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Protein Electrophoresis & Analysis

ProteoJET™ Mammalian Cell Lysis Reagent

Product information

Features

• High yield total protein extraction from mammalian cultured cells or tissue samples.
• Ready-to-use formulation.
• Convenient – room temperature.
• Easy-to-use – no sonication, freeze-thaw cycles.
• Compatible with downstream applications:
  – protein quantification,
  – 1D and 2D electrophoresis,
  – immunoprecipitation and Western blotting,
  – EMSA, enzymatic activity and reporter gene assays.

Applications

• Extraction of total native, non-denatured protein from mammalian tissues or cultured cells (adherent and suspension).

Description

ProteoJET™ Mammalian Cell Lysis Reagent is a ready-to-use solution designed for extraction of native, non-denatured proteins from mammalian cultured cells or tissue samples. The procedure, which is based on lysis with mild detergents at room temperature, is fast and simple: no need for sonication, freeze thaw cycles or scraping of adherent cells.

The procedure yields proteins in their native, non-denatured functional state. Therefore, isolated proteins can be used directly in many proteomic and enzymatic activity applications, including immunoprecipitation, affinity purification, reporter gene assays and electrophoretic mobility shift assays (EMSA). Isolated proteins are also compatible with quantification assays such as Bradford, Lowry and the BCA.

Quality Control

Reagent is functionally tested for total protein extraction from cultured mammalian suspension cells.

Storage

Store at 4°C or at room temperature. Stable for at least 1 year at room temperature.

Figure 1. Total protein extraction.
Protein was extracted from various cell lines (A) and mouse tissues (B). Extracts were separated in 12% SDS-PAGE, gel stained with PageBlue™ Protein Staining Solution.
A
1 – HEP2C cells
2 – human primary fibroblast cells
3 – NIH3T3 cells
4 – HEK293 cells
5 – Cos7 cells
6 – HeLa cells
7 – B50 cells
8 – CHO cells
9 – Raji cells
10 – Jurkat cells
B
11 – liver
12 – kidney
13 – spleen
14 – lung
15 – muscle

Figure 2. Comparison of total protein yields from 5x10⁶ cells.

Patents, Licenses, Trademarks

Protocols & recommendations

RECOMMENDATIONS FOR USE

ADDITIONAL PROTOCOLS

Protein Samples: Extraction and Quantification Guides

Sample/procedure	Extraction	Quantification
Mammalian cell and tissue samples	Extract total proteins using ProteoJET™ Mammalian Cell Lysis Reagent. Extract nuclear and cytoplasmic proteins using ProteoJET™ Cytoplasmic and Nuclear Protein Extraction Kit.	Use Bradford Reagent, ready-to-use for protein quantification. Easily create standard curves by using one of our convenient protein standard sets: Bovine Serum Albumin Standard Set, Bovine Gamma Globulin Standard Set, Protein Standard Solution. Create a standard curve by plotting the 595 nm values (y-axis) versus their concentration in µg/ml (x-axis). Use the standard curve to determine the protein concentration of each unknown sample.
Bacterial samples	To analyze total bacterial proteins in SDS-PAGE, cells can be treated directly with DualColor™ Protein Loading Buffer Pack or Protein Loading Buffer Pack.
Lyophilized proteins	To analyze proteins in SDS-PAGE, treat lyophilized proteins directly with DualColor™ Protein Loading Buffer Pack or Protein Loading Buffer Pack.

Protein Samples: Preparation for Loading on SDS-PAGE

Step	Sample preparation with the DualColor™ Protein Loading Buffer Pack	Sample preparation with the Protein Loading Buffer Pack
Thaw	Thaw the buffer pack components either at room temperature or at 37°C for a few minutes to dissolve precipitates.
Mix	Vortex gently, but thoroughly to ensure that the solution is homogeneous.
Dilution	2.0 µl of 20X Reducing Agent	2.5 µl of 20X Reducing Agent
	Protein sample (~0.5 ng – 2.5 µg) For Western blots or gels to be treated with Coomassie based stains, use up to 2.5 µg of total protein per minigel well. For silver staining applications use up to 10 ng of total protein per minigel well.
	10 µl of 4X DualColor™ Protein Loading Buffer Water, nuclease-free to 40 µl*	10 µl of 5X Protein Loading Buffer Water, nuclease-free to 50 µl*
Denaturation	Incubate samples at 95-100°C for 5 minutes.
Loading	Spin for a few seconds in a microcentrifuge. Apply directly to an SDS-polyacrylamide gel. Use ~10 µl per minigel well.

* The sample volume can be scaled up or down.

Related products

• ProteoBlock™ Protease Inhibitor Cocktail
• ProteoJET™ Membrane Protein Extraction Kit
• ProteoJET™ Cytoplasmic and Nuclear Protein Extraction Kit
• Bradford Reagent, ready-to-use
• Bovine Gamma Globulin Standard Set, ready-to-use
• Bovine Serum Albumin Standard Set, ready-to-use
• Protein Standard Solution
• Spectra™ Multicolor High Range Protein Ladder
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• Unstained Protein Molecular Weight Marker
• PageSilver™ Silver Staining Kit
• PageBlue™ Protein Staining Solution
• DualColor™ Protein Loading Buffer Pack
• Protein Loading Buffer Pack
• 10X Tris-glycine Electrophoresis Buffer
• 10X Tris-tricine-SDS Electrophoresis Buffer
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• DTT