Bio-Rad ReadyPrep™ 2-D Cleanup Kit - Manual (Page 12)

reduce possible spilling during vortexing and the incubation
at -20°C. A Nalgene Oak Ridge-type tube made of
polypropylene copolymer is an example of an appropriate
tube. The volume of the tube should be at least 12 times
the original volume of the sample.
Step 2. For each volume of sample use 3 volumes of
precipitating agent 1.
Step 3. For each volume of sample use 3 volumes of
precipitating agent 2.
Step 7. Wash the protein pellet by adding 3–4 times the
pellet volume of wash reagent 1.
Step 8. Add enough ReadyPrep proteomic grade water or
other ultrapure water to just cover the protein pellet.
Step 9. Add 1 ml of wash reagent 2 (prechilled at -20°C
for at least 1 hr) for every 0.1 ml of original sample.
Additionally, the volume of wash reagent 2 must be at least
10 times the volume of water used in Step 9. Add 5 µl of
wash 2 additive (use only 5 µl of wash 2 additive
regardless of the volume of sample).
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