Cell Plasma Membrane Protein Isolation Kit,IC-7920

Product Description  

The Cell Plasma Membrane Protein Isolation Kit is a high-efficiency solution for rapid isolation of plasma membrane proteins and cellular components (total membrane, cytosol, nucleus, and organelles) from cultured cells or tissues. Utilizing innovative filtration technology and differential centrifugation, the kit eliminates variability associated with traditional homogenization and density gradient methods. It is compatible with downstream applications such as SDS-PAGE, immunoblotting, ELISA, and more.  

Key Features  

1. Fast & Efficient: Complete protocol in under 45 minutes.  

2. High Purity: Proprietary filtration minimizes nuclear contamination, yielding highly enriched plasma membrane fractions.  

3. Multi-Fraction Isolation: Simultaneously isolates five cellular components (total membrane, plasma membrane, cytosol, nucleus, and organelles).  

4. Broad Compatibility: Works with cultured cells and tissue samples for diverse applications.  

5. User-Friendly: Requires only a standard tabletop microcentrifuge.  

Protocol Summary  

A. Total Membrane Protein Isolation  

1. Sample Preparation  

   - Cultured Cells: Harvest 1¨C50 ¡Á 10⁶ cells, wash with PBS, and resuspend in Buffer A (200¨C500 µL). Incubate on ice for 5¨C10 min, then vortex for 10¨C30 sec.  

   - Tissue Samples: Homogenize 10¨C30 mg fresh or 20¨C30 mg frozen tissue in 200 µL Buffer A using a plastic rod. Add 300 µL Buffer A, mix, and incubate on ice for 5 min.  

2. Filtration & Centrifugation  

   - Transfer lysate to a pre-chilled filter cartridge and centrifuge at 16,000 ¡Á *g* for 30 sec. Discard the filter.  

3. Fractionation  

   - Centrifuge at 700 ¡Á *g* for 1 min (pellet = nuclear fraction). Transfer supernatant to a new tube.  

   - Centrifuge at 16,000 ¡Á *g* for 10¨C30 min (supernatant = cytosol; pellet = total membrane fraction).  

B. Plasma Membrane Protein Isolation  

4. Plasma Membrane Purification  

   - Resuspend total membrane pellet in 200 µL Buffer B, centrifuge at 7,800 ¡Á *g* for 5 min (pellet = organelle membranes).  

   - Transfer supernatant to a new tube, add 1.6 mL PBS, and centrifuge at 16,000 ¡Á *g* for 30 min (pellet = purified plasma membrane).  

5. Solubilization  

   - Dissolve the plasma membrane pellet in detergent-containing buffers (e.g., for SDS-PAGE, ELISA, or MS).  

Important Notes  

1. Storage: Store Buffer A and B at ¨C20¡ãC. Thaw completely and keep on ice before use.  

2. Temperature Control: Perform all steps at 4¡ãC (use a refrigerated centrifuge or cold room).  

3. Inhibitors: Add phosphatase inhibitors (e.g., PhosStop) for phosphorylation studies; protease inhibitors are optional.  

4. Sample Load: Avoid overloading the filter cartridge. For sensitive cells (e.g., Jurkat), dilute Buffer A 1:1 with PBS.  

5. Quality Control: Use Na+/K+-ATPase antibody (Cat# IN-SM005AB) as a plasma membrane marker. For total protein extraction, use Minute™ Total Protein Extraction Kit (Cat# SD-001/SN-002).  

Troubleshooting  

Safety Information

Please wear gloves, lab coat and goggles while operating. Prevent contact product directly. In case of contacting, wash with large amount of water.

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