TALON Magnetic Beads
TALON Magnetic Beads
Brand: Takara Bio.
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TALON Magnetic Beads
High-throughput, micro-scale purification of his-tagged proteins—TALON Magnetic Beads
TALON Magnetic Beads provide the selective chemistry of TALON immobilized metal affinity chromatography (IMAC) resin in a magnetic bead format. The beads are designed for the microscale purification of high and low molecular weight recombinant his-tagged proteins under native or denaturing conditions, when placed on a magnetic separator.
Overview
- Quick and easy separation of his-tagged proteins
- Highly-selective TALON chemistry for increased purity
- Eluted samples ideal for small-volume applications
Applications
TALON Magnetic Beads provide simple, effective separation of recombinant his-tagged proteins for a wide range of applications, including:
- Microscale purification of his-tagged proteins
- Studying protein structure and function
- Preparing proteins for X-ray crystallography
- Performing assays that detect protein-protein and protein-DNA interactions
- Immunization to raise antibodies against a protein of interest
- Screening for protein expression
- Optimizing purification conditions for scale-up with TALON Resin
TALON Magnetic Beads can also be used to immobilize 6xhis-tagged proteins for affinity chromatography, in order to:
- Analyze protein-protein and protein-nucleic acid interactions
- Purify untagged subunits or nucleic acids that interact with the immobilized protein
- Perform antibody purification
- Study interactions between ligands and receptors
Advantages
- Choice of native or denaturing conditions to purify proteins
- Directed presentation of 6xhis-tagged proteins increases reproducibility and signal-to-noise ratios
- Varying the amount of beads per well allows for a wide range of binding capacities
- A powerful tool for analyzing interactions between biomolecules
Purification of his-tagged proteins using TALON Magnetic Beads
Purification of his-tagged proteins using TALON Magnetic Beads. Proteins were expressed in BL21 E. coli cells and extracted in xTractor Buffer (Cat. No. 635623). Then 200 μl of a 5% suspension of TALON Magnetic Beads were washed with water to remove storage buffer and equilibrated with loading buffer (50 mM sodium phosphate, 0.3 M NaCl, pH 7.2). The extract was mixed with TALON Magnetic Beads for 30 min at room temperature (6xHN-AcGFP1) or 60 min at 4°C (6xHN-LacZ). The beads were washed with loading buffer, followed by washing with 10 mM imidazole in loading buffer. The protein was eluted with 250 mM imidazole in the loading buffer. Panel A. SDS PAGE analysis (4–20% gradient gel) of 6xHN-AcGFP1 purification using TALON Magnetic Beads. Lane 1: Starting E. coli extract. Lane 2: Nonadsorbed material. Lane 3: Eluted protein. Panel B. SDS-PAGE analysis (4–20% gradient gel) of 6xHN-LacZ purification using TALON Magnetic Beads. Lane 1: Starting E. coli extract. Lane 2: Nonadsorbed material. Lane 3: Eluted protein. Lane M: Molecular weight markers. The SDS-PAGE results for both constructs indicate a high level of purification.
Native vs. denaturing purification procedures using TALON resin
Native vs. denaturing purification procedures using TALON resin.
Native purification with TALON resin preserves biological activity of proteins
Native purification with TALON resin preserves biological activity of proteins. Fresh cells (0.5 g) expressing 6xHis-GFPuv were extracted in 5 ml of 50 mM sodium phosphate; 0.3 M NaCl, pH 7.0 Panel A. Elution profile of GFP which was loaded, washed with the same buffer, and eluted with a step gradient of imidazole (150 mM). Panel B. Fractions were analyzed by SDS-PAGE. The fluorescent signal of green fluorescent protein (GFPuv) was completely enriched by TALON Superflow Resin.
Purification of 6xHis-GFPuv under denaturing conditions using TALON resin
Purification of 6xHis-GFPuv under denaturing conditions using TALON resin. The fusion protein was purified in 8 M urea using TALON resin. M=molecular weight markers.
Native purification of 6xHis protein using TALON resin in the presence of beta-mercaptoethanol
Native purification of 6xHis protein using TALON resin in the presence of beta-mercaptoethanol. N-terminal 6xHis-tagged mouse DHFR (19.5 kDa) was expressed in E. coli. 2 ml of lysate was purified using gravity flow on TALON resin in increasing concentrations of beta-mercaptoethanol. Even lanes: 20 μl of nonadsorbed material. Odd lanes: 5 μl of eluate
6xHN-tagged AcGFP1 and LacZ, purified using TALON Magnetic Beads
6xHN-tagged AcGFP1 and LacZ, purified using TALON Magnetic Beads. TALON Magnetic Beads bind his-tagged proteins ranging from low to high molecular weight with high specificity.
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Purification of his-tagged proteins using TALON Magnetic Beads
Purification of his-tagged proteins using TALON Magnetic Beads. Proteins were expressed in BL21 E. coli cells and extracted in xTractor Buffer (Cat. No. 635623). Then 200 μl of a 5% suspension of TALON Magnetic Beads were washed with water to remove storage buffer and equilibrated with loading buffer (50 mM sodium phosphate, 0.3 M NaCl, pH 7.2). The extract was mixed with TALON Magnetic Beads for 30 min at room temperature (6xHN-AcGFP1) or 60 min at 4°C (6xHN-LacZ). The beads were washed with loading buffer, followed by washing with 10 mM imidazole in loading buffer. The protein was eluted with 250 mM imidazole in the loading buffer. Panel A. SDS PAGE analysis (4–20% gradient gel) of 6xHN-AcGFP1 purification using TALON Magnetic Beads. Lane 1: Starting E. coli extract. Lane 2: Nonadsorbed material. Lane 3: Eluted protein. Panel B. SDS-PAGE analysis (4–20% gradient gel) of 6xHN-LacZ purification using TALON Magnetic Beads. Lane 1: Starting E. coli extract. Lane 2: Nonadsorbed material. Lane 3: Eluted protein. Lane M: Molecular weight markers. The SDS-PAGE results for both constructs indicate a high level of purification.
Native vs. denaturing purification procedures using TALON resin
Native vs. denaturing purification procedures using TALON resin.
Native purification with TALON resin preserves biological activity of proteins
Native purification with TALON resin preserves biological activity of proteins. Fresh cells (0.5 g) expressing 6xHis-GFPuv were extracted in 5 ml of 50 mM sodium phosphate; 0.3 M NaCl, pH 7.0 Panel A. Elution profile of GFP which was loaded, washed with the same buffer, and eluted with a step gradient of imidazole (150 mM). Panel B. Fractions were analyzed by SDS-PAGE. The fluorescent signal of green fluorescent protein (GFPuv) was completely enriched by TALON Superflow Resin.
Purification of 6xHis-GFPuv under denaturing conditions using TALON resin
Purification of 6xHis-GFPuv under denaturing conditions using TALON resin. The fusion protein was purified in 8 M urea using TALON resin. M=molecular weight markers.
Native purification of 6xHis protein using TALON resin in the presence of beta-mercaptoethanol
Native purification of 6xHis protein using TALON resin in the presence of beta-mercaptoethanol. N-terminal 6xHis-tagged mouse DHFR (19.5 kDa) was expressed in E. coli. 2 ml of lysate was purified using gravity flow on TALON resin in increasing concentrations of beta-mercaptoethanol. Even lanes: 20 μl of nonadsorbed material. Odd lanes: 5 μl of eluate
6xHN-tagged AcGFP1 and LacZ, purified using TALON Magnetic Beads
6xHN-tagged AcGFP1 and LacZ, purified using TALON Magnetic Beads. TALON Magnetic Beads bind his-tagged proteins ranging from low to high molecular weight with high specificity.
