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Antibody affinity chromatography

When there’s an immunospecific interaction, affinity chromatography (AC) is usually the first, and sometimes only, step required for research. However, a further, polishing step – often size exclusion chromatography – may be needed to obtain a sample with sufficient homogeneity. Multistep purifications are common for biomanufacturing.

With antibody affinity purification there’s a lot of information available about the properties of the target molecule and the major contaminants. This provides a big advantage when purifying antibodies and antibody fragments. Affinity purification offers high selectivity and often high capacity for the target molecule. The target molecule is concentrated into a smaller volume, which means it’s possible to achieve high purity levels – often over 95% – in a single step.

Protein A is commonly used in antibody chromatography, particularly in bioprocessing. Other affinity resin options are protein G for antibodies and protein L for antibody fragments.

Protein A resins

Used for capturing recombinant monoclonal antibodies (mAb) in biomanufacturing processes, protein A chromatography is a form of affinity chromatography for purification of various types and fragments of immunoglobulins (IgGs) from cell culture and other biological samples. This technique relies on antigen binding, which allows you to isolate and purify proteins of interest based on their binding properties to an immobilized ligand.

To purify antibodies from a sample, pass the complex mixture over a column that contains an immobilized protein A ligand attached to a solid support, typically a chromatography base matrix. Antibodies will bind specifically and reversibly to the protein A ligand, resulting in their separation and purification from the original culture.

Get the most out of protein A chromatography with our protein A resins. We feature prepackaged protein A resins for advanced protein separation during the downstream processing of antibody manufacturing. With our resins, you can quickly purify polyclonal, monoclonal antibodies, and other recombinant antibodies from cell cultures.

Browse our prepacked, single-use resin columns and other options for your research, process development, and biomanufacturing needs.

Protein A resins: FAQs

Here are some answers to common questions regarding protein A resins.

What does protein A bind to?

Protein A is a stable surface receptor that's capable of binding to the Fc portion of immunoglobulins from many human and animal species.

What is a protein A column?

Protein A is a cell wall protein from Staphylococcus aureus that forms a reversible bond with immunoglobulins from different species. Protein A columns support the purification of antibodies from complex biological samples – serum, ascites, and cell culture feed, in the case of recombinant antibodies. Since protein A has a high affinity for human IgGs, these columns are extensively used in biopharmaceutical manufacturing.

Protein G and L resins

Besides protein A resins, you can use protein G resins for antibody purification. Each antibody binding protein differs in its capability to bind antibodies of various species and subtypes. It's therefore important to match the binding capacity of the antibody protein to the species and subtype of the target antibody. Protein A and protein G feature binding sites for the Fc segment of mammalian IgGs. Compared to protein A-based resins, protein G binds a broader range of immunoglobulins from eukaryotic species and more classes of IgG. Protein L binds to kappa light chains from various species and is used to purify antibody fragments that contain kappa light chains.