|Size:||80mg, enough for 100ml of samples|
|Description:||Phosphatase Inhibitor Cocktail II inhibits Tyrosine protein phosphatases, acid and alkaline phosphatases. It is used to protect samples against endogenous phosphatase activies so that downstream assays can accurately study the phosphorylation profile of the sample.|
|Application:||WB sample preparation, phosphorylation studies|
Boster's phosphatase inhibitor cocktails provide a broad spectrum inhibition of phosphatase activities, and protect the sample's phosphorylation state against endogenous enzymes. Phosphorylation is one of the most commonly studied post-translational modificaiton. It is a reversible reaction, where kinases catalyze the tansfer of the gamma-phosphate from ATP to the hydroxyl side chain of serine (S), threonine (T) or tyrosine (Y). Phosphorylation is a key regulator to protein activities and is one of the main research focuses for cell signaling studies. More than 30% of proteins present in mamaliam cells are phosphorylated. Studying the protein-protein interations of these proteins, and their involvement in signal transduction require phosphatase inhibitors to "freeze" the phosphorylation states of these proteins, before downstream applications can be applied.
What Is A Phosphotase?
There are four most common types of phosphatases, categorized by their substrate specificity. Phosphatase Inhibitor Cocktail II specifically inhibits Tyrosine protein phosphatases; acid and alkaline phosphatases activities.
- 1. Alkaline phosphatases are non-specific phosphatases. They can remove phospho groups from proteins, nucleotides and alkaloids. Common inhibitors of alkaline phosphatases include the analogs of homoarginine, levamisole and imidazole.
- 2. Protein Serine/Threonine phosphatases are responsible for phosphorylation of Serine and Threonine in mammalian cells. Common inhibitors of protein Serine/Threonine phosphatases include okadaic acid, calyculin A, microcystin-LR, tautomycin, fostriecin and cantharidin. These molecules are all naturally occurring in a variety of marine and soil microbes and the each can inhibit a specific sub selection of Protein Serine/Threonine phosphatases.
- 3. Protein Tyrosine phosphatase remove phosphate groups from phosphorylated tyrosine residues on proteins. These enzymes are key regulatory components in signal transduction pathways. Common inhibitors of protein Tyrosine phosphatases include sodium fluoride, orthovanadate and related compounds.
- 4. Dual specificity (Tyr and Ser/Thr) phosphatases dephosphorylate both serine and threonine residues. They are involved in the regulation of key cell signaling pathways. Common inhibitors of Dual specificity (Tyr and Ser/Thr) phosphatases include vanadate.
|Product Name||Phosphatase Inhibitor Cocktail II|
|Synonyms||Cell Extraction and Lysis Phosphatase Inhibitor cocktail|
|Cite This Product||Phosphatase Inhibitor Cocktail II (Boster Biological Technology, Pleasanton CA, USA, Catalog # AR1120)|
|Description||Phosphatase Inhibitor Cocktail II is a Western blot related that is to be added to cell lysis buffer to protect native phosphoproteins from dephosphorylation during proteins purification and sample preparation used in WB, Co-IP, ChIP, and protein kinase assays.|
|Application||WB sample preparation, protein purification, Co-IP, protein kinase activity assay experiments|
|Pack Size||80mg, enough for 100ml of samples||Reagent Type||Western Blotting related reagent; Inhibitors||Usage||To preserve phosphorylation state and protein functionality following cell lysis||Content||Proprietary mix of: Sodium fluoride, Sodium orthavanadium, Imidazole, Sodium molybdate, Sodium sulphate, Sodium pyrophosphate, B-phosphoric acid glycerol||Target Specificity||Tyrosine protein phosphatases, acid and alkaline phosphatases||Target Sample||Cell lysis extracts|
Technical Details/Feature Benefits
Boster's Phosphatase Inhibitor Cocktail II feature the following benefits:
- Dialyzable: inhibitors can be removed by dialysis, before performing assays known to be incompatible with phosphatase inhibitors.
- 100X: effective as 1X. For samples with high level of phosphatase may need 2-3X.
- Detergent compatible:can be used with lysis buffers and do not crossreact with detergents in the buffers.
- Protein Assay Compatible:works with BCA and commassie blue assays.
- Multiple sizes/Bulk pricing: available. Please contact email@example.com for bulk pricing.
- How to use: reconstitute in 1ml of water, add to sample in ratio of 1:100.
|Tyr and alkaline phosphatases|
|Sodium molybdate||Acid and phosphoprotein phosphatases|
|Sodium tartrate||Acid phosphatase|
|Physical State||White lyophilized powders|
|Recommended working concentration||Reconstitute Phosphatase Inhibitor Cocktail II with 1ml of water.|
|Add Phosphatase Inhibitor Cocktail II solution into tissue or cell extracts in a ratio of 1:100. Optimal dilutions should be determined by end users.|
|Compatibility with reagents||Fully compatible with cell lysis buffers and Broad Spectrum Protease Inhibitor Cocktail|
|Compatibility with assays||Compatible with IEF/2D studies;|
|MS-compatible: not contain AEBSF;|
|Compatible with immobilized metal chelate affinity chromatography and 2D gel electrophoresis inhibitor components dialysis removal or desalting;|
|Storage||At -20˚C for 6 months.|
|Precautions||FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC AND CLINICAL USE|
Usage and Handling
The product is supplied as white lyophilized powders. Since phosphatase levels may vary among cell and tissue types, it may be necessary to increase the concentration of inhibitors.
Phosphatase inhibitors are used when phosphorylation (activation) states of target proteins need to be studied and the phosphorylated residues of interest must remain intact. They are chemicals that aid in the extraction of intact proteins in their native modification state by inhibiting endogenous phosphatases that would otherwise dephosphorylate the proteins present in cell lysates and tissue extracts.
Phosphatase Inhibitor Cocktail II contains individual components with specific inhibitory properties to provide an all-around protection of the protein phosphorylation state.
Dynamic protein phosphorylation is a key cellular signaling mechanism for cell processes regulation. When tissues are lysed to make whole cell extracts, the loss of natural compartmentalization causes normal regulation of cellular signaling to get distorted, and resident cell phosphatases within the cell extract are free to disorderly dephosphorylate proteins. The usual consequence of this unregulated state is biologically meaningless representation of protein activities (i.e. phosphorylation status) and false negative staining in anti-phosphoprotein immunostaining analyses. The addition of phosphatase inhibitors to the cell lysis buffer aids in the preservation of phosphorylated residues at the time of cell disruption.