Isolation of 3,4-dihydroxyphenylalanine-Containing Proteins by Boronate Affinity Chromatography (CAT#: STEM-ACM-0231-CJ)

When boric acid or borate is used as a ligand in affinity chromatography, the method is known as boronate affinity chromatography.

In general affinity chromatography is composed of a stationary phase (solid phase) and a mobile phase. The mobile phase is your cell lysate or any mixture that contains biomolecules. A ligand that binds the target molecule is attached covalently to the solid phase. The interaction between the solid and the mobile phase are exploited by affinity chromatography to get your desired substance in a pure form. The target molecule binds to the ligand, whereas most of the other molecules flow through. The target biomolecule is eluted by changing conditions (pH or salt concentrations) or by competition with a free ligand.The most important property which the solid phase should have is ligand immobilization. Various materials like acrylates or silica gels are appropriate. To prevent steric interference of the target molecule to the ligand, an inhibitor is attached to the solid phase. This inhibitor is known as the spacer. The classic spacer is an inhibitor containing a hydrocarbon chain (CH2 spacer).

Analytical Biochemistry

1. Preparation of Column: The column is loaded with solid support such as sepharose, agarose, cellulose etc.. Ligand is selected according to the desired isolate. Spacer arm is attached between the ligand and solid support.
2. Loading of Sample: Solution containing a mixture of substances is poured into the elution column and allowed to run at a controlled rate.
3. Elution of Ligand-Molecule Complex: Target substance is recovered by changing conditions to favor elution of the bound molecules.

• Sample: Plants; Natural Food; Protein; Drug; Pollutants; Blood; Saliva; Serum; Plasma; Antibodies; Viruses & More
• Equipment: Agarose; Silica gel; Aluminium oxide; Acrylate; Organic polymers; Wash Buffer
• (Optional): Ligand; Spacer arm; Column

1. Most boronate derivatives are known to covalently bind substances with cis-diol groups at a pH greater than 8.
2. Due to the cis-diol groups of the sugars, separation of glycoproteins from non-glucoprotein structures is achievable by the boronate affinity method.
3. Boronate affinity chromatography (BAC) is commonly used in sample preparation techniques for its specific selectivity for cis-diol-containing compounds.