Both of these two types of support are of 4% agarose matrix. Agarose 4FF is rigid, highly cross-linked polymer. It exhibits excellent chemical and physical stability and allows applications using medium pressure (up to 0.3 MPa). Therefore, the recommended flow rate is 300 - 500 cm/hour. Glutathione Agarose is non-cross-linked medium. It offers excellent binding capacity for targeted proteins. It is suitable for applications using gravity flow (with flow rate of 100 - 300 cm/hour).
Efficient, single-step chromatography purification of Glutathione S-Transferase (GST) tagged fusion proteins with high yield and purity. Suitable for other glutathione transferases and glutathione binding proteins. ... Read more
PROTEINDEX™ Glutathione Agarose 4FF is an affinity chromatography medium developed for easy, one-step purification of recombinant glutathione S-transferase (GST) fusion proteins and other glutathione binding proteins expressed in E. coli, insect and mammalian cells. The rigid, highly cross-linked support is suitable for purification scale-up applications.
Our proprietary ProteIndex™ coupling technology ensures stable immobilization of active ligand and minimal ligand leakage in a wide range of purification buffer conditions. The resins can be reused multiple times to purify the same target protein.
- High selectivity for high purity
- > 10 mg GST/mL settled resins
- Stable coupling of ligand, low leakage; can be re-used to purify the same protein
- Versatile, ready-to-use with convenient formats
Purification performance of Glutathione 4FF resins.
50 mL of E.coli lysate containing a GST-tagged fusion protein was loaded on 0.5 mL packed Glutathione 4FF resins at 0.5 mL/mL. The column was then washed with 10 mL of PBS. The target GST-tagged protein was eluted with 7.5 mL of elution buffer (50 mM Tris-HCl, 10 mM GSH, pH 8.0). Fractions of lysate, flow-through, and eluate were analysed by SDS-PAGE. Lanes: (1) Raw cell lysate; (2) Flow-through; (3) Eluate.
|Ligand||Glutathione coupled with a 12-atom linker|
||120 - 320 μmol Glutathione/mL medium|
|Binding capacity||>10 mg GST (26 kDa)/mL medium|
|Matrix||4% highly cross-linked agarose supplied as 50% slurry|
|Particle size||45 μm - 165 μm|
||0.3 MPa (3 bar, 43 psi)|
|Recommended flow rate
||300 - 500 cm/hour|
||2 - 8°C|
||20% ethanol as preservative|
Product is shipped at ambient temperature. Upon receipt store at 2 - 8°C. Do not Freeze.
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Q&As and Reviews
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Customers Questions & Answers
Yes. Glutathione Agarose has high binding capacity for GST-tagged fusion proteins or other glutathione binding protein. The medium can be directly added to pre-treated cell lysates to capture target proteins.
There are many factors that can affect the final purification product. First of all, the composition of the buffers being used in binding, washing, and elution may affect greatly the yield and the purity of the final product. These buffers to should be compatible with the medium. Chemicals, such as 1% Triton X-100, 1% Tween-20, 1% CTAB, 0.03% SDS, or 0.1% NP-40, will not significantly affect the binding of GST or GST-fusion protein to Glutathione Beads. Therefore, these chemicals may be used to reduce non-specific binding. Adding DTT to a final concentration of 1 - 10 mM may greatly increase binding of certain GST-tagged proteins to Glutathione Agarose medium. However, while 1 - 10 mM DTT can be included in the binding and elution buffer to increase purity, it may result in lower yield of some target proteins. Binding of GST-tagged proteins to the chromatography medium depends on its size, conformation, as well as its concentration in the sample loaded. Binding of GST to glutathione is also flow dependent. Lower flow rates often increase the binding capacity, whereas higher flow rates decrease the binding efficiency. Therefore, lower flow rates are recommended, in particular, during sample loading. Furthermore, protein characteristics, pH and temperature may also have impact on the binding capacity.
The answer is no if the target protein is GST-tagged fusion protein and it requires functional GST to bind to the glutathione ligand. The reason is that the GST tag will be denatured in 8 M urea (the same case is for guanidine-HCl) and, even the denaturant is removed, will not be refolded into the functional conformation to bind to the glutathione ligand. Therefore, neither 8 M urea nor guanidine-HCl is recommended in any solutions in the purification procedure. If the presence of denaturant is indispensable for certain applications, urea can be added to a final concentration of up to 1 M, however, at a cost of lower yield of the final product.
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