Interpreting Kinetic Rate Constants from Optical Biosensor Data Recorded on a Decaying Surface

Authors: Joss L.¹; Morton T.A.²; Doyle M.L.³; Myszka D.G.¹

Source: Analytical Biochemistry, Volume 261, Number 2, August 1998 , pp. 203-210(8)

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Abstract:

A capturing assay was used to monitor a Fab-antigen interaction using a BIACORE optical biosensor. The antigen, a truncated single-site mutant (F43V) version of the CD4 receptor, was captured onto the sensor surface using an immobilized nonneutralizing monoclonal antibody. While this assay design created an oriented antigen surface, the antigen slowly dissociated during subsequent binding of the Fab, thus complicating the binding responses. In this paper, we illustrate how binding events occurring on a decaying surface can be accurately described by globally fitting the response data to a model that accounts for the background surface decay. Support for the method was obtained by showing the equilibrium dissociation constant calculated from the kinetic rate constants (K_d = 2.20 ± 0.01 nM) was similar to the value measured in solution using titration calorimetry (K_d = 2.6 ± 0.5 nM). The ability to interpret rate constants from decaying surfaces significantly extends the types of experimental systems that can be quantitatively studied on optical biosensors. Copyright 1998 Academic Press.

Language: English

Document Type: Research article

Affiliations: 1: Huntsman Cancer Institute, University of Utah, 15 N. 2030 E. Rm 2100, Salt Lake City, Utah, 84112-5330 2: John Curtin School of Medical Research, Australian National University, Canberra, ACT 0200, Australia 3: Department of Structural Biology, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania, 19406

Publication date: 1998-08-01