Bio-Rad Bio-Beads SM-2 Adsorbents - Manual (Page 6)

Section 4
Sample Protocols
4.1 Triton X-100 Detergent Removal
Triton X-100 detergent is useful for membrane
solubilization. However, the usefulness of Triton X-100
detergent has been limited by the difficulty experienced
in removing the detergent from the sample. Holloway
reported a simple, rapid, and mild procedure for
removing Triton X-100 detergent on columns of Bio-
Beads SM-2 adsorbents.
1 The following protocol is based
on Holloway’s published procedure. Refer to his paper
for complete details.
1. Pack a 1 x 8 cm column (or 1 x 10 cm Econo-
Column
® chromatography column) with 5 g moist
Bio-Beads SM-2 adsorbent. Equilibrate with 10 mM
potassium phosphate, pH 7.2, at 4 °C. (1 g of Bio-
Beads SM-2 adsorbent will adsorb 0.07 g of Triton
X-100 detergent.)
2. Layer a 1 ml sample of protein solution onto the top
of the bed.
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3.3 Batch Method
The batch method is the addition of adsorbent
directly into the sample followed by stirring to achieve
the adsorption. This is a very simple and common
method, though it requires 90–120 minutes, as opposed
to 15 minutes for the column procedure.
1. Weigh 5 g of Bio-Beads SM adsorbent for every 25
ml of solution.
2. Add the adsorbent to the solution.
3. Use either a stir bar or a mechanical agitator to mix
the solution at room temperature for 2 hours.
4. The sample may be recovered by decanting, or by
removing it with a pipette. For smaller samples, it
may be more convenient to centrifuge the sample
before pipetting.
Both columns and batches can be stored. For short
term storage (<1 week), use the elution solvent. For
longer storage, use a 0.05% sodium azide solution or a
20% solution of an organic solvent such as methanol or
isopropanol to prevent microbial growth.
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LIT195B 6/17/98 11:37 AM Page 6