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Description:
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Investigations of specific and nonspecific interactions of biomolecules at liquid /solid interfaces are presented . To investigate specific multivalent ligand -receptor interactions , bivalent antibodies and haptens bound to solid supported membrane were used as models for ligand -receptor coupling . Novel microfabrication strategies , which included spatially addressed bilayer arrays and heterogeneous microfluidic assays , in conjunction with total internal reflection microscopy , was employed to achieve this goal . These high throughput techniques allow thermodynamic data of binding interactions to be acquired with only a few microliters of analyte and superior signal to noise . The results yield both the first and second dissociation constant for bivalent IgG antibodies with membrane bound hapten molecules . Studies were conducted both as a function of hapten density and cholesterol content in the membrane .
Another research area of this dissertation is the molecular level description of nonspecific adsorption and displacement of the model protein , fibrinogen , onto hydrophilic surfaces . Techniques such as atomic force microscopy , immunochemical assays , fluorescence microscopy , and vibrational sum frequency spectroscopy were employed to probe this system . The results demonstrate that the protein's & #945 ;C domains play the critical role . When fibrinogen is adsorbed to a hydrophilic surface via these moieties , its displacement rate in the presence of human plasma is approximately 170 times faster than when these domains are not in direct surface contact . Even more significantly , spectroscopic studies show evidence for highly aligned Arg and Lys residues interacting with the negatively charged substrate only when the & #945 ;C domains make direct surface contact . The interfacial ordering of these residues appears to be the hallmark of a weak and labile electrostatic attraction between the substrate and the adsorbed macromolecule . |