Bio-Rad ProteoMiner Protein Enrichment Kits - Manual (Page 11)

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5. Incubate column at room temperature, lightly vortex several times over a
period of 15 min.
6. Remove caps, place in a clean collection tube labeled E1 and centrifuge
at 1,000 x g for 30–60 sec. This elution contains your eluted proteins. Do
not discard.
7. Repeat steps 5–6 two more times. (Elutions may be pooled or analyzed
individually. If analyzed individually, you will need additional collection
tubes not provided with kit.)
8. Store elution at –20°C or proceed with downstream analysis. (For best
results, we recommend a clean up of your sample prior to analysis. See
Section 11 for more information on preparing sample for analysis.)
Section 8
Instructions for Use With Sequential
Elution Kit (Large-Capacity)
The ProteoMiner™ Sequential Elution large-capacity kit combines the
ProteoMiner kit (catalog #163-3007) and the ProteoMiner sequential elution
reagents (catalog #163-3003) and is available for researchers using SELDI or
other downstream protein separation analysis methods who wish to access
additional proteins. This kit is NOT compatible with 2-D gel electrophoresis.
This protocol has been optimized for plasma and serum samples with
protein concentration of >50 mg/ml (requires total protein load >50 mg). For
other sample types, please refer to Section 5: Sample
Considerations.
Step 1 – Column Preparation
(Reagents and columns required for this step are included in the ProteoMiner
kit, catalog #163-3007.)
Vacuum (at 16 mm Hg) can replace centrifugation for column preparation,
sample binding and sample wash steps if desired. (Vacuum manifold is
available through Bio-Rad, catalog #732-6470.)
1. First remove the top cap and then snap off the bottom cap from each of
the spin columns you will be using.
Note: Do not discard top or bottom caps, they will be reused
throughout the protocol. If beads settle in top cap, replace after removing
bottom plug and centrifuge with top cap on column. To use bottom cap
as a plug, invert and firmly place in bottom of spin column.
2. Place the column in a capless collection tube and centrifuge at 1,000 x g
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