Conjugating Ligands to Agarose Beads for Affinity Purification
Affinity purification is a frequently used technique for purifying proteins. During affinity purific...
Biomolecules can be immobilized by binding them to a support under conditions that will then determine the characteristics of the complex. Immobilization is a technique that binds a ligand (enzyme, antibody, affinity proteins like Protein A or Protein G) to a support, giving high stability and making it easier to reuse the immobilized molecule.
Glyoxal beads allow a covalent binding of agarose to lysine amino groups (similar orientation to CNBr). Glyoxal agarose beads are ready to use and require no activation. The resin has a high amount of aldehydes groups (15-25 µmoles Glyoxal / ml gel) providing a very good option to conjugate affinity ligands improving the stability of the ligand.
The choice of Glyoxal or Aminoethyl will depend on the biomolecule to be immobilized, the accessibility of the reactive groups, and the direction/ orientation required for the binding to the support.
Glyoxal 6% agarose beads are suitable for use with FPLC, but with pressures no more than 20 kPa. Too much pressure or too fast of a flow rate will result in diminished performance.
High Density Resin |
Low Density Resin |
High Crosslinking |
Low Crosslinking |
Best for smaller proteins |
Best for larger proteins (e.g. antibodies) |
Best for repeated use or FPLC systems |
Best for batch incubations or gravity flow |
Small pores |
Large pores |
Stronger covalent binding of ligands improves ligand stability |
Weaker covalent binding improves ligand mobility after immobilization |
Store at 4°C. Do not freeze.
Catalog ID | G-302 |
---|---|
Name(s) | Glyoxal (Low Density) 6% Agarose Beads, Crosslinked |
Bead Size | Spherical, Standard: ~50-150 μm (approx.) |
Cross-linking | Yes |
Binding/Loading Capacity | 5-10 (mg BSA/ml gel) |
Activity | 15-25 (μmol glyoxyl/ml gel) |
Agarose % | 6% agarose |
Chemical Stability | Stable in all commonly used reagents. |
Temperature Stability | Autoclavable at 120°C for 20 min in H2O |
Storage Buffer | 20% Ethanol |
Storage/Handling | store at 4°C. Do NOT freeze. |
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