Bio-Rad ReadyPrep Protein Extraction Kit (Cytoplasmic/Nuclear) User Manual
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4.2 Protocol B
4.2.1 Extraction of Cytoplasmic/Nuclear Proteins From
Tissue Samples
Note: For best results, use a Wheaton Dounce tissue
homogenizer or Potter-Elvehjem tissue grinder. See Section 2 for
catalog information.
1. Chill the Dounce in ice before beginning. Add ~50 mg
of tissue into the chilled Dounce homogenizer. Add
0.75 ml of CPEB to the tissue (~15 ml/gm of tissue).
Notes: If using a tight-fitting microcentrifuge-pestle
combination, use 0.5 ml of CPEB per 50 mg of tissue and
completely homogenize the tissue (5–15 gentle strokes).
Proceed to step 3.
Protease inhibitors may be added to CPEB immediately
prior to use to prevent proteolysis during extraction.
Insufficient volume of CPEB may result in poor cell lysis,
low cytoplasmic protein yield, and contamination of the
nuclear pellet with cytoplasmic proteins.
2. If using a Dounce homogenizer, break up the tissue
with 5–15 strokes using the loose-fitting pestle. Then,
complete the release of the nuclei from the cells with
8–10 strokes using the tight-fitting pestle. If using a
Potter-Elvehjem type tissue grinder, homogenize the
tissue completely with 15–25 strokes of the tight-fitting
pestle. In either case, do not twist the pestle while
raising and lowering, as this may rupture the nuclei
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