Bio-Rad ReadyPrep Protein Extraction Kit (Total Protein) - Manual (Page 8)

2. In a 2.0 ml microcentrifuge tube add 1 ml of complete
2-D Rehydration/Sample Buffer 1 (see Step 1) per
50–100 mg of animal tissue or 0.05 ml of wet cell
pellet from sources such as cell culture, yeast, or bac-
teria. For plant tissue add 2–3 ml of 2-D Rehydration/
Sample Buffer 1 per each gram of tissue in, for
example, a disposable 14 ml round-bottom tube. The
sample-to-buffer volume ratio indicated above is only a
guide and may be adjusted depending upon the
desired scale of the preparation and type of sample
used.
Insufficient volume of 2-D Rehydration/Sample Buffer 1 may
result in poor cell lysis and incomplete solubilization of all
protein types.
Plant tissue should be ground to a fine powder using a
mortar and pestle in liquid nitrogen before addition of 2-D
Rehydration/Sample Buffer 1.
3. Place the sample on ice and sonicate the suspension
with an ultrasonic probe to disrupt the cells and
fragment the genomic DNA. Sonicate the sample
using 30 sec bursts, typically 3–4 times, or until lysis is
complete. Chill the suspension on ice briefly between
each ultrasonic treatment. If necessary, transfer the
extract to a microcentrifuge tube when this step is
complete.
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