SKOV-3 (human adenocarcinoma) Nuclear extract lysate, Denatured; Abnova

Description

Western Blotting

Specifications

• Tissue: Ovary
• For Use With (Application): Western Blot
• Description: Nuclear extract cell lysate (denatured)
• Lysis Buffer: Buffer A: 10mM HEPES pH 7.9, 1.5mM MgCl2, 10mM KCl, 0.5 mM DTT. Buffer C: 20mM HEPES pH 7.9, 25%(v/v) Glycerol , 0.42M NaCl , 1.5mM MgCl2, 0.2 mM EDTA, 0.5 mM DTT & 0.5 mM PMSF. Buffer D : 20mM HEPES pH 7.9, 20%(v/v) glycerol, 50mM KCl, 0.2 mM EDTA, 0.5 mM DTT & 0.5 mM PMSF.
• Preparation Method: Nuclear extract was prepared by using a modified protocol of Dignam et al. Cells were Harvested and homogenized in Buffer A, and then centrifugated at 25,000 g for 20 minutes to remove cytoplasm and pellet the nuclei. The pellet was re-suspended in Buffer C, and then the suspensions were centrifuged to collect nuclear extract. The supernatant was dialyzed against Buffer D. The dialysate was then centrifuged, divided into aliquots, and stored at -80°C. The protein concentration was determined by the method of Bradford (Bio-Rad protein assay, microplate standard assay). The lysate was adjusted to 2.5 mg/ml, and then mixed with 5X Sample Buffer to become final 2 mg/ml in 1X Sample Buffer. The lysate was heated at 95°C for 5 min, and cooled rapidly.
• Quality Control Testing: 12.5% SDS-PAGE Stained with Coomassie Blue.
• Quantity: 50 μg
• Storage Buffer: In ready-to-use 1X Sample Buffer (50 mM Tris-HCl, 2% SDS, 10% glycerol, 300 mM 2-mercaptoethanol, 0.01% Bromophenol blue).
• Host Species: Human
• Content And Storage: Store at -80°C. Aliquot to avoid repeated freezing and thawing.