Procedure

Preparation of column: The solid support (bead matrix) is a gel loaded into an elution column. Sepharose, agarose and cellulose are the most commonly used solid support, because the hydroxyl groups on the sugar residues can be easily manipulated to accept a ligand. The ligand is then selected according to the desired isolate. In case a researcher planning to isolate antibodies specific for antigen A from an antiserum, antigen A can be used as ligand. There are several methods of immobilizing protein ligand on these solid support and some examples are given below (Fig. 1).

Figure 1: Immobilizing protein ligand on solid support

Sometimes flexible spacer - arm is attached between ligand and solid support to render better flexibility to ligand. For example NHS-activated Sepharose (agarose beads with 10-atom spacer arms (6-aminohexanoic acid) attached by epichlorohydrin and activated by N-hydroxysuccinimide) is used for protein ligand immobilization (Fig. 2)