Use the ReadyPrep Protein Extraction Kit (Membrane I) for simple, rapid and reproducible preparation of cellular protein fractions highly enriched in membrane and hydrophobic proteins. Protein isolated using this kit can be used for many applications including 2-D gel analysis, SDS-PAGE, and western blotting.

• Isolates proteins containing 1–2 transmembrane domains, membrane-associated proteins, and hydrophobic proteins
• Protocol can be used with a wide variety of biological samples
• Does not require the use of ultracentrifugation or density gradients
• Contains a strongly chaotropic extraction buffer to efficiently solubilize proteins
• Includes the components of the ReadyPrep 2-D Cleanup Kit to remove constituents of the extraction buffer that can interfere with IEF

The ReadyPrep Protein Extraction Kit (Membrane I) separates membrane proteins by temperature-dependent phase partitioning using Triton X-114 detergent (Bordier 1981, Santoni et al. 2000). Proteins anchored to the membrane or proteins containing one or two transmembrane domains are efficiently partitioned to the detergent-rich phase. Cytoplasmic proteins partition into the aqueous phase. An insoluble pellet contains more complex membrane proteins. Proteins extracted into the two phases are treated using the included ReadyPrep 2-D Cleanup Kit to remove kit components that interfere with IEF. Pellets are then efficiently solubilized using the strongly chaotropic extraction buffer. The solubilized proteins can then be immediately used in 2-D gel electrophoresis or other applications.

Kit Components

For 50 extractions of 25–50 mg of cells or tissue:

• Membrane protein extraction buffer 1 (M1), 50 ml
• Membrane protein extraction buffer 2 (M2), 50 ml
• Protein solubilization buffer (PSB), 25 g
• PSB diluent, 30 ml
• ReadyPrep 2-D Cleanup Kit (1632130)
• Instruction manual

Bio-Rad offers a range of ReadyPrep Kits for Fractionation by Cellular Location. These kits reduce sample complexity, increase detection and identification of low-abundance proteins, and simplify proteomic studies.

Related Products

• ReadyPrep Kits for Fractionation by Solubility
• Aurum™ Ion Exchange Mini Kits for Fractionation by Charge
• Rotofor^® Liquid-Phase IEF Cells for Fractionation by pI
• Model 491 Prep Cell and Mini Prep Cell for Fractionation by Solubility
• ReadyPrep™ 2-D Starter Kit
• PROTEAN^® i12™ IEF System
• ReadyStrip™ IPG Strips
• ReadyPrep Reduction-Alkylation Kit (1632090)
• PowerPac™ HV High-Voltage Power Supply

Related Categories

• Protein Gels
• Electrophoresis Chambers
• 2-D Electrophoresis Buffers and Reagents
• Ampholytes
• Tube Gel IEF Systems
• Gel Imaging Systems
• 2-D Electrophoresis Workflow

Supporting Documentation

• ReadyPrep Protein Extraction Kit (Membrane I) Instruction Manual
• Sample Preparation: Tools for Protein Sample Extraction, Cleanup, Fractionation, and Depletion Brochure
• ReadyStrip IPG Strip Instruction Manual
• PROTEAN i12 IEF System Brochure
• Host Cell Protein (HCP) Workflow Quick Guide

References

Bordier C (1981). Phase separation of integral membrane proteins in Triton X-114. J Biol Chem 256, 1604–1607. PMID: 6257680
Santoni V et al. (2000). Membrane proteomics: Use of additive main effects with multiplicative interaction model to classify plasma membrane proteins according to their solubility and electrophoresis properties. Electrophoresis 21, 3329–3344. PMID: 11079553