Biomolecular Patterning on Micro- and Nanofabricated Surfaces for Biosensor Applications and Cell-Based Research

摘要

In this study, biomaterials are patterned at the micro- and nanometer scale for biosensor applications and cell-based research. Two enabling technologies were photoactivatable biotin and the polymer lift-off technique. Avidin-biotin complexes have been immobilized on planar silicon substrates and to the inner surfaces of capillary tubes. The latter serves as a model for patterning inside microfluidic systems. Several biomaterials have been patterned using the polymer lift-off method, including lipids, metal, microspheres, biotin, and proteins (e.g., antibodies, protein A, and NeutrAvidin). The patterned lipids formed supported lipid bilayers with diffusion coefficients comparable to cell membranes as confirmed by fluorescence recovery after photobleaching (FRP). The author's tailored patterns comprised 2, 4- dinitrophenyl (DNP) or biotin functionalized lipids with features ranging from 1.3 micrometers to 76 micrometers. Non-lipid molecules were patterned in features as small as 700 nanometers. A characterization of the surfaces has shown a highly uniform patterning. Several of these materials also were applied in patterned regions inside microtrenches. Micro- and nanometer-scale patterns of functionalized biomaterials served as spatially controlled stimuli for cell surface ligands in this research. This technology has been applied to rat basophilic leukocyte (RBL) cells, eosinophils, lymphocytes, macrophages, and fibroblasts. For RBL experiments, DNP-conjugated lipids have been incorporated in the lipid bilayers with fluorescent lipid DiIC16 and used to stimulate RBL cells sensitized with anti-DNP IgE on the Fc receptors. Receptor aggregation was observed with confocal microscopy. This technique allows researchers to study the effects of antigen density on cellular response, binding kinetics, and the redistribution after individual binding events. (1 table, 71 figures).