TALON Magnetic Beads

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.