Bio-Rad Profinity Epoxide Resin - Manual (Page 7)

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After coupling, ligand-coupled resin needs to be washed thoroughly with coupling
buffer to remove excess ligand, but sometimes small amounts of ligand are still
trapped on the resin through ionic interactions. Additional wash cycles alternating
acidic and basic buffers are recommended. Each cycle should consist of a wash
with a pH 4.0 buffer (100 mM acetate, 500 mM NaCl) followed by a wash with a
pH 8.0 buffer (100 mM phosphate, 500 mM NaCl).
Section 3
Column Packing
General handling
Profinity base resin is a rigid support that can operate under high flow rates and
pressures. However, bead damage due to excessive physical force is possible.
Magnetic stirrers or excessive stirring may cause mechanical damage and fracture
of some beads. Fine particles generated in this manner may clog the column and
increase the column backpressure; in addition, they may enter the pores of intact
beads and reduce their binding capacity.
Small column packing — slurry packing
Slurry packing is the preferred packing method for small columns. Use this
method for packing ligand-coupled Profinity epoxide resin into 5–15 mm ID
columns. For best results, a bed height of 5 to 30 cm should be used. Since the
slurry volume may exceed the column volume, a packing reservoir may be
necessary.
1. Make a 50–70% slurry of ligand-coupled Profinity epoxide resin in degassed
binding buffer of choice. Resuspend the slurry by gently swirling or stirring
with a glass or plastic rod. Do not use a magnetic stirrer.
2. Connect the packing reservoir to the column with the outlet valve closed.
3. Fill the column to about 10% of its volume with buffer. Ensure that the bed
support is fully hydrated and free of any bubbles.
4. Pour the slurry into the reservoir and insert the flow adaptor. Allow buffer to
back-flow out of the adaptor to remove trapped air.
5. To pack the column bed, open the column outlet and pump 5–10 CV of
buffer through the column at 125–200% of the selected operating flow rate.
For optimal performance, pack the column at the maximum flow rate allowed
by the column hardware and resin.
6. Turn off the pump and close the column outlet. Reposition the flow adaptor
firmly against the bed. Additional pumping may be required for the final
adjustment of the flow adaptor.
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