Affinity purification of natural ligands.
Immobilization of proteins, nucleic acids, and other bioligands is not always straightforward since they are often large molecules with numerous chemically reactive groups that can all participate in the immobilization process through physical adsorption, ionic binding, or covalent linkage. Protocols for some of the most frequently used matrix-activation systems are described in this unit. For agarose, protocols are given for cyanogen bromide, p-nitrophenyl chloroformate, tresyl chloride, and cyanuric chloride. Tosyl chloride is used to activate cellulose, and cyanuric chloride is also used to activate aminopropyl silica gel. Activation of magnetic beads with cyanogen bromide is described, and a protocol is provided for reacting the aldehyde groups of glyoxal agarose beads with the primary amine groups of ligands, with subsequent reduction of the formed Schiff base to yield a stable matrix-ligand bond.
Luong
John H T
JH
National Research Council Canada, Biotechnology Research Institute, Montreal, Quebec, Canada.
Scouten
William H
WH
D002846:Chromatography, Affinity / Q000379:methods*; D019995:Laboratory Chemicals; D008024:Ligands; D008722:Methods
D019995:Laboratory Chemicals; D008024:Ligands