Mem-PER™ Plus Membrane Protein Extraction Kit - FAQs

The kit reagents can be scaled up or down as long as the ratio of reagents to cells is maintained. However, we don't have specific guidelines for the range of cell numbers that can be used because protein yields can vary depending on the cell line. It is important to note that the process of scaling the kit reagents up or down and determining the appropriate cell numbers for optimal protein yield will require empirical optimization by the investigator.

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The composition of the Permeabilization Buffer, including the identity and concentration of any detergent therein, is proprietary. However, the Permeabilization Buffer contains a very small amount of a weak non-ionic detergent to facilitate the permeabilization of the cells and release of the cytosolic proteins.

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The Permeabilization buffer in the Mem-PER Plus Membrane Protein Extraction Kit (Cat. No. 89842) creates pores in the plasma membrane of cells that releases cytosolic proteins. The permeabilized cells are centrifuged, and the supernatant contains the cytosolic protein fraction. The pellet is then resuspended in the Solubilization buffer containing a proprietary detergent that solubilizes plasma, mitochondrial, and ER membrane proteins. Subsequent centrifugation yields a supernatant containing solubilized membrane and membrane-associated proteins.

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The Mem-PER Plus Membrane Protein Extraction Kit is not suitable for any organism that has a cell wall, including algae, plants, yeast, or bacteria.

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M-PER Mammalian Protein Extraction Reagent can extract some membrane or cytoskeletal proteins, but the extraction efficiency is not consistent. The reagent was not intended to specifically extract these proteins. We recommend using Mem-PER Plus Membrane Protein Extraction Kit (Cat. No. 89842) for membrane protein extraction or Subcellular Protein Fractionation Kit for Cultured Cells (Cat. No. 78840) for compartmental extraction.

Find additional tips, troubleshooting help, and resources within our Cell Lysis and Fractionation Support Center.

Yes, frozen tissues can be processed with the Mem-PER Plus Membrane Protein Extraction Kit.

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The kit uses a mild detergent-based selective extraction protocol to extract membrane proteins. First, the cells are permeabilized with a mild detergent to allow the release of soluble cytosolic proteins. A second detergent then solubilizes membrane proteins. Extraction efficiencies will vary depending on the number of times the integral membrane protein(s) of interest spans the lipid bilayer. Membrane proteins with at least 1-2 transmembrane domains are typically extracted with an efficiency of up to 90%. Cross-contamination of cytosolic proteins into the membrane fraction is usually less than 10%.

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Yes, membrane proteins from various organelles will fractionate with the membrane fraction. We have evaluated COX-IV (mitochondrial membrane protein) and Calnexin (ER membrane protein) which fractionated with the membrane fraction.

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Different cell lines will give different yields. Some cells may be more difficult to permeabilize and some proteins may be more difficult to solubilize. For example: Jurkat cell yields from 5 million cells are around 100 µg of membrane protein and 500 µg of cytosolic protein while HeLa cell yields from 5 million cells are around 450 µg of membrane protein and 750 µg of cytosolic protein.

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The Mem-PER Plus Membrane Protein Extraction Kit has been evaluated for use with cultured mammalian cells and tissues (both hard and soft). Cell lines tested include HeLa, HEK293, A431, PC3, LNCaP, A549, HEPG2, and Jurkat cells.

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We offer the following kits for cell fractionation:

Mem-PER Plus Membrane Protein Extraction Kit, Cat. No. 89842
NE-PER Nuclear and Cytoplasmic Extraction Reagents, Cat. No. 78835
Subcellular Protein Fractionation Kit for Cultured Cells, Cat. No. 78840
Subcellular Protein Fractionation Kit for Tissues, Cat. No. 87790
Mitochondrial Isolation Kit for Cultured Cells, Cat. No. 89874
Mitochondrial Isolation kit for Tissue, Cat. No. 89801
Lysosome Enrichment Kit for Tissues and Cultured Cells, Cat. No. 89839

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Isolation of subcellular fractions and concentration of low-abundantce proteins allows for more efficient identification and study of the proteins of interest.

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Certain detergents can be used to selectively extract and isolate membrane (hydrophobic) proteins from cytosolic (hydrophilic) proteins. The Mem-PER Plus Membrane Protein Extraction Kit (Cat. No. 89842) enriches for integral membrane proteins and membrane-associated proteins from cultured mammalian cells or tissue via selective solubilization using a mild detergent-based protocol. The use of selective detergent extraction eliminates the hassle of phase separation based on hydrophobicity, allowing better reproducibility and higher throughput. The cells are first permeabilized with a mild detergent, allowing the release of soluble cytosolic proteins, after which a second detergent solubilizes membrane proteins.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.