Bio-Rad ProteoMiner Protein Enrichment Kits - Manual (Page 12)

for 30–60 sec to remove the storage solution. Discard collected material.
Note: Kit contains one capless collection tube per spin column for the
following steps: column preparation, sample binding, and sample wash.
Kit contains one capped collection tube per spin column to be used for
the elution step, allowing for easy storage of your eluted sample.
3. Replace the bottom cap and add 600 µl wash buffer, then replace top cap.
4. Rotate column end-to-end several times over a 5 min period.
5. Remove bottom cap, place the column in a capless collection tube and
centrifuge at 1,000 x g for 30–60 sec to remove water. Discard collected
material.
6. Repeat steps 3 and 4.
7. Remove bottom cap, place the column in a capless collection tube and
centrifuge at 1,000 x g for 30–60 sec to remove the wash buffer. Discard
collected material.
8. Replace bottom cap on spin column. The column is now ready for sample
binding.
Step 2 – Sample Binding
Samples should be free of precipitate. Centrifuge samples at 10,000 x g for
10 min to clarify. Take precautions to avoid the bottom aggregate proteins and
top lipid layer when recovering your sample. It is recommended that at least
1 ml of sample (protein concentration
≥
50 mg/ml) is added to the column,
lower volumes may not achieve optimal results. For other sample types,
please refer to Section 5: Sample Considerations.
If using plasma: Add 900 µl plasma (nonheparinized) and 100 µl plasma
preparation buffer to spin column. Replace top cap and rotate column on a
platform or rotational shaker for 2 hr at room temperature. After approximately
1 hr of binding you may see clumping; this is expected and will not negatively
impact your sample preparation. Heparinized plasma is not compatible with
this kit.
For all other sample types: Add 1 ml of sample to column and replace
top cap. Rotate column on a platform or rotational shaker for 2 hr at room
temperature.
Step 3 – Sample Wash
(Reagents required for this step are included in the ProteoMiner kit, catalog
#163-3007.)
1. Remove bottom cap, place column in a capless collection tube and
centrifuge at 1,000 x g for 30–60 sec. Discard collected material.
Centrifuge again at 1,000 x g for 30–60 sec to remove any remaining
material. Discard collected material.
10
10010636D:4110137B-4.0.GR.qxd 3/12/2009 7:51 AM Page 10