Affinity chromatography lectin purification

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... 

Lectin affinity chromatography may be used to purify glycoproteins Immobilized antibodies may be used as affinity absorbants for the antigen that stimulated their production (e.g. purification of factor VIII using immobilized anti-factor VIII antibodies)... 

Partial purification of membrane-bound GH receptors from rabbit liver has been achieved after solubilization with detergents [36,38]. Affinity chromatography on immobilized GH or lectins proved a particularly powerful way of isolating the receptors. However, antibodies raised against the partially-purified receptors failed to block the growth-promoting actions of the hormone. 

Shibuya, N., Berry, J. E., and Glodstein, I. J., (1988). One-step purification of murine IgM and human a2-macroglobulin by affinity chromatography on immobilized snowdrop bulb lectin. Arch. Biochem. Biophys. 267, 676-680. 

Affinity chromatography Has a wide number of uses and can be applied to the isolation and purification of virtually all biomolecules. Specific applications include nucleic acid purification, protein purification from cell and tissues extracts, and antibody purification from blood serum. There are a number of matrices used for the construct, and some examples of these and their uses are as follows heparin columns to separate cholesterol lipoproteins, lectin columns to separate carbohydrate groups, and phenyl boronate columns to separate glycated haemoglobins. 

Nearly all current methods of isolation and purification of lectins rely on affinity chromatography. Naturally, the characteristic ligand must be determined in advance. The properties of lectins can be used to precipitate macromolecules and to agglutinate some types of cells, be they plant or animal. The driving force of this reaction is the association with certain bound residues, generally monosaccharides, from the macromolecule or the cellular periphery. When this type of reaction is observed, the problem is to find the sugar that can inhibit activity at the lowest possible molar concentration. As in the case of immunochemical precipitations, this inhibition is due to the occupation of the recognition site by the small soluble molecule. 

Suitable pairs of protein and ligand combination for affinity chromatography are antigen-antibody, hormone-receptor, glycoprotein-lectin or enzyme-substrate/cofactor/ effector (Table 3.6). The affinity of lectins for specific carbohydrate moieties (Lis and Sharon, 1973 Barondes, 1981) has made them particularly useful for the purification of distinct groups of glycans and glycoproteins (Table 3.7). 

Purification of lectin II from Ulex europaeus 196 Affinity chromatography of L-fucosidase 197... 

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