Isolation of Cholera Toxin by Affinity Chromatography on Porous Silica Beads with Covalently Coupled Ganglioside GM1
The great interest of ion exchange and affinity chromatography for protein purification has prompted us to design new particles able to meet the technical requirements for industrial scale fractionation. Porous silica beads “Spherosil ® ” were impregnated with DEAE Dextran. Like other aminated macromolecules with a positive electric charge, it has a very strong affinity for the silica surface. In its presence, the strong and irreversible adsorption properties of silica completely disappear and DEAE Dextran gives the support the physicochemical properties of anion exchangers. After cross-linking with bisepoxy reagents, the DEAE Dextran coating is completely stable even in acid or alkaline solutions. Furthermore this new chromatographic support has outstanding mechanical properties which makes large columns very easy to prepare. The bed height is completely incompressible and allows very high flow-rates from 100 to 400 ml/cm2/h under reasonable pressures. For all these reasons, Spherosil-DEAE Dextran is well adapted to the large scale fractionation of proteins by ĭon-exchange (Tayot et al., 1978-a) and immunoaffinity chromatography (Tardy et al., 1978). We now describe the technique used for the preparation and attachment of the ganglioside GP41 to this spherosil derivative and its application to cholera toxin purification by affinity chromatography. A part of this work is described in a preliminary report (Tayot et al., 1978-b).
KeywordsSialic Acid Affinity Chromatography Cholera Toxin Radial Immunodiffusion Scale Fractionation
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