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Binding isotherms by continuous-flow dynamic dialysis.

Abstract

The classical dynamic dialysis technique for the determination of a protein-ligand binding isotherm has been modified by the introduction of a flow cell in which the dialysate on the sink side of the membrane is continuously eluted with a constant flow of eluting buffer and its ligand concentration measured. This new experimental method is termed continuous-flow dynamic dialysis (CFDD). A transfer function procedure for extracting the binding isotherm from the dialysis data is described. This is a more general technique (requiring only a verifiable assumption of linearity) than that previously used, in which the system was modelled using Fick's first law and which relied on the establishment of quasi-steady state conditions across the membrane. The present analysis uses the Laplace transform to effect deconvolution of the impulse response function of the cell from the dialysis data and, using a Fourier series approach, directly yields numerical data representing the free ligand concentration in equilibrium with the protein-ligand complex. The protein-ligand binding isotherm is obtained in parametric form, with time as the parameter.

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BibTeXRIS

N A Sparrow, L Glasser, D Glasser, A E Russell. 1986. Binding isotherms by continuous-flow dynamic dialysis.. https://doi.org/10.1016/0731-7085(86)80067-x

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