Cyanogen bromide activation

March, S.C., Parikh, I., and Cuatrecasas, P. (1974) A simplified method for cyanogen bromide activation of agarose for affinity chromatography. Anal. Biochem. 60, 149-152. 

The immobilization of any other protein to cyanogen bromide-activated or N-hydroxysuccinimide- (NHS) activated gels is done the same way. 

Immunoaffinity chromatography cleanup has also been applied as an ideal and reliable strategy for residue analysis. Immunoaffinity columns prepared by coupling the antibodies to a cyanogen bromide-activated support were used to analyze avermectin BI residues in cattle tissues (359) and ivermectin in sheep serum (376). An immunoaffinity column prepared by an alternative activation/ coupling procedure with carbonyl diimidazole was also employed to analyze ivermectin residues in swine liver (361) since the earlier-reported methods did not work well in the analysis of this matrix. This recent work demonstrated the high specificity of tire antibody-mediated cleanup, but also showed that the immunoaffinity procedures could not always or completely eliminate matrix interference of samples. Therefore, application of additional cleanup steps before or after these procedures is often inevitable. 

The addition of phenylalanine to the previously described starch base materials (Control and Oxidized starches) was done on a lOX (w/w) level for covalent attachment. The Oxidized starch was apparently more conductive to cyanogen bromide activation for amino acid attachment than was the Control starch (92X attachment versus 67X, respectively). The addition of aspartame was also completed for the Oxidized (ii) starch polymer on a level of... 

An immunoadsorbent has been prepared by attaching BSA, containing p-diazophenyl l-thio-/3-D-galactopyranosides, to cyanogen bromide-activated Sepharose.175 This affinity material was used to isolate mouse immunoglobulins (myeloma proteins) that bound carbohydrate.178... 

Antibody affinity chromatography is employed to isolate antigen-specific antibodies. The most common affinity matrix for coupling of molecules is cyanogen bromide-activated... 

Sprinkle 1 g of cyanogen bromide-activated Sepharose 4B (Pharmacia-LKB, Piscataway, NJ) over 20 ml of 10 mM HC1. The gel swells immediately. One gram of dry gel yields 3.5 ml of hydrated matrix. 

Sepharose 4B, cyanogen bromide-activated-Sepharose, or protein A, G, or L Sepharose. 

Coupling of Protein to Cyanogen Bromide-Activated Agarose... 

If purchased, cyanogen bromide-activated agarose will be in a dried form in the presence of stabilizers and will require swelling and washing before use. The activated groups are liable to hydrolysis at alkaline pH and thus the swelling and washing is carried out at acid pH. 

Efficiencies less than 70% indicate that inefficient IgG immobilization has occurred. The cause of inefficient coupling is often the presence of a buffer component (free amines in Tris-HCl buffers, for example) that competes with primary amines of the antibody for binding at the active sites of the cyanogen bromide activated Sepharose beads. 

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