Purification by affinity chromatography

EC 1.1.1.27]. 40-Fold purification by affinity chromatography using Sepharose 4B coupled to 8-(6-aminohexyl)amino-5 -AMP or -NAD. [Lees et al. Arch Biochem Biophys 163 561 7974 Pesce et al. J Biol Chem 239 1753 7964.]... 

Cuatrecasas, P. (1970). Protein purification by affinity chromatography. Derivatizations of agarose and polyacrylamide beads. J. Biol. Chem. 245, 3059-3065. 

Lefkowitz, R. J., Haber, E., and O Hara, D. (1972). Identification of the cardiac beta-adrenergic receptor protein Solubilization and purification by affinity chromatography. Proc. Natl. Acad. Sci. USA 69, 2828-2832. 

The demonstration that 3-alkoxypyridines are metalated in the 2-position (Scheme 91) (82S235) allowed the preparation of a series of ribo-furanosyl pyridines 565 as potential deazapyrimidine nucleosides for evaluation as thymidylate synthetase inhibitors (Scheme 170) (86MI2). Thus, metalation of the 3-alkoxypyridines 291 followed by condesation at lower temperatures with a protected D-ribose aldehyde afforded diaster-eoisomeric mixtures of compounds 564 which, upon mesylation and acid-catalyzed cyclization, delivered the ribofuranosyl pyridines 565 in high yields. Purification by affinity chromatography afforded the a- and /3-anomers, which showed insignificant antileukemic activity. 

In consequence of the extensive bibliography of purifications by affinity chromatography, the authors found themselves with a problem regarding which reference to cite for a given application and which applications to choose. 

Kabir S. Immunoglobulin purification by affinity chromatography using protein A mimetic Ugands prepared by combinatorial chemical S5mthesis. Immunol. Invest. 2002 31 263-278. 

EC 1.1.1.27]. 40-Fold purification by affinity chromatography using SepI... 

A. D. Gounaris, M. A. Brown, and A. J. Barrett. Human plasma alpha-cysteine proteinase inhibitor. Purification by affinity chromatography, characterization and isolation of an active fragment. Biochem. J. 221 445 (1984). 

Cuatrecasas P 1970 Protein purification by affinity chromatography J. Biol. Chem. 

In Figure 3 is documented the resolution of a polyclonal antibody sanple exhibiting only a few closely spaced isoelectric bands. The antibodies were raised to the bacterial carbohydrates derived from Micrococcus lysodeiktikus in a rabbit from a colony outbred for simplicity of their clonotype patterns. After purification by affinity chromatography, the antibodies exhibited three major bands, with very close isoelectric points. To effectuate their separation, commercial Airpholine was subfractionated in the RIEF and a narrow cut, pH range 7.5 to 8.5, was used for the fractionation (8). This illustrates the resolution achievable in critical separations. 

In contrast to enzymatic catalyst optimization, artificial metalloenzymes require purified protein samples in milligram quantities. These requirements severely slow down the screening process, the bottleneck being the protein purification by affinity chromatography, followed by dialysis and lyophilization. To accelerate this process, a straightforward extraction-immobilization protocol with biotin-sepharose was implemented to capture functional Sav from crude cellular extracts (Fig. 8) [39, 57], This procedure significantly hastened the optimization process, at the cost of a slight erosion in selectivity. 

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