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NEW! InnoZyme™ Calpain1/2 Activity Assay Kit (Cat. No. CBA054)
 | | | Calpains, calcium ion-dependent papain -like cysteine proteases, are widely distributed in all mammalian cells. Calpain 1 and 2 are heterodimeric proteins comprised of a large 78-80 kDa catalytic subunit encoded by capn 1 and 2, and the smaller 28-30 kDa regulatory subunit, common for both enzymes, encoded by the capn4 gene. In vitro, calpain 1 ( m-calpain) and 2 (m-calpain) are activated by micromolar (10-50 mM) or millimolar (0.2-0.35 mM) Ca2+, respectively. There is evidence to suggest that calpains are tightly regulated post-transcriptionally by calpastatin (the endogenous calpain inhibitor), calcium requirements, and auto-proteolytic cleavage.
Calpains are believed to participate in intracellular signal processing via limited proteolysis of their targets. As irreversible modifiers of a variety of proteins, calpains are likely to be involved in the regulation of many life processes. They may play a central role in the execution of apoptosis either upstream or downstream of caspases in glucocorticoid-treated and irradiated thymocytes, neuronal cells exposed to various stresses, or MCF-7 breast cancer cells treated with b-lapachone. Calpains also mediate calcium-induced activation of the ERK1/ERK2 MAPK pathway resulting in aberrant phosphorylation of the neuronal cytoskeleton, an early event in Alzheimer’s disease.
The Calbiochem® InnoZyme™ Calpain 1 & 2 Activity Kit is a sensitive, homogeneous assay optimized for measuring active calpain 1 and 2. The Substrate used in the assay (DABCYL)-TPLK|SPPPSPR-(EDANS) is based on the finding that the amino acid preferences for calpain recognition/cleavage extend to 11 positions around the scissile amide bond. Calpain-containing samples are incubated with the Substrate in the presence of calcium ions and Reducing Agent. Cleavage of the scissile amide bond, K-S, releases the fluorophore (EDANS) from the internal quenching molecule (DABCYL), resulting in an increase in fluorescence. Fluorescence is measured at an excitation wavelength of 320 nm and an emission wavelength of 480 ± 20 nm. All biological samples are assayed in parallel with the calpain inhibitor, Z-LLY-FMK (included). | | | The InnoZyme™ Calpain1/2 Activity Assay Kit is: | • | Sensitive: detects calpain activity at 63-1000 ng/ml | • | Specific: Substrate and inhibitor optimized for specific detection of calpain activity | • | Versatile: Homogeneous 96-well format can be used with biological samples or for inhibitor screens | • | Convenient: Contains all reagents to assay samples in 30 minutes |
| | | | Rollover a description below to view appropriate figure. | | | |
| | | | | Related Products  | | Calpain Products | | Calcium Signaling/Calcium Metabolism Products | | | | | |
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