Abstract
Artificial lipid bilayers are powerful tools that can be used to model the interactions between platelets and membrane-bound ligands. To mimic the interaction of platelets with membrane-bound ligands, biotinylated lipids can be used to couple monobiotinylated recombinant ligands to the upper leaflet of an artificial lipid bilayer using streptavidin to bridge the two. Artificial lipid bilayers are generated by preparing liposomes, treating glass coverslips to make them hydrophilic and by assembling the bilayer in a specialized flow chamber. Finally platelets can be added to the flow chamber and the localization of fluorescently labeled molecules followed using microscopy.
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Fooksman DR, Vardhana S, Vasiliver-Shamis G, Liese J, Blair DA, Waite J, Sacristan C, Victora GD, Zanin-Zhorov A, Dustin ML (2010) Functional anatomy of T cell activation and synapse formation. Annu Rev Immunol 28:79–105. https://doi.org/10.1146/annurev-immunol-030409-101308
Groves JT, Dustin ML (2003) Supported planar bilayers in studies on immune cell adhesion and communication. J Immunol Methods 278(1-2):19–32
Fleire SJ, Batista FD (2009) Studying cell-to-cell interactions: an easy method of tethering ligands on artificial membranes. Methods Mol Biol 462:145–154. https://doi.org/10.1007/978-1-60327-115-8_9
Pollitt AY, Poulter NS, Gitz E, Navarro-Nunez L, Wang YJ, Hughes CE, Thomas SG, Nieswandt B, Douglas MR, Owen DM, Jackson DG, Dustin ML, Watson SP (2014) Syk and Src family kinases regulate C-type lectin receptor 2 (CLEC-2)-mediated clustering of podoplanin and platelet adhesion to lymphatic endothelial cells. J Biol Chem 289(52):35695–35710. https://doi.org/10.1074/jbc.M114.584284
Dustin ML, Starr T, Varma R, Thomas VK (2007) Supported planar bilayers for study of the immunological synapse. Curr Protoc Immunol Chapter 18:Unit 18.13. https://doi.org/10.1002/0471142735.im1813s76
Watson SP, Herbert JM, Pollitt AY (2010) GPVI and CLEC-2 in hemostasis and vascular integrity. J Thromb Haemost 8(7):1456–1467. https://doi.org/10.1111/j.1538-7836.2010.03875.x
Nye JA, Groves JT (2008) Kinetic control of histidine-tagged protein surface density on supported lipid bilayers. Langmuir 24(8):4145–4149. https://doi.org/10.1021/la703788h
Valvo S, Mayya V, Seraia E, Afrose J, Novak-Kotzer H, Ebner D, Dustin ML (2017) Comprehensive analysis of immunological synapse phenotypes using supported lipid bilayers. Methods Mol Biol 1584:423–441. https://doi.org/10.1007/978-1-4939-6881-7_26
Schutz GJ, Huppa JB (2017) Forster resonance energy transfer to study TCR-pMHC interactions in the immunological synapse. Methods Mol Biol 1584:207–229. https://doi.org/10.1007/978-1-4939-6881-7_14
Su X, Ditlev JA, Rosen MK, Vale RD (2017) Reconstitution of TCR signaling using supported lipid bilayers. Methods Mol Biol 1584:65–76. https://doi.org/10.1007/978-1-4939-6881-7_5
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Dustin, M.L., Pollitt, A.Y. (2018). The Study of Platelet Receptors Using Artificial Lipid Bilayers. In: Gibbins, J., Mahaut-Smith, M. (eds) Platelets and Megakaryocytes . Methods in Molecular Biology, vol 1812. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8585-2_8
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DOI: https://doi.org/10.1007/978-1-4939-8585-2_8
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