Bio-Rad Media Sampler Pack - Manual (Page 7)

Section 6
Column Packing
Columns can be packed in different ways depending on the type of column and
equipment to be used. Macro-Prep ion exchange media can be packed using a
combination of axial compression with constant pressure, constant flow, or
suction-packing methods. To pack efficient columns, a 50% slurry is
recommended. General guidelines for packing small and large columns are given
below. Follow the instructions for the column hardware to be used. If you are
unsure about a particular column, contact the column manufacturer or your local
Bio-Rad Laboratories representative. Throughout this instruction, flow rate is
expressed in centimeters per hour (cm/hr). The relationship between linear and
volumetric flow rates as delivered by a pump is:
Linear flow rate (cm/hr) x Cross-sectional area of the column (cm 2 ) =
Volumetric flow rate
Example: 300 cm/hr in a 14 cm diameter column = 300 cm/hr x πr 2 or
300 cm/hr x (3.14 x 7 2 ) cm 2 or 46,158 cm 3 /hr or 769.3 ml/min.
Packing Small Columns
The following slurry packing method was designed to pack 25 ml of Macro-Prep
media into 5–15 mm ID, low- to medium-pressure lab columns. Ideally, buffers are
degassed prior to packing. Use a packing reservoir that holds ~100 ml.
1. Mix the prepared medium with an equal volume, 1:1 (v/v), of degassed
packing buffer. For optimal column packing, use packing buffer containing
0.5–1.0 M salt with the same counterion as the start buffer.
Caution: Do not mix with a magnetic stirbar, to avoid formation of small
fines. Larger volumes of slurry may be mixed with an overhead stirrer at low
to moderate speed.
2. Close the outlet of the column. Fill 10% of the column with degassed packing
buffer. Remove any air bubbles that might be trapped in the bed support or
column flow adaptor.
3. Add an appropriate amount of the media in a homogenous slurry to the
column.
4. Fill the remainder of the column with packing buffer.
5. Connect the flow adaptor to the pump, fill it with buffer, and make sure it is
free of air bubbles. Insert the flow adaptor to the column at a slight angle to
avoid trapping air bubbles.
6. Open the column outlet and pump 4–5 column volumes through the column
at a flow rate of about 100 cm/hr. Gradually increase the flow rate from 100
cm/hr to the flow rate at which the maximum allowable pressure for the
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