Physico-chemical boundaries in the continuous and one-step discontinuous affinity-chromatographic isolation of proteins.
From a physico-chemical point of view, affinity chromatography has no unambiguous definition. It is generally understood as the one-step chromatographic isolation of a protein from a biological sample. For such processes the protein recovery and the adsorption capacity for a given adsorption time is limited by static and dynamic physico-chemical properties of the system. The protein recovery is limited by the ratio of the static capacity, n(s), and the dissociation constant, K, for the interaction with the immobilized binding site. The limits of these quantities for 90% and 99% protein recovery were estimated. The residence time required to reach 90% of the adsorptive capacity of an adsorbent is a function of the above static properties, the pore-diffusion coefficient, D(p), and the diffusion distance in the adsorbent. It was estimated and was found to correlate well with experimental data. The one-step discontinuous or continuous chromatographic isolation of one protein from a biological sample by means of adsorbents that separate with respect to different properties is reviewed. This is only possible with selective specific adsorbents and, in special cases, also with bifunctional adsorbents that use hydrophobic interactions for the adsorption, and electrostatic repulsion for the desorption.
Kasche
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AB Biotechnologie II, Technical University Hamburg-Harburg, Denickestrasse 15, D-21071 Hamburg, Germany. kasche@tu-harburg.de
D000042:Absorption; D001665:Binding Sites; D002846:Chromatography, Affinity / Q000379:methods*; D004058:Diffusion; D007700:Kinetics; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*
D011506:Proteins