Description
Protein A Agarose Resin is an affinity chromatography resin for antibody purification, isolation, and immunoprecipitation. Recombinant Protein A is covalently immobilized on cross-linked agarose beads for selective capture of compatible immunoglobulins. The resin has a binding capacity of approximately 25 mg human IgG per mL and is supplied as a Protein A 4% agarose suspension in 20% ethanol.
Protein A binds primarily to the Fc region of IgG and has strong affinity for antibodies from many mammalian species. Binding varies by species and IgG subclass, so antibody compatibility should be considered when choosing between Protein A and other affinity ligands such as Protein G.
This resin is useful for isolating antibodies from serum, cell culture supernatants, and other biological samples, as well as for antibody-based immunoprecipitation experiments.
The Protein A ligand is covalently attached to the cross-linked agarose support, providing a stable affinity matrix designed to minimize Protein A ligand leakage.
When properly handled, the resin can be regenerated and reused for multiple purification cycles. Bound antibodies can be washed to remove unbound sample components and then eluted for downstream biochemical, analytical, or immunological research.
Binding Capacity
Approximately 25 mg human IgG per mL
Supplied as a Protein A 4% agarose suspension in 20% ethanol
Mechanism
Protein A Agarose Resin captures antibodies through the natural affinity of Protein A for immunoglobulins. Protein A contains multiple antibody-binding domains that interact primarily with the Fc region of IgG. Immobilizing Protein A on agarose allows compatible antibodies to bind to the resin while other sample components are removed during washing.
After binding and washing, captured antibodies are typically released by lowering the pH of the elution buffer. The purified antibody can then be neutralized and prepared for downstream analysis or experimental use.
Common Applications
(Click each for more information)
Antibody and IgG Purification
-
Purpose: To selectively isolate IgG antibodies from serum, cell culture supernatants, and other antibody-containing samples.
-
How It Works: Immobilized Protein A binds primarily to the Fc region of compatible IgG molecules. Unbound proteins and other sample components are removed by washing, and the captured antibodies are then eluted from the resin.
-
Applications: IgG purification, monoclonal antibody purification, polyclonal antibody purification, antibody production research, and preparation of antibodies for downstream assays.
Hober, S., Nord, K., & Linhult, M. (2007). Protein A chromatography for antibody purification. Journal of Chromatography B, 848(1), 40–47. https://doi.org/10.1016/j.jchromb.2006.09.030
Immunoprecipitation
-
Purpose: To isolate a target protein or protein complex from a biological sample using a specific antibody.
-
How It Works: Protein A Agarose Resin captures the Fc region of a compatible antibody. The immobilized antibody can then capture its target antigen from a cell lysate or other sample, allowing the antibody-antigen complex to be separated from unbound material.
-
Applications: Immunoprecipitation, protein isolation, antigen enrichment, protein interaction studies, and preparation of samples for Western blot analysis.
Phizicky, E. M., & Fields, S. (1995). Protein-protein interactions: Methods for detection and analysis. Microbiological Reviews, 59(1), 94–123. https://doi.org/10.1128/mr.59.1.94-123.1995
Antibody Purification from Cell Culture Supernatants
-
Purpose: To recover secreted antibodies from conditioned media following antibody expression or hybridoma culture.
-
How It Works: Compatible IgG molecules in clarified culture supernatants bind to immobilized Protein A, allowing media proteins and other contaminants to be washed away before antibody elution.
-
Applications: Purification of monoclonal antibodies, hybridoma-derived antibodies, recombinant antibodies, and other compatible IgG from cell culture supernatants.
Fahrner, R. L., Knudsen, H. L., Basey, C. D., Galan, W., Feuerhelm, D., Vanderlaan, M., & Blank, G. S. (2001). Industrial purification of pharmaceutical antibodies: Development, operation, and validation of chromatography processes. Biotechnology and Genetic Engineering Reviews, 18(1), 301–327. https://doi.org/10.1080/02648725.2001.10648017
Antibody-Based Protein Complex Isolation
-
Purpose: To enrich proteins associated with a target protein for analysis of molecular interactions and protein complexes.
-
How It Works: A target-specific antibody binds its antigen and associated proteins within a sample. Protein A Agarose Resin captures the antibody-containing complex, allowing it to be separated, washed, and analyzed for interacting proteins.
-
Applications: Co-immunoprecipitation, protein-protein interaction studies, signaling complex analysis, and downstream proteomic characterization.
Free, R. B., Hazelwood, L. A., & Sibley, D. R. (2009). Identifying novel protein-protein interactions using co-immunoprecipitation and mass spectroscopy. Current Protocols in Neuroscience, 46, 5.28.1–5.28.14. https://doi.org/10.1002/0471142301.ns0528s46
Key Benefits
· Selective IgG Capture: Protein A provides strong affinity for the Fc region of compatible IgG antibodies, enabling efficient separation from other proteins and sample components.
· High Antibody Binding Capacity: Provides approximately 25 mg human IgG binding capacity per mL of resin, supporting antibody purification across a range of research-scale sample volumes.
· Stable Protein A Immobilization: Protein A is covalently attached to the cross-linked agarose matrix, helping minimize ligand leakage during antibody purification and immunoprecipitation.
· Suitable for Purification and Immunoprecipitation: The same affinity chemistry can be used for routine antibody purification as well as antibody-mediated capture of antigens and protein complexes.
· Reusable Resin: The resin can be regenerated and reused when properly handled, making it practical for laboratories performing repeated antibody purification.
· Flexible Sample Compatibility: Can be used to capture compatible antibodies from serum, cell culture supernatants, and other antibody-containing biological samples.
Storage/Handling
Storage/Handling: Store at 4°C. Do not freeze.
Powered by Bioz
