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Example sentences for: hitrap

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Recombinant V80R and A61D mutants were expressed in E. Coli BL21 (DE3) pLysS cells (Novagen), and purified by Ni-NTA (Qiagen) and HiTrap SP (Pharmacia) affinity column chromatography followed by ammonium sulfate precipitation as described by Kim et al [ 7 ] .
The pellet was resuspended in 10 ml of buffer B (20 mM HEPES, pH 8.5, 1 mM EDTA, 1 mM DTT), desalted on a Sephadex G-25 column equilibrated with buffer B, then subjected to sequential chromatography on a 5 ml HiTrap heparin column (Pharmacia) and then a 5 ml HiTrap Q column (Pharmacia).
Pooled Macroprep Q fractions containing GFP-BLM were loaded onto a 5 ml HiTrap metal chelate column (Amersham Pharmacia) charged with NiS0 4 equilibrated in Buffer B. The metal chelate resin was washed with successive 10 column volumes of Buffer B + 10mm imidazole, Buffer B + 50 mM imidazole, and Buffer B + l00 mM imidazole.
The pellets from the ammonium sulfate fractionation were resuspended in 60 ml of buffer B (20 mM HEPES pH 8.5 4°C, 1 mM EDTA, and 0.1% 2-mercaptoethanol), desalted on a G-25 sephadex column equilibrated with buffer B, then subjected to sequential chromatography on a 5 ml HiTrap Heparin column (Pharmacia), a 5 ml HiTrap Q column (Pharmacia) and a HiTrap SP column (Pharmacia) respectively.