Thermo Scientific Pierce Glutathione Superflow Agarose consists of glutathione that has been immobilized to a crosslinked 6% beaded agarose that is designed for high-purity, large-scale purification of GST-tagged proteins.
Pierce Glutathione Superflow Agarose is a highly crosslinked, durable resin that does not compress at flow rates required for medium- to large-scale FPLC purifications. The resin holds up well to a variety of chemical and pH values, and it is compatible with common clean-in-place procedures. The specific binding of glutathione to the GST-tag results in high purity and high yield in a single step purification process.
- High dynamic binding capacity – binds 10mg of pure GST per mL of resin at a linear flow rate of 150cm/hour; can bind greater than 20mg/mL at 30cm/hour flow rate
- High purity – provides greater than 90% purity when used to purify overexpressed GST or GST-fusion proteins from lysates
- Robust – highly crosslinked 6% agarose Superflow beaded support tolerates linear flow rates up to 1200cm/hour
- Compatible – maintains function after exposure to a wide variety of chemicals and pH values
- Cost effective – resin is stable through multiple cycles of cleaning and reuse
- Medium- to large-scale FPLC purification of GST-tagged proteins
Properties of Thermo Scientific Pierce Glutathione Superflow Agarose.
||Superflow 6 Resin, highly crosslinked 6% agarose
||60 to 160μm
||Recommended: 150cm/hour (binding, wash, elution)
||Static: approx. 30mg GST/mL resin
Dynamic‡: approx. 10mg GST/mL resin
||1M acetic acid pH 2, 1% SDS pH 7, 6M guanidine-HCl, 70% ethanol for ≥1 week at 37°C; 8M urea, 10mM DTT, 5mM TCEP for ≥2 hours at 22°C
||pH 3 to 9 for ≥1 week at 4°C
pH 2 to 3 or 9 to 12 for ≥ 2 hours at 22°C
||0.05% sodium azide in water
||Up to 25 times
|† Maximum Linear Flow Rate Conditions: Column Dimensions (w x h): 13mm x 38mm (5mL resin); Sample: deionized water at room temperature; Linear Flow Rate: volumetric flow rate (mL/min) x 60 (min/hr)/cross-sectional area (cm2).
‡ Dynamic Binding Conditions (10% breakthrough): Column Dimensions (w x h): 5mm x 50mm (1mL resin); Sample: 1mg/mL GST (26kDa) pure protein in 50mM Tris pH 8.0, 150mM NaCl; Flow Rate: 0.5mL/min.
Affinity chromatography is often used as a quick and easy approach for the purification of recombinant proteins. One of the more common recombinant protein purification methods involves expressing a target protein as a fusion with glutathione S-transferase (GST) and then purifying it via the strong and highly specific interaction of GST to glutathione which has been immobilized on a beaded support.
Pierce Glutathione Superflow Agarose enables high-purity purification of GST fusion proteins using medium to high flow rates. Glutathione has been immobilized by its central sulfhydryl group via a 12-atom spacer arm to a highly crosslinked 6% beaded Superflow agarose. To demonstrate performance, GST was overexpressed and purified on a 48L scale. Biomass (170g) containing overexpressed GST was lysed with 1.7L of lysis buffer. The lysate was clarified using centrifugation and split into two fractions. The target protein was then purified using Pierce Glutathione Superflow Agarose and another leading supplier’s resin in equivalent 200mL packed bead columns. The total yield, recovery, and purity (>95%) were nearly identical for both resins.
|Large scale FPLC purification of GST produces >95% purity of target protein. Biomass (170g) containing overexpressed GST was lysed with 1.7L of lysis buffer and then 0.75L of lysate was loaded onto equilibrated 200mL columns (50mm x 100mm) of Thermo Scientific Pierce Glutathione Superflow Agarose (left) or Glutathione Sepharose™ 4 Fast Flow (GE Healthcare, right) at a linear flow rate of 30cm/hour. Columns were washed with binding buffer until the UV280 reached baseline; then bound protein was eluted with elution buffer, and fractions containing purified GST were pooled. Load, flow-through, wash, and eluate fractions were separated by SDS-PAGE, stained with Imperial Protein Stain (Part No. 24615) and evaluated using myImageAnalysis Software (Part No. 62237) to determine purity. Total yield, recovery, and purity were nearly identical for both resins.
Review of GST-tagged fusion protein production and purification
Other Glutathione Resins for GST-tagged Protein Purification
Affinity Resins for His-tagged Protein Purification
B-PER Bacterial Protein Extraction Reagent
Protease Inhibitor Solutions and Tablets
Slide-A-Lyzer Dialysis Flasks (250mL capacity)