Extending the range of rate constants available from BIACORE: interpreting mass transport-influenced binding data.
Surface-based binding assays are often influenced by the transport of analyte to the sensor surface. Using simulated data sets, we test a simple two-compartment model to see if its description of transport and binding is sufficient to accurately analyze BIACORE data. First we present a computer model that can generate realistic BIACORE data. This model calculates the laminar flow of analyte within the flow cell, its diffusion both perpendicular and parallel to the sensor surface, and the reversible chemical reaction between analyte and immobilized reactant. We use this computer model to generate binding data under a variety of conditions. An analysis of these data sets with the two-compartment model demonstrates that good estimates of the intrinsic reaction rate constants are recovered even when mass transport influences the binding reaction. We also discuss the conditions under which the two-compartment model can be used to determine the diffusion coefficient of the analyte. Our results illustrate that this model can significantly extend the range of association rate constants that can be accurately determined from BIACORE.
Myszka
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DG
Huntsman Cancer Institute, University of Utah, Salt Lake City, Utah 84112-5330 USA.
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Dembo
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Morton
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Goldstein
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B
D001665:Binding Sites; D001703:Biophysics / Q000379:methods; D015374:Biosensing Techniques; D003198:Computer Simulation; D004058:Diffusion; D007700:Kinetics; D008962:Models, Theoretical