MDA-MD-435S Whole Cell Lysate

• Cat.No.: MDA-MD-435S-01HL
• Product Overview: MDA-MD-435S Whole Cell Lysate

Specification

• Species: Human
• Source: mammary gland ductal carcinoma
• Lysate preparation: The cells were grown in Leibovitz''s L-15 medium with 0.01 mg/mL insulin, 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 and 0.1% SDS, 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 2× SDS-PAGE sample buffer was added.
• Recommended Usage: Ready-to-use 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).
• Storage: Store at 2-8 centigrade for continuous use. For extended storage, freeze working aliquots at -70 centigrade. Repeated freezing and thawing is not recommended. Under proper storage conditions the shelf life is half a year from the date of receipt.
• Storage Buffer: 1X SDS-PAGE Sample Buffer (62.5 mM Tris HCl, 2% SDS, 10% Glycerol and 0.005% bromophenol blue, pH 6.8)
• Concentration: 2 mg/mL