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Calcineurin cellular activity assay kit

Provides everything needed to assay calcineurin cellular activity, including recombinant calcineurin control
 
BML-AK816-0001 96 wells 482.00 USD
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  • Safe and non-radioactive assay
  • Convenient 1-step detection
  • Excellent sensitivity
A complete colorimetric assay kit for measuring cellular calcineurin (PP2B) phosphatase activity. It employs a convenient 96-well plate format with all reagents necessary for measuring calcineurin (PP2B) phosphatase activity in tissue/cellular extracts PLUS human recombinant calcineurin is included as a positive control. The RII phosphopeptide substrate, supplied with this kit is the most efficient and selective peptide known for calcineurin. The detection of free-phosphate released is based on the classic malachite green assay.
BML-AK816 std curve
BIOMOL GREEN™ Phosphate Standard Curve
BML-AK816 Fig2
Cellular CaN Assay Using Mouse Brain Phosphatase activity from a freshly prepared mouse brain extract. Prior to gel filtration the extract was diluted 1:1 in lysis buffer containing protease inhibitors. After gel filtration and prior to the phosphatase assay, the extract was diluted 1/25 in lysis buffer. Well contents were as in manual. The reaction was incubated for 30 min at 30°C.
BML-AK816 Fig3
CaN Positive Control. Purified calcineurin (BML-SE163-9090) positive control. The CaN was incubated 1 hr at 30°C under various buffer conditions. The results demonstrate that CaN activity is inhibited by the EGTA buffer, but not inhibited by 100nM or 500nM concentrations of okadaic acid
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BML-AK816 std curve BML-AK816 Fig2 BML-AK816 Fig3

Product Specification

Applications:Colorimetric detection
 
Use/Stability:Store all components except the microtiter plate at -70°C for the highest stability. The calcineurin enzyme component SE163-9090 must be handled particularly carefully in order to retain maximal enzymatic activity. Thaw it quickly in a RT water bath or by rubbing between fingers, then immediately store on an ice bath. The remaining unused enzyme should be quickly refrozen by placing at -70°C. To minimize the number of freeze/thaw cycles, aliquot the calcineurin into separate tubes and store at -70°C.
 
Shipping:Shipped on Dry Ice
 
Long Term Storage:-80°C
 
Kit/Set Contains:

Calcineurin enzyme, Calmodulin (human, recombinant), Substrate (RII phosphopeptide), 2x assay buffer, EGTA Buffer, Lysis Buffer, Protease inhibitor cocktail, Biomol Green™ Reagent, Phosphate standard, Okadaic acid, Desalting column and resin, ½-Volume microplate

 

Product Literature References

Pulsed electromagnetic field inhibits RANKL-dependent osteoclastic differentiation in RAW264.7 cells through the Ca2+-calcineurin-NFATc1 signaling pathway: J. Zhang, et al.; Biochem. Biophys. Res. Commun. 482, 289 (2017), Abstract;
CRTC1 nuclear localization in the hippocampus of the pilocarpine-induced status epilepticus model of temporal lobe epilepsy: D. Dubey & B.E. Porter; Neuroscience 320, 57 (2016), Application(s): Measure calcineurin levels in hippocampal lysates, Abstract;
KRAS mutation leads to decreased expression of regulator of calcineurin 2, resulting in tumor proliferation in colorectal cancer: H. Niitsu, et al.; Oncogenesis 5, e253 (2016), Application(s): Calcineurin activity assay, cell lysates, Abstract; Full Text
New strategy of tacrolimus administration in animal model based on tacrolimus-loaded microspheres: J.J. Zamorano-Leon, et al.; Transplant Immunol. 36, 9 (2016), Abstract;
Differential effects of short-term β agonist and growth hormone treatments on expression of myosin heavy chain IIB and associated metabolic genes in sheep muscle: K.M. Hemmings, et al.; Animal 9, 285 (2015), Application(s): Use with skeletal muscle homogenates, Abstract; Full Text
Myocyte enhancer factor (MEF)-2 plays essential roles in T-cell transformation associated with HTLV-1 infection by stabilizing complex between Tax and CREB: P. Jain, et al.; Retrovirology 12, 23 (2015), Application(s): Assay, Abstract; Full Text
Regulation of GAP43/calmodulin complex formation via calcineurin-dependent mechanism in differentiated PC12 cells with altered PMCA isoforms composition: T. Boczek, et al.; Mol. Cell. Biochem. 407, 251 (2015), Application(s): Assay using cell lysate, Abstract; Full Text
The Ca2+-activated cation channel TRPM4 is a negative regulator of angiotensin II-induced cardiac hypertrophy: M. Kecskés, et al.; Basic Res. Cardiol. 110, 43 (2015), Application(s): Assay using mouse heart supernatant, Abstract; Full Text
TRPV6 channel modulates proliferation of insulin secreting INS-1E beta cell line: M. Skrzypski, et al.; Biochim. Biophys. Acta 1853, 3202 (2015), Application(s): Calcineurin activity in rat pancreatic cells, Abstract;
Calcineurin suppresses AMPK-dependent cytoprotective autophagy in cardiomyocytes under oxidative stress: H. He, et al.; Cell Death Dis. 5, e997 (2014), Application(s): Activity measured in rat cardiomyocyte cellular extracts, Abstract; Full Text
Calcineurin upregulates local Ca2+ signaling through ryanodine receptor-1 in airway smooth muscle cells: C. Savoia et al.; Am. J. Physiol. Lung Cell. Mol. Physiol. 307, 781 (2014), Abstract; Full Text
Calcineurin activity is required for cardiac remodelling in pregnancy: E. Chung, et al.; Cardiovasc. Res. 100, 402 (2013), Abstract; Full Text
Calcium phosphatase calcineurin influences tau metabolism: C.M. Karch, et al.; Neurobiol. Aging 34, 374 (2013), Application(s): Calcineurin phosphatase activity in extracted brain and cell lysates, Abstract;
IF1 limits the apoptotic-signalling cascade by preventing mitochondrial remodelling: D. Faccenda, et al.; Cell Death Differ. 20, 686 (2013), Application(s): Calcineurin phosphatase activity in HeLa cells and mouse embryonic fibroblasts, Abstract;
Intracellular two-phase Ca2+ release and apoptosis controlled by TRP-ML1 channel activity in coronary arterial myocytes: M. Xu, et al.; Am. J. Physiol. Cell Physiol. 304, C458 (2013), Abstract; Full Text
The R740S mutation in the V-ATPase a3 subunit increases lysosomal pH, impairs NFATc1 translocation, and decreases in vitro osteoclastogenesis: I. Voronov, et al.; J. Bone Miner. Res. 28, 108 (2013), Abstract;
Leptin-induced cardiomyocyte hypertrophy reveals both calcium-dependent and calcium-independent/RhoA-dependent calcineurin activation and NFAT nuclear translocation: V. Rajapurohitam, et al.; Cell Signal. 24, 2283 (2012), Application(s): Calcineurin phosphatase activity in rat cardiac myocytes, Abstract;
Molecular mechanisms underlying effects of neural stem cells against traumatic axonal injury: E. Wang, et al.; J. Neurotrauma 29, 295 (2012), Application(s): Calcineurin phosphatase activity in neural stem cells, Abstract;
Sustained postexercise increases in AS160 Thr642 and Ser588 phosphorylation in skeletal muscle without sustained increases in kinase phosphorylation: G.G. Schweitzer, et al.; J. Appl. Physiol. (1985) 113, 1852 (2012), Abstract; Full Text
The antimicrobial peptide, LL-37, inhibits in vitro osteoclastogenesis: C. Supanchart, et al.; J. Dent. Res. 91, 1071 (2012), Application(s): Calcineurin phosphatase activity in human osteoclast progenitor cells, Abstract;
Beneficial effects of adenylyl cyclase type 6 (AC6) expression persist using a catalytically inactive AC6 mutant: M.H. Gao, et al.; Mol. Pharmacol. 79, 381 (2011), Application(s): Calcineurin phosphatase activity in adult rat cardiac myocytes, Abstract;
Calcineurin/nuclear factor of activated T cells (NFAT) signaling in cobalt-chromium-molybdenum (CoCrMo) particles-induced tumor necrosis factor-α (TNFα) secretion in MLO-Y4 osteocytes: V. Orhue, et al.; J. Orthop. Res. 29, 1867 (2011), Application(s): Calcineurin phosphatase activity in osteocytes, Abstract;
Ginseng inhibits cardiomyocyte hypertrophy and heart failure via NHE-1 inhibition and attenuation of calcineurin activation: J. Guo, et al.; Circ. Heart Fail. 4, 79 (2011), Application(s): Calcineurin phosphatase activity in rat cardiac myocytes, Abstract;
Regulator of calcineurin 1 (Rcan1) is required for the development of pulmonary eosinophilia in allergic inflammation in mice: Y.J. Yang, et al.; Am. J. Pathol. 179, 1199 (2011), Application(s): Calcineurin phosphatase activity in murine bone marrow cells, Abstract;
A conserved calcineurin-binding motif in human T lymphotropic virus type 1 p12I functions to modulate nuclear factor of activated T cell activation: S.J. Kim, et al.; J. Biol. Chem. 278, 15550 (2003), Application(s): Jurkat T cells, Abstract; Full Text
Targeted inhibition of calcineurin prevents agonist-induced cardiomyocyte hypertrophy: T. Taigen, et al.; PNAS 97, 1196 (2000), Application(s): Rat cardiomyocytes, Abstract; Full Text

General Literature References

Overexpression and purification of human calcineurin alpha from Escherichia coli and assessment of catalytic functions of residues surrounding the binuclear metal center: A. Mondragon et al.; Biochemistry 36, 4934 (1997), Abstract;
Characterization and kinetic analysis of the intracellular domain of human protein tyrosine phosphatase beta (HPTP beta) using synthetic phosphopeptides: K. W. Harder et al.; Biochem. J. 298 , 395 (1994), Abstract;
Dephosphorylation of phosphopeptides by calcineurin (protein phosphatase 2B): A. Donella-Deana et al.; Eur. J. Biochem. 219, 109 (1994), Abstract;
Nonradioactive assay for protein phosphatase 2B (calcineurin) activity using a partial sequence of the subunit of cAMP-dependent protein kinase as substrate: A. Enz et al.; Anal. Biochem. 216, 147 (1994), Abstract;
Use of fluorinated tyrosine phosphates to probe the substrate specificity of the low molecular weight phosphatase activity of calcineurin: B. Martin et al.; J. Biol. Chem. 260, 14932 (1985), Abstract;

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