Bio-Rad Nuvia™ IMAC Resin - Manual (Page 31)

Nuvia IMAC Ni-Charged Resin 27
Part 2: Washing the Resin
1. Insert micro spin column into new collection vessel.
2. Wash the resin with at least 5 column volumes of binding
buffer containing imidazole.
Pipet up and down at least 5 times.
Note: If previously determined, you may use an optimized
concentration of imidazole that is slightly less than the
concentration necessary to elute the target protein. See
Section 8.
3. Centrifuge at 1,000 x g for 1 min.
Remove remaining unbound proteins by centrifuging. The
wash step can be repeated if necessary.
Part 3: Eluting the Histidine-Tagged Protein
1. Insert micro spin column into a new, clean collection vessel.
2. Elute bound proteins with 5 column volumes of elution buffer.
Pipet up and down at least 5 times and incubate for up to
5 min.
The choice of elution buffer will vary depending on the
procedure used. For example, a range of imidazole
concentrations (150–500 mM) may be used to elute bound
protein from the Nuvia IMAC resin.
3. Analyze fractions from above steps by A
280
, SDS-PAGE,
ELISA, etc.