Bio-Rad Affi-Gel Protein A Media - Manual (Page 4)

1.3 Additional Items Required
Column rack The Econo-Pac 10 (12 place) acrylic rack is
ideal for holding the Econo-Pac columns.
Other benchtop or lattice mount racks may
also be used.
Test tubes General purpose tubes for fraction collection
are recommended.
pH meter A pH meter is required to check the pH of
the binding and elution buffers after reconsti-
tution.
Mixer Standard laboratory magnetic stirrer and bar
for buffer mixing.
Balance Standard laboratory scale for weighing out
buffer solids.
Filters 0.22 micron filters for buffer preparation.
1.4 Buffer Preparation
Binding Buffer Preparation
The binding buffer is supplied as a premixed, preweighed solid.
Reconstitution and filtration are required prior to use. Dissolve 31.4 g
binding buffer solids in distilled, deionized water for a final volume
of 100 ml. (Use the full 471 g for 1,500 ml.) Stir for 10 minutes.
Filter through a 0.22 µm nylon filter and check the pH. The pH
should be 9.0 ± 0.2. If the pH is not in this range, adjust the pH with
10 N NaOH or 6 N HC1. Store buffer solids at room temperature.
Stoer reconstituted buffer at 4 °C. If desired, sodium azide may be
added to 0.05% (w/v).
Elution Buffer Preparation
The elution buffer is supplied as a preweighed, premixed solid.
Reconstitution and filtration are required prior to use. These salts
are hygroscopic. Any material in clumps should be broken up before
you weight the solids. Dissolve 2.2 g in distilled, deionized water for
a final volume of 100 ml (use the full 25 g for 1,100 ml). Stir for 10
minutes. Filter through a 0.22 µm filter and check the pH. The pH
should be 3.0 ± 0.2. If the pH is not in this range, adjust the pH with
10 N NaOH or 6 N HC1. Store buffer solids at room temperature.
Reconstituted buffer should be stored at 4 °C, and if desired,
Thimerosal bacteriostat may be added to a final concentration of
one part per 10,000. The use of sodium azide in low pH buffers is haz-
ardous.
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