Bio-Rad Profinity IMAC Resins - Manual (Page 49)

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No protein is eluted
(continued from
previous page)
Elution conditions are too
mild or protein may be in
an aggregated or multimer
form Elute with pH or an
imidazole step-elution
Protein precipitates
during purification
Temperature is too low
Aggregate formation
Perform the purification at
room temperature
Add solubilization agents
to the samples and/or
buffers: 0.1% Triton X-100,
Tween-20, 20 mM
β-mercaptoethanol and
≤20% glycerol to maintain
protein solubility
Poor recovery of target
protein
Protein is found in the
flow-through
Binding capacity of the
column has been
exceeded
Target protein was not
detected in the flow-
through
Strong nonspecific
adsorption of the target
protein to the matrix
See recommendations in
“No protein eluted” section
Increase the column size
or reduce the sample
volume application
Capillary sample loop is
too small
Reduce hydrophobic
adsorption by including
detergents, or organic
solvents, or by increasing
the concentration of NaCl
Problem Possible Cause Solution
10001677B.qxd 1/28/2005 12:42 PM Page 48