Bio-Rad DEAE Affi-Gel Blue Gel - Manual (Page 4)

1.4 Buffer Preparation
Human Application Buffer Preparation
The human buffer is supplied as a premixed, preweighed solid. Reconstitution
and filtration are required prior to use. Dissolve 1.3 grams human buffer solids
per 300 ml (final volume) distilled, deionized water. (Use the full 13 grams for
3 liters final volume.) Filter through a 0.45 µm filter and check the pH. The pH
should be 8.0 ± 0.2. If the pH is not in this range, adjust the pH with 10 N KOH
or 6 N HCl. Store buffer solids at room temperature. Store reconstituted buffer
at 4 °C. If desired, sodium azide may be added to 0.05% (w/v).
Rabbit Application Buffer Preparation
The rabbit buffer is supplied as a premixed, preweighed solid. Reconstitution
and filtration are required prior to use. Dissolve 1.3 grams rabbit buffer solids
per 300 ml (final volume) distilled, deionized water. (Use the full 13 grams for
3 liters final volume.) Filter through a 0.45 µm filter and check the pH. The pH
should be 8.0 ± 0.2. If the pH is not in this range, adjust the pH with 10 N
NaOH or 6 N HCl. Store buffer solids at room temperature. Store reconstitut-
ed buffer at 4 °C. If desired, sodium azide may be added to 0.05% (w/v).
Regeneration Buffer Preparation
The regeneration buffer is supplied as a premixed, preweighed solid.
Reconstitution and filtration are required prior to use. Dissolve 12.2 grams per
100 ml (final volume) distilled, deionized water. (Use the full 121 grams for 1 liter
final volume.) Filter through a 0.45 µm filter. No pH adjustment is necessary. Store
buffer solids at room temperature. Store reconstituted buffer at 4 °C.
1.5 Sample Preparation
Sample preparation is simplified by using the Econo-Pac 10DG columns. Each
Econo-Pac 10DG column can process up to 3 milliliters of serum or ascites fluid
per cycle. The columns can be re-used. It is recommended that one Econo-Pac
10DG column be used for each (different) serum to avoid cross contamination.
To prepare up to 3 ml of serum for purification on the serum IgG purifica-
tion columns:
1. Discard the buffer above the top frit of one Econo-Pac 10DG column.
2. Add 20 ml of application buffer (either human or rabbit buffer) to the column
(fill to the top), and snap off the bottom tip to start the column flowing.
3. Allow the buffer to drain to the top frit. The column will not run dry. Flow
will stop when the buffer level reaches the top frit.
4. Add 3.0 ml of serum to the column. If the serum sample is less than 3.0
ml, add application buffer to reach a total sample volume of 3.0 ml.
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