HeLa Whole Cell Lysate (TNFa Stimulated)
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- Protein Species
- Human
- Species of Lysate
- Human Cells
- Application
- Western Blotting (WB)
- Specificity
- The cells were grown in Dulbecco’s medium supplemented with 10% fetal bovine serum. Cells were treated with 0.2 µg/ml TNF-alpha for 30 min. 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.
- Characteristics
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Cell Line: HeLa - Human epidermoid carcinoma
Induction: Tumor Necrosis Factor-alpha (0.2 µg/ml) - Lysed Cells
- HeLa Cells
- Lysate Type
- Cell Lysate
- Lysate Fraction
- Whole Cell Lysate
- Lysate Treatment
- TNF-α stimulated
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VNN1 293T Cell Transient Overexpression Lysate(Denatured)
VNN1 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Human
CYP26A1 293T Cell Transient Overexpression Lysate(Denatured)CYP26A1 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Human
APLP2 293T Cell Transient Overexpression Lysate(Denatured)APLP2 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Human
BIRC3 293T Cell Transient Overexpression Lysate(Denatured)BIRC3 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Human
BAG1 293T Cell Transient Overexpression Lysate(Denatured)BAG1 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Human
Hep2 Whole Cell LysateWB Whole Cell Lysate Cell Lysate HepG2 Cells Reactivity: Human
NIH/3T3 Whole Cell Lysate (PDGF Stimulated)WB Whole Cell Lysate Cell Lysate 3T3 Cells PDGF Stimulated Reactivity: Mouse
DYKDDDDK Positive Control LysatePC, WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells Reactivity: Tag
A431 Whole Cell Lysate (EGF Stimulated)WB Whole Cell Lysate Cell Lysate A431 Cells EGF treated Reactivity: Human
CD177 (human): 293T LysateCD177 WB Whole Cell Lysate Overexpression Lysate HEK 293T Cells
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- Application Notes
- 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.
- Comment
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Lysate Fractionation: Whole Cell Lysate
Lysate Stimulation: TNF alpha
Lysate Tissue Culture: Tissue Culture - Restrictions
- For Research Use only
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- Format
- Liquid
- Concentration
- 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)
- Handling Advice
- 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.
- Storage
- -80 °C
- Expiry Date
- 3 months
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- Background
- 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.
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