Bio-Rad Profinity IMAC Resins - Manual (Page 34)

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3. Apply a linear gradient from 100% refolding buffer to 100% phosphate
elution buffer, with high imidazole.
Note: Elution is done by a gradient from 20 mM to 500 mM imidazole.
4. Access protein purity and recovery of fractions containing His-tagged
proteins.
Use an activity assay (Bio-Rad’s protein assay kit), SDS-PAGE, or western blot
analysis with anti-histidine antibodies or antibodies specific to the target
protein.
5. If necessary, add an additional chromatography step.
Size exclusion chromatography may be a good choice because aggregates of
un-refolded protein can be removed and the buffer composition of the purified
material can be changed simultaneously.
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