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Cell were then harvested, centrifuged and their pellets were permeabilized for 5 min on ice with 200 g/ml digitonin (Sigma-Aldrich GmbH, Buchs, Switzerland) dissolved in 200 l digitonin buffer (250 mM sucrose 70 mM KCl, 137 mM NaCl, 4.3 mM Na 2 HPO 4 , 1.4 mM KH 2 PO 4, pH 7.2 ) and protease inhibitor cocktail tablets (Roche Diagnostics GmbH, Mannheim, Germany)
It is completely normal to feel a bit anxious for results, especially with all the buzz surrounding these medications
To denature the proteins, 120 L of 8 M urea in 50 mM Tris-HCl (pH 8) was added
Caspase-3 and NF-B expressions were evaluated immunohistochemically
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