HeLa Whole Cell Lysate

• Catalog Number: orb348668
• Category: Tools
• Description: HeLa Whole Cell Lysate
• Tested applications: SDS-PAGE, WB
• Concentration:

  1.0 mg/ml

• Dilution range: WB: User Optimized
• Form/Appearance: Liquid
• Purity: The Hela cells were grown in Dulbecco’s medium supplemented with 10% fetal bovine serum. Cells were washed with PBS and then incubated on ice in modified RIPA buffer, containing 150 mM sodium chloride, 50 mM Tris HCl, pH 7.4, 1 mM EDTA, 1.0% NP-40, 0.5% sodium deoxycholic acid , 0.1% SDS and 0.01% (w/v) sodium azide to lyse the cells. Protein integrity was ensured using a cocktail of protease inhibitors with broad specificity for the inhibition of aspartic, cysteine, and serine proteases as well as aminopeptidases (0.1 mM AEBSF HCl, 0.08 µM Aprotinin, 5 µM Bestatin, 1.5 µM E-64, 2 µM Leupeptin Hemisulfate, 1 µM Pepstatin A). Phosphatase inhibitors 1 mM NaF and 1 mM Na3VO4 were also added. Cell debris was removed by centrifugation. Protein concentration was determined by a modified Lowry assay using a commercially available kit. Protein concentration was adjusted to 2 mg/ml and then an equal volume of 2X SDS-PAGE sample buffer was added.
• Conjugation: Unconjugated
• Hazard Information: Non-Toxic
• Source: Human
• Storage: Store vial at -70° C or COLDER. For extended storage, aliquot contents to minimize freeze/thaw cycles.
• Buffer/Preservatives: Preservative: None. Stabilizer: 10% (v/v) Glycerol. 1X SDS-PAGE Sample Buffer (62.5 mM Tris HCl, 2% SDS, 10% Glycerol and 0.005% bromophenol blue, pH 6.8)
• Alternative names: Hela Cells, HeLa Lysate, Human Derived Whole Cell
• Note: For research use only
• Application notes: ready-to-use HeLa whole cell lysates are especially prepared as positive controls for separation by SDS-PAGE and subsequent western blot analysis. Lysates are prepared in denaturing buffer WITHOUT dissociating agents (i.e. no 2-mercaptoethanol or dithiothreitol has been added). Heat lysate to 95°C for 5 minutes and rapidly cool. If dissociating conditions are desired, add reducing agent prior to heating. The recommended loading volume per lane is 10-20 μl depending on the size format of your gel.
• Expiration Date: 12 months from date of receipt.