Mem-PER Eukaryotic Membrane Protein Extraction Kit 

The Thermo Scientific Mem-PER Eukaryotic Membrane Protein Extraction Kit enables small-scale solubilization and enrichment of hydrophobic membrane-associated proteins in a simple reagent-based procedure. Traditional methods for isolation of membrane proteins are tedious, time-consuming and typically require gradient separations with the aid of expensive ultracentrifugation equipment. The Mem-PER Kit effectively isolates membrane proteins from cultured cells and tissues using a simple benchtop microcentrifuge procedure. Hydrophobic proteins having three or fewer membrane-spanning domains are effectively solubilized and isolated from hydrophilic （cytoplasmic） proteins using the three-reagent system. The kit makes possible the extraction and selective enrichment of integral and attached membrane proteins from cultured mammalian and yeast cells, as well as from hard and soft mammalian tissues.

• Extraction and isolation – produces minimal cross-contamination （typically less than 10%） of hydrophilic protein into the hydrophobic （membrane） protein fraction
• Cells or tissues – effect for extraction from cultured mammalian and yeast cells and mammalian tissues
• Downstream compatibility – analyze membrane protein extracts by SDS-PAGE, Western blotting, and protein assays
• Fast and simple – isolate membrane proteins in approximay one hour
• No special equipment required – only a benchtop microcentrifuge, tubes and pipettors are needed

Protocol summary for the Mem-PER Membrane Protein Extraction Kit. Preparation of mammalian membrane protein extracts using Mem-PER Mammalian Membrane Protein Extraction Kit can be done quickly and efficiently. First, cells are lysed with an initial detergent. A second detergent is then added to solubilize the membrane proteins. After a quick centrifugation, the cocktail is incubated at 37°C to separate the hydrophobic proteins from the hydrophilic proteins through phase partitioning.

	Yeast membrane protein solubilization and isolation. The Thermo Scientific Mem-PER Kit was used to extract membrane proteins from Saccharomyces cerevisiae strain EGY-194. Wet cell paste （10 to 15mg） was vortexed for 10 minutes at room temperature in 80μL of Mem-PER Reagent A and 100 to 150mg of acid-washed glass beads （405-600μm dia.） to disrupt the yeast cell wall. After pelleting the beads by centrifugation, the cell lysate was transferred to a new tube and kept on ice. The beads were washed with 720μL of Mem-PER Reagents B and C, and the wash was then combined with the cell lysate. Finally, the solution was incubated on ice for 30 minutes. Samples were resolved on 4-12% Bis-Tris gels and examined by Western blot using target-specific antibodies and chemiluminescent substrate. Abbreviations: mitochondrial porin （MP）, 3-phosphoglycerate kinase （PGK）, alkaline phosphatase （AP）, Dol-O-Man synthase （Dpm1p）, solubilized membrane protein （M） and hydrophilic protein fraction （H）.
	Effective extraction and partitioning of membrane and cytoplasmic proteins. Proteins from three cell lines were solubilized and extracted using the Thermo Scientific Mem-PER Kit. Each set of membrane （M） and hydrophilic （H） protein fractions were normalized to one another and analyzed by Western blot for four proteins from the cellular locations noted. The Thermo Scientific Pierce SDS-PAGE Sample Prep Kit （Part No. 89888） was used to remove the detergent from the membrane fraction before electrophoresis analysis of COX4. A negligible amount of protein was found in all debris fractions. AchE is acetylcholinesterase; COX4 is cytochrome oxidase subunit 4; hsp90 is heat shock protein 90.

Membrane protein extraction from rat liver and heart tissue. Hydrophobic （membrane protein, M） fractions and corresponding hydrophilic （H） fractions were prepared from rat liver （A） and rat heart （B） tissue using the Thermo Scientific Mem-PER Kit. Extracts were analyzed by Western blot for VDAC （31 kDa）, cellugyrin （29 kDa） and flotillin-1 （48 kDa） using specific antibodies and chemiluminescent substrate.

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Ordering Information
Product # Description Pkg. Size Instructions MSDS CofA Price 89826 Mem-PER Eukaryotic Membrane Protein Extraction Reagent Kit Formulation: Multi-component kit Sufficient For: 50 to 100 samples each containing 5 million cells, 15mg yeast cell paste or 20mg tissue Kit Contents: Mem-PER Reagent A, 10mL Mem-PER Reagent B, 25mL Mem-PER Reagent C, 40mL   75mL kit Local contact