EpiQuik™ Total Histone Extraction Kit for Tissues (Treated and Untreated
Re-suspend cell/tissue pellet in 3 volumes (approximately 200 µl/107 cells or 100 mg of tissue) of Lysis Buffer and incubate on ice for 30 min.
Centrifuge at 12,000 rpm for 5 min at 4°C and transfer the supernatant fraction (containing acid-soluble proteins) into a new vial.
Prepare Balance-DTT Buffer by adding DTT Solution to Balance Buffer at a 1:500 ratio (e.g., 1 µl of DTT Solution + 500 µl of Balance Buffer).
Add 0.3 volumes of the Balance-DTT Buffer to the supernatant immediately (e.g., 0.3 ml of Balance-DTT Buffer to 1 ml of supernatant).
Quantify the protein concentration with an OD reading.
BSA can be used as a standard.
Aliquot and store the extract at –20°C for several days, or –80°C for long-term storage.
Avoid repeated thawing and freezing.
Weigh the sample and cut the sample into small pieces (1-2 mm3 ) with a scalpel or scissors.
Transfer tissue pieces to a Dounce homogenizer.
Dilute 10X Pre-Lysis Buffer into 1X Pre-Lysis Buffer with distilled water at a 1:10 ratio (e.g., 1 ml of 10X Pre-Lysis Buffer + 9 ml of water).
Add the Diluted 1X Pre-Lysis Buffer at 1 ml per 200 mg of tissue, and disaggregate tissue pieces by 50-60 strokes.
Transfer homogenized mixture to a 15 ml conical tube and centrifuge at 3000 rpm for 5 min at 4°C.
Remove supernatant.
If total mixture volume is less than 2 ml, transfer mixture to a 2 ml vial and centrifuge at 10,000 rpm for 1 min at 4°C.
