Affinity chromatography immobilization

Metal-chelate affinity chromatography (Immobilized-metal (Ion) affinity chromatography)... 

Photoswitchable sub strate-protein / cofactor- enzyme/antigen-antibody interactions Light-stimulated affinity chromatography Immobilization of photoactive separation component on solid matrices... 

Analysts have attempted to use immobilized enzymes for over 30 years. The original motivations were two-fold to conserve enzymes that were expensive and difficult to isolate, and to incorporate enzymes in reusable sensors, such as enzyme electrodes. During such researches, other advantages and applications of immobilized enzymes were found, and immobilization technologies were extended to other areas of biospecific analysis such as immunoassays (see below) and affinity chromatography. Immobilized enzymes have also been u.sed extensively in manufacturing processes and for therapeutic purposes. 

Interaction with ligands Metal binding Pseudo-affinity chromatography Immobilized metal ion affinity chromatography (IMAC)... 

Holzman, T. F., and Baldwin, T. O. (1982). Isolation of bacterial luciferases by affinity chromatography on 2,2-diphenylpropylamine-Sepharose phosphate-mediated binding to immobilized substrate analogue. Biochemistry 21 6194-6201. 

Active derivatives of forskolin include 7-deacetyl-forskolin (EC5o 20(llM), 6-acetyl-7-deacetyl-forskolin (EC5o 40(llIVI), 7-deacetyl-7-0-hemisuccinyl-forskolin (EC50 50(llIVI). The last of these has been used as an immobilized affinity chromatography ligand for the purification of adenylyl cyclases from tissues. 

The fact that adenosine and its derivatives are azo coupling components is used for immobilizing nicotinamide-adenine nucleotide (NAD+) for affinity chromatography purposes. In 12.58 NAD+ is bonded to a matrix through an azo bond. Compound 12.58 is used for the purification of dehydrogenase enzymes (Hocking and Harris, 1973). 

Affinity chromatography A form of chromatography in which separation is achieved by utilizing highly specific biochemical interactions, such as steric-or charge-related conditions, between me analyte and a molecule immobilized on a column. It is different from most forms of chromatography in mat analytes do not continuously elute from me column - only mose mat interact wim me stationary phase are retained and mus separated from omer components of me mixture under investigation. These immobilized materials are eluted from me column after all omer materials have been removed. 

The ELP expression system was compared to the conventional oligohistidme fusion, which is traditionally applied for purification by immobilized metal affinity chromatography (IMAC). Both techniques were shown to have a similar yield of the recombinant protein. The temperature-triggered approach offers a fast and inexpensive nonchromatographic separation with the possibility for larger scale purification. Although the ELP expression system may not be applicable to all types of recombinant proteins, numerous examples have already been shown [40]. 

Crude chloroform-methanol-water (30 60 8, v/v) extracts of immunostainedTLC bands were analyzed without further purification by nanoelectrospray low-energy mass spectrometry. The authors showed that this effective PLC/MS-joined procedure offers a wide range of applications for any carbohydrate-binding agents such as bacterial toxins, plant lectins, and others. Phenyl-boronic acid (PBA) immobilized on stationary support phases can be put to similar applications. This technology, named boronate affinity chromatography (BAC), consists of a chemical reaction of 1,2- and 1,3-diols with the bonded-phase PBA to form a stable... 

Suresh, V., Gallant, S., and Cramer, S., Immobilized metal affinity chromatography displacer characteristics of traditional mobile phase modifiers, Biotechnol. Prog., 12, 84, 1996. 

Li, S. and Dass, C., Iron(III)-Immobilized Metal Ion Affinity Chromatography and Mass Spectrometry for the Purification and Characterization of Synthetic Phosphopeptides, Anal. Biochem., 270, 9, 1999. 

Patwardhan, A.V. and Ataai, M.M., Site accessibility and the pH dependence of the saturation capacity of a highly cross-linked matrix. Immobilized metal affinity chromatography of bovine serum albumin on Chelating Superose, /. Chromatogr. A, 767, 11, 1997. 

Wu, H. and Bruley, D.F, Homologous Human Blood Protein Separation Using Immobilized Metal Affinity Chromatography Protein C Separation from Prothrombin with Application to the Separation of Factor IX and Prothrombin, Biotechnol. Prog., 15, 928, 1999. 

Andersson, L., and Porath, J. (1986). Isolation of phosphoproteins by immobilized metal (Fe3+) affinity chromatography. Anal. Biochem. 154, 250-254. 

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