Description
Isolation of microsomal membranes from plant tissues is a common laboratory procedure.
Microsomal fraction of plant cell lysate is the focus of interest in many plant research projects.
Microsomal fraction is believed to be enriched for plasma membranes, endoplasmic reticulum,
Golgi apparatus, vacuolar membranes and other components of membrane system. Traditional
method for microsomal fraction isolation involves so called differential pelleting protocol where a
series of centrifugation steps are required to obtain various membrane fractions. Traditional
protocol for microsomal fraction isolation requires large amount of starting material and employs
tedious ultracentrifugation steps. MM-018 offers a simple, rapid and user friendly approach for
microsomal membrane extraction using small amount of starting material (200 mg). Water
soluble cytosolic proteins are removed during the procedures and water insoluble microsomal
fraction, especially plasma membrane fraction, is extracted with optimized buffers in a table-top
microcentrfuge. The procedure is simple, rapid and no special instrument required. Native
microsomal proteins can be isolated from plant tissue in about one hour without
ultracentrifugation. The protein yield is in the range of 100-200 µg/sample.
Applications
Microsomal membranes extracted with this kit can be used for many downstream applications
such as SDS-PAGE analysis, Western blotting, IP. ELISA, enzyme activity assays, proteomics
and membrane trafficking analysis.
Kit components
1. 25 ml buffer A
2. 10 ml buffer B
3. Filter cartridge with 2.0 ml collection tube (50)
4. Plastic rods (4)
Storage: Store the kit at -20oC.
Additional Materials Required
Table-Top Microcentrifuge with a maximum rpm of 14,000-16,000. 1 X PBS.
Important Product Information Prior to plasma membrane isolation addition of protease inhibitor cocktail to buffer A is recommended. For determination of protein concentration, BCA kit (Pierce) is recommended. To study protein phosphorylation, phosphatase inhibitors (such as PhosStop from Roche) should be added to buffer A prior to use.