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    Biomaterials Friday Sessions
       Session BI-FrM

Paper BI-FrM8
Study of the Formation and Function of the Cell Membrane Hybrid Layers Containing G-Protein Coupled Receptors at SAM Surfaces

Friday, November 2, 2001, 10:40 am, Room 102

Session: Biosensors
Presenter: V. Silin, National Institute of Standards and Technology
Authors: V. Silin, National Institute of Standards and Technology
R. Madhusudhana, Institute of Biochemical Research, India
J.T. Woodward, National Institute of Standards and Technology
K.D. Ridge, National Institute of Standards and Technology
A. Plant, National Institute of Standards and Technology
Correspondent: Click to Email
Cell membrane vesicles can reorganize at an alkanethiol/lipid mixed monolayers or alkanethiol monolayer-coated surface resulting in the formation of cell membrane hybrid (CMH) layers. The ability to form CMH layers at the surface from various cell types expressing G-protein coupled receptors offers a promising method for the rapid screening of potential membrane receptor ligands. In our work CMH layers were formed using membrane vesicles from monkey kidney COS-1 cells that had been transiently transfected with synthetic human CCR5 chemokine receptors. CMHs were formed on a thiahexa-(ethylene oxide)-octadecane (THEO-C18) thiol surface or on a mixed POPC/THEO-C18 surface. AFM and SPR technique showed that the membrane form continuous layers at the surfaces with a thickness around 4 to 5nm for different samples. CMH surfaces were tested using specific antibodies against CCR5 receptors to check the surface concentration of the receptors at the surfaces and their orientation. The specific binding of the chemokine ligand was detected at this surface over different concentration. The kinetics of the CMH formation, as well as antibody and ligand binding to the surfaces was measured using the SPR technique, which showed extremely good sensitivity for this application. A number of control experiments were carried out with nonspecific antibodies, ligands and membranes to support data for the specificity of the CCR5 receptor response.