Improved method of preparation of supported planar lipid bilayers as artificial membranes for antigen presentation
- 11 May 2011
- journal article
- research article
- Published by Wiley in Microscopy Research and Technique
- Vol. 74 (12), 1174-1185
- https://doi.org/10.1002/jemt.21012
Abstract
T cell activation is the result of direct cell‐cell contact between T cells and antigen presenting cells (APCs), and of interactions between membrane‐bound ligands and receptors at the contact interface, the “immunological synapse.” Model APCs based upon supported fluid lipid bilayers have been used to dissect these complex molecular interactions and to facilitate real‐time microscopic observations. Nearly all studies have used liposome fusion‐based methods to make supported bilayers, and the biophysical properties of these membranes were not characterized in detail. Here, using both Langmuir‐Blodgett and liposome fusion techniques, we explored five different methods of lipid bilayer preparation on glass, mica, or dextran cushion substrates and characterized the stability, homogeneity, and fluidity of the bilayers with fluorescence microscopy and fluorescence recovery after photobleaching (FRAP). Most combinations of techniques and substrates led to unsatisfactory results, notably, a lack of homogeneity for liposome fusion on glass, low stability of bilayers on mica, and loss of fluidity of dextran‐cushioned bilayers in solutions containing protein. To overcome these deficits, we developed a technique that combines liposome fusion on glass and thermally enhanced bilayer expansion. The newly expanded pristine bilayer showed high degrees of stability, homogeneity, and fluidity. MHC and ICAM‐1 molecules anchored on the bilayer diffused freely and stimulated T cell calcium flux and adhesion, respectively. Microsc. Res. Tech., 2011.Keywords
This publication has 36 references indexed in Scilit:
- Pattern Formation and Molecular Transport of Histidine-Tagged GFPs Using Supported Lipid BilayersLangmuir, 2010
- The Glycosylphosphatidylinositol Anchor: A Complex Membrane-Anchoring Structure for ProteinsBiochemistry, 2008
- MHC II molecules and invariant chain reside in membranes distinct from conventional lipid raftsJournal of Leukocyte Biology, 2005
- Agonist/endogenous peptide–MHC heterodimers drive T cell activation and sensitivityNature, 2005
- Identification of Self Through Two-Dimensional Chemistry and SynapsesAnnual Review of Cell and Developmental Biology, 2001
- Wetting of phospholipid membranes on hydrophilic surfaces - Concepts towards self-healing membranesZeitschrift für Physik B Condensed Matter, 1999
- The Immunological Synapse: A Molecular Machine Controlling T Cell ActivationScience, 1999
- Lateral Diffusion of Lipids in Silane-, Dextran-, and S-Layer-Supported Mono- and BilayersLangmuir, 1999
- Lateral Diffusion of Proteins in MembranesAnnual Review of Physiology, 1987
- The diffusion of ions from a phospholipid model membrane systemProtoplasma, 1967