Thio-Link Gel Kit
The Thio-link Gel Kit contains all the buffers, blocking agents, poly-propylene columns and materials required to make 5 x 2ml affinity purification columns.
This is for linking Haptens/Peptides/Immunogens (also called ligands) to produce Affinity Columns for the direct purification of antisera. Thio Link Geltm is an activated support that reacts and specifically binds to sulphydryl (thiol) groups on proteins and peptides.
The -S-S- bonds in immunoglobulin antibodies (e.g IgG) can be reduced with dithiothreitol (DTT), 2-mercaptoethanol or 2-mercaptoethylamine to form -SH groups that will bind directly to the gel within the column. This does not affect the heavy or light chains of the immunoglobulin molecule and binds the antibody irreversibly to the gel column.
It is also ideal for immobilising peptides via a cysteine Sulphydryl group. The gel can then be used to affinity purify the corresponding anti-peptide antibody.
Thio-Link Geltm conains a 12-atom hydrophilic spacer arm to improve the accessibility of the binding sites to large proteins. This increases its efficiency, because the linker molecule is long enough to prevent any problems with steric hindrance, lodo-acetyl chemistry is used which is specific for free thiols and gives an irreversible linkage to the ligand molecule. It binds approximately 6mg cysteine per ml bed volume.
The schematic shows stepwise how to use the gel kit. From pouring the gel into the column through to binding the hapten to blocking unreacted sites on the gel,capturing the antibody of interest and eluting the affinity purified anti-sera molecules from the gel.
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