HDAC Activity Sex Differences

Materials

EpiQuik Nuclear Extraction Kit Epigentek OP-0002-1 100 Extractions $184.00

Epigenase HDAC Activity/Inhibition Direct Assay Kit (Colorimetric) Epigentek P-4034-96 96 assays $415.00

Tissue Nuclear Extraction

For Tissue samples

1. Weigh tissue and cut it into small pieces.

2. Add to 2ml homogenizer tube with NE1 buffer (5000 ul/mg of tissue)

3. Homogenize (50-60 strokes).

4. Incubate on ice for 15mins

5. Centrifuge for 10mins at 12000 rpm at 4C

6. Remove the supernatant

Nuclear Extract Preparation

1. Add 2 volumes (based on pellet size) of NE2 containing DTT and PIC to nuclear pellet (2500 ul / mg of initial tissue)

2. Incubate the extract on ice for 15mins with vortex (5secs) every 3 mins. (The tissue extract can be further sonicated for 3 x 10 seconds to increase nuclear protein extraction).

3. Centrifuge the suspension for 10mins at 14000 rpm at 4C and transfer the supernatant into a new micro-centrifuge vial.

4. Use immediately or aliquot and freeze supernatant at -80C until further use. Be sure to save aliquot for protein concentration.

5. Measure the protein concentration of the nuclear extract