RIPA buffer is one of the most useful protein extraction buffers and maintains most native structures of proteins. Nevertheless, RIPA buffer is not convenient enough to extract membrane proteins and membrane-associated proteins concentrated in lipid raft.
Lipid raft is a highly unique microdomain on the lipid bilayer which contains special lipids, cholesterol and functional proteins. In this layer, lipid raft-enriched proteins are usually insoluble in mild detergent buffers such as 1% Triton X-100 and RIPA buffer. Consequently, it was, up to now, difficult to analyze the functions of this kind of proteins.
Whato do then?
UltraRIPA kit for Lipid Raft provides a solution to extract and analyze membrane proteins or membrane-associated proteins enriched in lipid rafts with native structure and function.
Features of UltraRIPA kit
- Extract membrane / membrane associated proteins enriched in lipid rafts, with native structure and fully retained function
- Easy and simple procedure : only a centrifuge is required
- Only two components: A buffer and B buffer
Extraction of RIPA-insoluble proteins
Left : Quantification of extracted total proteins by BCA protein assay. UltraRIPA kit could constantly extract over 70% of RIPA-insoluble proteins from the mouse brain tissue.
Right : Western blotting of lipid raft markers. Some lipid raft markers among proteins or a ganglioside extracted were dramatically increased in RIPA-insoluble fraction by UltraRIPA kit.
Enzyme activity assay of proteins extracted by UltraRIPA kit
Left : Lactate Dehydrogenase activity of proteins extracted by using 1% Triton X-100, A and B buffer. Equivalent enzyme activity are obtained when using A and B buffer.
Right : Total protein phosphatase activity. Protein extracts from RIPA-insoluble fraction of the mouse whole brain by UltraRIPA kit B-buffer, RIPA, and 2% SDS buffer were applied to total protein phosphatase assay. Proteins extracted by UltraRIPA kit had high phosphatase activity.
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