Abstract
Three different coupling chemistries that have been tried and tested for use in affinity chromatography are described. These methods are particularly recommended for use by workers who do not have access to, or do not wish to use, complex organic chemical synthetic procedures. They have been demonstrated repeatedly to be reliable, efficient, low cost, and easily scaleable up or down in size. The periodate oxidation method works best with Sephacryl type gels and uses only low toxicity reagents and couples well to proteins with both high efficiency and high capacity. The vinyl sulfone method is more reactive and couples both carbohydrates and proteins. The bis-epoxide method, although less reactive, can be used under more extreme conditions of pH to couple otherwise unreactive molecules, such as synthetic polymers, drugs, and so forth.
Similar content being viewed by others
References
Porath, J., Laas, T., and Jansson, J.-C. (1975) Agar derivatives for chromatography, electrophoresis and gel-bound enzymes. III Rigid agarose gels, cross linked with divinyl sulfone (DVS).J. Chromatogr. 103, 49–62.
Sairam, M. R., Clarke, W. C., Chung, D., Porath, J., and Li, C. H. (1974) Purification of antibodies to protein hormones by affinity chromatography on divinyl-sulfonyl Sepharose.Biochem. Biophys. Res. Commun. 61, 355–359.
Fornstedt, N. and Porath, J. (1975) Characterization studies on a new lectin found in seeds ofVicia ervilia.FEBS Letts. 57, 187–191.
Sairam, M. R. and Porath, J. (1976) Isolation of antibodies to protein hormones by bioaffinity chromatography on divinylsulfonyl Sepharose.Biochem. Biophys. Res. Commun. 69, 190–196.
Allen, J. J. and Johnson, E. A. Z. (1977) A simple procedure for the isolation of L-fucose-binding lectins fromUlex europeus andLotus tetragonolobus.Carbohydrate Res. 58, 253–265.
Lihme, A., Schafer-Nielsen, C., Larsen, K. P., Muller, K. G., and Bog-Hansen, T. C. (1986) Divinylsulfone activated agarose-formation of stable and non-leaking affinity matrices by immobilisation of immunoglobulins and other proteins.J. Chromatogr. 376, 299–305.
Jorgensen, T. (1987) High performance liquid chromatography on hydroxyethyl methacrylate.Biochem. H.S. 368, 752.
Wright, J. F. and Hunter, W. M. (1982) A convenient replacement for cyanogen bromide-activated solid phase in immuno-radiometric assays.J. Immunol. Methods 48, 311–325.
Hornsey, V. S., Prowse, C. V., and Pepper, D. S. (1986) Reductive amination for solid-phase coupling of protein. A practical alternative to cyanogen bromide.J. Immunol. Methods 93, 83–88.
Sundberg, L. and Porath, J. (1974) Preparation of adsorbants for biospecific affinity chromatography. 1. Attachment of group-containing ligands to insoluble polymers by means of bifunctional oxiranes.J. Chromatogr. 90, 87–98.
Gelsema, W. J., De Ligny, C. L., Roozen, A. M. P., and Wilms, G. P. (1981) Optimal conditions for the coupling of aromatic amines to epoxy activated Sepharose 6B.J. Chromatogr. 209, 363–368.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Pepper, D.S. Some alternative coupling chemistries for affinity chromatography. Mol Biotechnol 2, 157–178 (1994). https://doi.org/10.1007/BF02824808
Issue Date:
DOI: https://doi.org/10.1007/BF02824808