This is a proposal to purchase the BIAcore biosensor (Pharmacia Biosensor AB, Uppsala, Sweden), and instrument that allows real time biospecific interactions to be analyzed by the detection of the optical phenomenon of surface plasmon resonance (SPR). This system presents several advantages to the study of macromolecular interactions: The interaction of molecules can be followed in real time, which allows the measurement of not only the equilibrium binding constant, but also measurement of the association and dissociation constants. SPR detection can be used more generally than fluorescence and absorption methods since it is sensitive to chances in mass and since no labelling of reagents is required. In addition, only one of the molecules involved in the interaction is required to be in a purified state. Projects initiated by 11 investigators at the Blood Research Institute propose to make use of the requested instrumentation. Several of these projects entail use of this instrument for epitope mapping or for defining the structural determinants that underlie receptor-ligand interactions. Other projects take advantage of the capacity of this system to detect specific binding interactions when one reactive component is contained in plasma or other multicomponent system. There are experiments that require the level of sensitivity for detection provided by the BIAcore or take advantage of this instrument's capacity to kinetically monitor on and off rates and to derive equilibrium affinity constants. Project Tilts include: Function of a C5b-9 Inhibitor in Blood and Vascular Cells; Molecular Variants of von Willebrand's Disease; Monoclonal Antibodies; von Willebrand Proteins and their Cellular relationships; Molecular and Structural Determinants of Wf Interactions; Detection of Drug-dependent, Platelet-reactive Antibodies; Molecular Basis of Heparin-induced Thrombocytopenic Purpura Caused by Quinidine, Quinine, Sulfonamide Antibiotics and Other Drugs; Assessment of Levels of Binding of Antigenic Peptides to Allelic Variants of HLA-DR; Analysis of HLA-DR Structure and Function by Mutagenesis; Immunogenetics of B cell Responses to Platelet Alloantigens; Polymorphism of HLA-DR Expression; Molecular Biology and Function of Human Platelet Integrins; Molecular Biology and Function of PECAM-1; Mechanisms of Platelet Activation by Complement C5b-9; Structural Basis of the Interaction of Wf and FVIII.
