Waters Advanced Purification Glass Columns - Manual (Page 4)

[ Care and Use ManUal ]
Waters Advanced Purification Columns 4
Preparing slurry solutions
To make slurry solutions, suspend one part of packing material
in three parts of degassed solution. The packing funnel can hold
40 mL of slurry.
Slurry-packing
To slurry-pack the column (Figure 4):
1. Unscrew the inlet connector from the inlet cap.
2. Screw the packing funnel into the inlet cap.
3. Clamp the column upright to a rigid support. Drain the outlet tubing to
waste.
4. Fill the column with the slurry. If the slurry does not flow into the
column, insert a pipette into the tip of the funnel to break the surface
tension.
5. When all of the packing material is in the column, plug the outlet
tubing.
6. Let the packing material settle. Remove excess solvent from the top of
the bed with a pipette. Do not disturb the bed.
7. Remove the funnel, and reattach the inlet connector.
8. Adjust the inlet connector until the plunger contacts the packed bed.
Ensure that the packed bed does not exceed the nominal column
length.
Figure 4: Slurry-Packing the Column
Dry-Packing the Column
Amount of packing material required
To determine the amount of packing material required, multiply the
column volume, V (mL), by the bulk density of the packing material,
d (g/mL):
Amount of packing material required = V x d
Waters Protein-Pak HR must be slurry-packed (see “Slurry-packing the
column”)
Determining density
To determine the ulk density of the packing material, put 19 g of
packing material into a 10 mL graduated cylinder. Tap the graduated
cylinder on a flat surface until the material is completely settled.
Note the volume and calculate the density.
Dry-packing
To dry-pack the column:
1. Unscrew the fine adjustment knob from the inlet cap.
2. Screw the packing funnel into the inlet cap.
3. Remove the outlet tubing.
4. Holding the column vertically, gradually fill the column with packing
material. While filling, tap the column lightly on a flat surface until the
material settles.
5. Remove the funnel and reattach the inlet connector and the outlet
tubing.
6. Adjust the inlet connector until the inlet plunger contacts the packed
bed.
Ensure that the packed bed does not exceed the nominal column
length.
Packing Material Solution
Ion-exchange chromatography High concentration buffer
Reversed-phase chromatography Methanol
Hydrophobic-interaction
chromatography (HIC) Water or low concentration buffer
Slurry
Funnel
Inlet cap
To waste
Packing Material Bulk Density of Packing Material
Waters AccellPlus silica-based
packing materials 0.5 g/mL
Other packing materials Use procedure below to determine density