Purification of Heart Hexokinase by Dye Ligand Chromatography (CAT#: STEM-ACM-0200-CJ)

Dye-ligand affinity chromatography is one of the Affinity chromatography techniques used for protein purification of a complex mixture. Like general chromatography, but using dyes to apply on a support matrix of a column as the stationary phase that will allow a range of proteins with similar active sites to bind to, refers to as pseudo-affinity. Synthetic dyes are used to mimic substrates or cofactors binding to the active sites of proteins which can be further enhanced to target more specific proteins. Follow with washing, the process of removing other non-target molecules, then eluting out target proteins out by changing pH or manipulate the salt concentration. The column can be reused many times due to the stability of immobilized dyes. It can carry out in a conventional way by using as a packed column, or in high-performance liquid chromatography (HPLC) column.

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. This approach is particularly popular for purifying enzymes and proteins. Dye-ligand adsorbents are desirable because they are affordable, readily available, and simple to immobilise. These adsorbents can be utilised for analytical, preparative, and large-scale research.
2. The dye-ligand affinity approach for pharmaceuticals has been discontinued due to leakage and toxicity issues.