Bio-Rad UNO® Monolith Cation Exchange Columns - Manual (Page 5)

Sample Preparation
Proper adjustment of the sample pH and ionic strength is critical for consistent
and reproducible chromatography. For best results, the sample should be exchanged
into the start buffer or diluted to the start buffer’s concentration. Buffer exchange can
be accomplished using Bio-Spin
® 6 or Bio-Spin 30 columns, Econo-Pac ® 10DG
desalting columns, Bio-Gel ® P-6DG size exclusion gel or the Econo-Pac P6 car-
tridge. The choice of product depends on sample volume. Always centrifuge or fil-
ter the sample (0.2–0.45 mm filter) to remove particulates. Application of turbid or
lipid-containing samples may reduce the column lifetime.
Sample Load
The recommended sample load for each column is shown in Table 1. This
amount may vary somewhat depending on the actual sample composition. We do not
recommend overloading the column as both resolution and column lifetime will
decrease. For larger loads, either change to the UNO 1, 6, or 12 ml columns or per-
form several chromatographic runs with a reduced loading. Ideally, samples should
be bound in a concentrated zone at the top of the column. Higher sample loads pro-
duce a broad application zone in which components with less charge are displaced
by more highly charged components. This may result in a shift of certain peaks to an
earlier elution position in the gradient.
Choice of Elution Salt
Sodium or potassium chloride are the most common elution salts and are rec-
ommended for use with UNO columns. Other ions may be used and may show dif-
ferent selectivities based on their relative elution strengths and chaotropic nature.
The following ions are shown below in order of elution strengths:
Cations for UNO S Columns
Barium > Calcium > Magnesium > Potassium > Sodium > Lithium
Anions for UNO Q Columns
Citrate > Sulfate > Iodide > Chloride > Formate > Acetate
See Reference 1 for a more detailed explanation of ion selectivity in chromato-
graphic separations.
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