Boster Immunoleader

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Membrane Protein Extraction Kit

AR0155

1 kit

100 USD

AR0155

Membrane Protein Extraction Kit

1 kit

100 USD

Western Blotting Related Reagent

Store at 4˚C for one year

This Membrane Protein Extraction Kit can be used to extract the membrane protein efficiently from mammal tissues and cells with optimized buffer and reagent. It can enrich the membrane protein via moderate lysis buffer. Firstly, the cell is lysed by moderate lysis buffer. Then add lysis buffer to dissolve membrane protein by incubating at 37˚C, and separate through water and decontamination phase. Here, the hydrophilic protein dissolved in water phase, while the water-repellent membrane protein is soluble in decontamination phase. In this way, the membrane protein can be extracted successfully. And the extracted membrane protein can be used in the Western Blotting, Two Dimensional Electrophoresis and Enzymatic Activity Analysis etc.

1. Anchorage-dependent cell: Remove the nutrient solution with PBS buffer, or deal it with EDTA solution to loose cells, then stroke cells in pipette (digesting with pancreatin is not recommended, it may degrade the protein). Centrifuge at 500xg for 2 -3min, then collect cells. Try to separate sediment from supernatant for stand-by. 2. Suspending cell: Wash with PBS buffer, centrifuge at 500xg for 2-3min, then collect cells. Try to separate sediment from supernatant for stand-by. 3. Suspend the precipitated cells on the 150 ul of Protein Extraction Solution A, shake and incubate it for 10 min on ice. 4. Place the lysed cell on ice, add 450ul of Protein Extraction Solution B and incubate on ice for 1 hour, vortex frequently. 5. Centrifuge at 10000xg at 4˚C for 10 min, transfer the supernatant to a new tube and give up precipitate. Incubate supernatant at 37˚C for 10 min to separate membrane protein. To improve the separative efficiency, incubation time can be prolonged to 20 min. 6. Centrifuge at 10000xg at room temperature for 10 min to separate the water-repellent membrane protein in decontamination phase from hydrophilic protein in water phase. 7. Transfer the supernatant water phase (bottom is decontamination phase) to a new tube quickly. 8. Place the isolated protein solution on ice, most membrane protein is located in the bottom decontamination phase. Notes The collected membrane protein contains detergent, before WB analysis, it should be diluted and dialysed to remove the detergent. Protein precipitation also can work to remove.

For extracting the membrane protein efficiently from mammal tissues and cells

50-100 assays