humans
imaging
Immunological Synapse
Immunological synapses
Lymphocytes T
Macromolecules
Microclusters
Micropits
Muromonab-CD3
T-Cell
T-cell receptor
T-Lymphocytes - physiology
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:T-cell activation via antigen presentation is associated with the formation of a macromolecular membrane assembly termed the immunological synapse (IS). The genesis of the IS and the onset of juxtacrine signalling is characterized by the formation of cell membrane microclusters and the organization of such into segregated microdomains. A central zone rich in T-cell receptor (TCR)–major histocompatibility complex microclusters termed the central supramolecular activation cluster (cSMAC) forms the bullseye of this structure, while the cellular interface surrounding the cSMAC is characterized by regions enriched in adhesion and co-stimulatory molecules. In vitro, the study of dynamic TCR microcluster coalescence and IS genesis in T-cell populations is hampered by cell migration within the culture system and resolution constraints resulting from lateral cell–cell contact. Here, we detail a novel system describing the fabrication of micropit arrays designed to sequester single T-cell–antigen presenting cell (APC) conjugates and promote IS formation in the horizontal imaging plane for high-resolution studies of microcluster dynamics. We subsequently use this system to describe the formation of the cSMAC in T-cell populations and to investigate the morphology of the interfacial APC membrane.
ISSN:1742-5689
1742-5662
DOI:10.1098/rsif.2011.0025