HeLa Whole Cell Lysate Etoposide Stimulated
HeLa Whole Cell Lysate - SDS-Page
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Datasheet

HeLa Whole Cell Lysate Etoposide Stimulated

W09-001-A80
500 µg
SDS-PAGE
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Description

Background

Ready-to-use whole cell lysates produced by Rockland Immunochemicals 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.

Application Note

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.

Purity/Specificity

The cells were grown in DMEM supplemented with 10% FBS (Fetal Bovine Serum). Cells were treated with 10 µg/ml Etoposide for 3 h. Cells were washed in PBS and incubated on ice in modified RIPA buffer containing 150 mM sodium chloride, 50 mM Tris Cl, 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 is 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 and 1 µM Pepstatin A). Phosphatase inhibitors sodium fluoride, sodium orthovanadate, sodium pyrophosphate and -glycerophosphate were also added. Cell debris was removed by centrifugation. Protein concentration was determined by Lowry assay using a commercially available kit. The protein concentration was adjusted to 2 mg/ml and then an equal volume of 2X SDS-PAGE sample buffer was added.

HeLa Whole Cell Lysate - Etoposide Stimulated - W09-001-A80
HeLa Whole Cell Lysate Etoposide Stimulated, HeLa Etoposide Stimulated Whole Cell Lysate, HeLa Etoposide Stimulated WCL, HeLa WCL Etoposide Stimulated
SDS-PAGE
Human
HeLa - Human epidermoid carcinoma
Whole Cell Lysate
Etoposide
Tissue Culture
Etoposide (10 µg/ml)
Liquid (sterile filtered)
1.0 mg/mL by modified Lowry assay
1X SDS-PAGE Sample Buffer (62.5 mM Tris HCl, 2% SDS, 10% Glycerol and 0.005% bromophenol blue, pH 6.8)
None
10% (v/v) Glycerol
Dry Ice
Store vial at -70° C or COLDER. For extended storage, aliquot contents to minimize freeze/thaw cycles.
Expiration date is three (3) months from date of opening.

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This product is for research use only and is not intended for therapeutic or diagnostic applications. Please contact a technical service representative for more information. All products of animal origin manufactured by Rockland Immunochemicals are derived from starting materials of North American origin. Collection was performed in United States Department of Agriculture (USDA) inspected facilities and all materials have been inspected and certified to be free of disease and suitable for exportation. All properties listed are typical characteristics and are not specifications. All suggestions and data are offered in good faith but without guarantee as conditions and methods of use of our products are beyond our control. All claims must be made within 30 days following the date of delivery. The prospective user must determine the suitability of our materials before adopting them on a commercial scale. Suggested uses of our products are not recommendations to use our products in violation of any patent or as a license under any patent of Rockland Immunochemicals, Inc. If you require a commercial license to use this material and do not have one, then return this material, unopened to: Rockland Inc., P.O. BOX 5199, Limerick, Pennsylvania, USA.

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