MCF-7 Whole Cell Lysate (H2O2 Stimulated)

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Proteinspezies
Human
Lysatspezies
Human Cells
Applikation
Western Blotting (WB)
Optionen
Hersteller
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Spezifität The 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.
Produktmerkmale Cell Line: Human Epithelial (Mammary)
Induction: 300 µM Hydrogen Peroxide (H2O2)
Sterilität Sterile filtered
Lysate Fraction Whole Cell Lysate
Lysate Treatment H2O2 Stimulated
Lysate Type Cell Lysate
Lysed Cells MCF7 Cells
Hintergrund Ready-to-use whole cell lysates produced are derived from cell lines or tissues using highly refined extraction protocols to ensure exceptionally high quality, protein integrity and lot-to-lot reproducibility.  All extracts are tested by SDS-PAGE using 4-20% gradient gels and immunoblot analysis using antibodies to key cell signaling components to confirm the presence of both high molecular weight and low molecular weight proteins.
Applikations-hinweise 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). 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.
Kommentare

Lysate Fractionation: Whole Cell Lysate
Lysate Stimulation: H2O2
Lysate Tissue Culture: Tissue Culture

Beschränkungen Nur für Forschungszwecke einsetzbar
Format Liquid
Konzentration 1.0 mg/mL
Buffer 1X SDS-PAGE Sample Buffer (62.5 mM Tris HCl, 2% SDS, 10% Glycerol and 0.005% bromophenol blue, pH 6.8)
Handhabung 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). 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.
Lagerung -80 °C
Haltbarkeit 3 months
Bilder des Herstellers
 image for MCF-7 Whole Cell Lysate (H2O2 Stimulated) (ABIN964026) MCF-7 Whole Cell Lysate (H2O2 Stimulated)