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Principle & Advantages
Proteins A, G and L can be good ligands to perform total IgG fraction purification from a sample. Nevertheless it is often required to dispose of the unspecific antibodies population and retain only the antigen-specific antibodies, which then tend to show lower background or non-specific binding levels.
This can be performed by immobilising the antigen of interest (e.g., the peptide used to raise the antibody) on a solid phase so that the antibodies that bind specifically to the antigen are retained upon loading of the serum, while other impurities and unspecific IgG are discarded in the flow-through. However, optimisation of antigen orientation may be a concern especially in the case of peptides, which could be not easily recognised by antibodies if major binding sites are sterically hindered by the chemical bonds with the agarose beads. Thanks to our expertise in purification and peptide chemistry, we are able to determine the best conditions to allow the most effective antigen-antibody interaction.
Immunoaffinity provides many advantages such as very high purity levels (>99% can be achieved in one step), very high selectivity, and therefore very high resolution, including certitude that the antibody is specific for the antigen of interest. Anti-post-translational modification antibodies are purified using a specific 3-step antigen affinity purification procedure, for those antibodies require an extremely specific separation between antibodies which are specific for the control peptide and those which are specific to the modified peptide.
Learn more about our anti-PTM antibody-specific purification procedure.
Purification of antigen-specific antibodies using antigen affinity chromatography
We purify your antibodies by using the following steps:
- immobilisation of 1 mg of antigen on agarose beads
- loading of the antiserum on the affinity column
- elution of bound antigen-specific antibodies
- ELISA on both crude serum and immunopurified antibodies samples
Immunopurified antibodies are supplied in a glycine buffer (glycine 0.1 M, Tris 0.1 M, pH 7.8) containing 0.02% sodium azide. If you do not wish azide to be added, please inform us while placing your order. Average yields obtained range between 50 to 100 µg/ml.