Protein A-Agarose
Protein A-Agarose
I. Introduction
The Imgen BioSceinces’ Protein A-Agarose is 4% agarose beads that have been covalently coupled with recombinant protein A. It is a high quality immunoaffinity matrix for the purification of IgG fractions from biological fluids and from cell culture media. It is also an idea matrix for the isolation of antibody-antigen complexes by immunoprecipitation.Native protein A, a bacterial cell wall protein from Staphylococcus aureus, binds specifically to the Fc region of immunoglobulin molecules of many mammalian species without disturbing their binding of antigen. A native protein A has five IgG binding domains. The recombinant protein A, used to produce Imgen BioSceinces’ Protein A-Agarose, has an exact amino acid copy of the native protein A from S. aureus, but expressed in and purified from E. coli. The purified recombinant protein A functions same as its native form, but with low levels of endotoxin contamination.
II. Features
1. High IgG binding capacity2. Stable affinity medium
3. Milder elution condition than protein G resin
4. Resin is reusable up to 10 - 20 times
III. Characteristics
The matrix is a 4% agarose beads that have been covalently coupled with recombinant protein A by a cyanogen bromide method. The swollen agarose medium has approximately 6 mg of recombinant protein A/ml of drained resin. It’s total binding capacity for human IgG is about 25 -30 mg/ml of drained resin. It has high excellent chromatographic and immunoaffinity isolation qualities in high binding capacity, low leakage, and stable affinity medium. It withstands a wide range of pH, and many common denaturing reagents such as 4 M urea, 4 M thiourea and 6 M guanidine hydrochloride. The product characteristics are summarized in Table 1.Table 1. Technical Specifications
| Ligand | Recombinant protein A (E. coli) |
| Ligand coupling | Cyanogen bromide covalently coupling |
| Ligand density | 6 mg recombinant protein A/ml resin |
| Matrix | 4% agarose beads |
| Average beads size | 90 μm (50 – 150 μm) |
| Binding capacity | ≥25 mg human IgG/ml medium |
| pH stability | 2 - 14 (short term); 3-12 (long term) |
| Flow rate | 1 - 4 ml/min |
Contents
Supplied as 50% aqueous suspension (e.g., 5 ml of settled resin is equivalent to 10 ml of 50% slurry) in buffer containing 20% ethanol, 20 mM sodium phosphate pH 7.4, 0.15 M NaCl, and 0.02% NaN3 (w/v). Product is shipped at ambient temperature.Storage
+4°C to +8°C. Do not freeze.|
Sample Fig. 1. (right) Comparison of SDS-PAGE of purified antibodies from normal human serum by chromatography between with Imgen BioSciences’ Protein A-Agarose and with a leading competitor’s.Lane 1: Molecular weight marker Lane 2: Blank Lane 3: Loaded sample Lane 4: Flow through Lane 5: Elute Lane 6: Blank Lane 7: Loaded sample Lane 8: Flow through Lane 9: Elute |
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