Bioaffinity separation

The described bioaffinity separations demonstrate that polyacrylamide spacers aid the selective binding of highly complex and delicate biomacromolecules and their associates. Moreover, these solutes remain biologically active after desorption probably due to the high inertness and flexibility of the surrounding polymer chains fixed on the solid support. The unbound parts of serum usually show no loss of the activities of their constituents. Thus we evaluate the surface of inorganic supports coated with chemisorbed iV-hydroxyethyl polyacrylamide and its derivatives as being biocompatible. 

Woll, J. M. Hatton, T. A. Yarmush, M. L. "Bioaffinity separations using reversed micellar extraction" Biotechnology Progress. 1989, 5, pp 57-62. 

Figure 8 Examples of reactive supports for the preparation of bioaffinity separation media.

Complementary structures of biological materials, especially those of proteins, often result in specific recognitions and various types of biological affinity. These include many pairs of substances, such as enzyme-inhibitor, enzyme-substrate (analog), enzyme-coenzyme, hormone-receptor, and antigen-antibody, as summarized in Table 11.2. Thus, bioaffinity represents a useful approach to separating specific biological materials. 

Figure 6.10 Procedure for bioaffinity screening of combinatorial drug candidates libraries with two cycles of iterative size exclusion chromatography, using spin columns to separate the receptor and receptor-binder complexes from unbound ligands. (Reprinted with permission from Davis et al., 1999. Copyright 1999 American Chemical Society.)...

Membranes used for separation are thin selective barriers. They may be selective on the basis of size and shape, chemical properties, or electrical charge of the materials to be separated. As discussed in previous sections, membranes that are microporous control separation predominantly by size discrimination, charge interaction, or a combination of both, while nonporous membranes rely on preferential sorption and molecular diffusion of individual species. This permeation selectivity may, in turn, originate from chemical similarity, specific complexation, and/or ionic interaction between the permeants and the membrane material, or specific recognition mechanisms such as bioaffinity. 

Figure 13.11. Bioaffinity electrospray FTICR mass spectrometry. The isolation and mass spectrometricidentificationof receptor-specific ligands are carried out entirely in the mass spectrometer without chromatography or other separation steps.

The kinetics of adsorption-desorption is rarely slow in preparative chromatography, and most examples of a slow kinetics are foimd in bioaffinity chromatography, or in the separation of proteins. Thus, there are few cases in which a reaction-kinetic model is appropriate. This model is important, however, because there are many cases where it is convenient to model the finite rate of the mass transfer... 

Ohlson et al. (05) were the first to describe the combination of a bioaffinity support and HPLC. They used a silica support coated with amino-hexyl AMP groups to separate horse liver alcohol dehydrogenase and porcine lactate dehydrogenase. Antibodies to human serum albumin bonded to silica separated bovine from human serum albumin within 5 minutes. The recovery of human serum albumin added to the column was quantitative (>95%). 

In this section we discuss the choice of a chromatographic system on the basis of, firstly, the type of stationary phase and, secondly, the composition of the eluent. A division in subsections is made from the kinds of stationary phases available, bearing in mind that the more specialized phases for ion-exchange, gel permeation (size exclusion), and bioaffinity chromatography are treated in separate chapters (4.5 to... 

A widely applicable, non-chaotropic elution technique in bioaffinity chromatography is electrophoretic desorption [60,74,75]. If the affinant is coupled to the solid support by an azo bond or by thiol- or alcohol-ester bonds, the complex of the affinant with the isolated substance can be detached from the solid matrix and then the affinity ligand separated by dialysis or gel filtration. 

Turkova, J. (1984). Bioaffinity Chromatography. In Deyl, Z. (ed). Separation Methods. Elsevier, Amsterdam. 

Bioaffinity chromatography means that solute components which have a very specific and selective interaction with the adsorbent are separated into fractions with a high purity. Sometimes this selective bonding is based on a steric effect (key-lock-interaction) but also an equilibrium or kinetic effect can be applied for separation. As a mle also a specific eluent is necessary for regeneration. 

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