Creatine Kinase Assay Kit, BioAssay(TM)

Creatine Kinase (CK), also known as creatine phosphokinase (CPK), is an enzyme (EC 2.7.3.2) expressed predominantly in skeletal muscle, smooth muscle and the brain. The CK enzyme consists of two subunits, which can be either B (brain type) or M (muscle type), and hence three different isoenzymes: CK-MM, CK-BB and CK-MB. CK catalyzes the conversion of creatine to phosphocreatine, consuming adenosine triphosphate (ATP) and generating adenosine diphosphate (ADP) and the reverse reaction. CK is often determined routinely in emergency patients with chest pain and acute renal failure. Elevation of CK is an indication of damage to muscle and has been associated with injury, rhabdomyolysis, myocardial infarction, myositis, myocarditis, malignant hyperthermia and neuroleptic malignant syndrome, etc. Lower levels can be an indication of alcoholic liver disease and rheumatoid arthritis. \n\nSimple, direct and automation-ready procedures for measuring CK activity are very desirable. Creatine Kinase Assay Kit is based on enzyme coupled reactions in which creatine phosphate and ADP is converted to creatine and ATP by CK, the generated ATP is used to phosphorylate glucose by hexokinase to generate glucose-6-phosphate, which is then oxidized by NADP in the presence of glucose-6-phosphate dehydrogenase. The produced NADPH, measured at 340 nm, is proportionate to the CK activity in the sample.\n\nApplications:\nDirect Assays: CK in serum, plasma and other biological samples.\nPharmacology: effects of drugs on CK activity.\n\nKey Features:\nSensitive and accurate. Detection range: 5 to 300 U/L creatine kinase in 96-well plate assay.\nConvenient. The procedure involves adding a single working reagent, and reading the optical density at 20 min and 40 min at room temperature or 37 degrees C.\nHigh-throughput. Can be readily automated as a high-throughput 96-well plate assay for thousands of samples per day.\n\nKit Contents: (100 tests in 96-well plates)\nAssay Buffer: 12ml Substrate Solution: 1.0ml\nEnzyme Mix: 120ul Calibrator: 150ul\nStorage conditions. This kit is shipped on dry ice. Store all reagents at -20 degrees C. Shelf life: 6 months after receipt.\nPrecautions: reagents are for research use only. Normal precautions for laboratory reagents should be exercised while using the reagents. Please refer to Material Safety Data Sheet for detailed information.\n\nProcedures:\nSample Preparation. Use non-hemolyzed samples. Samples should be assayed within 4 hours of blood collection if they remain at room temperature, or within 12 hours if stored at 4 degrees C. Samples can be stored at -80 degrees C and should be thawed only once. If turbidity is observed, centrifuge sample and use clear supernatant for assay. Reagent Reconstitution. Bring all components to room temperature. For each reaction well, mix 10ul Substrate Solution, 100ul Assay Buffer and 1ul Enzyme Mix. Fresh reconstitution is recommended. If the assay is to be carried out at 37 degrees C, warm up the Reconstituted Reagent at 37 degrees C.\n1. Calibrator: transfer 110ul water and (10ul Calibrator+100ul water) into separate wells of a clear bottom 96-well plate.\nSamples: transfer 10ul samples into separate wells. Add 100ul Reconstituted Reagent and tap plate to mix.\n2. Reaction. Incubate at room temperature or 37 degrees C. CK is fully activated within 20 min by glutathione provided in the Substrate Solution. Read OD340nm at 20 min and again at 40 min.\n3. Calculation:. Calculate sample CK activity using the equation,\nCK (U/L) =\nOD40min

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SPECIFICATIONS

Catalog Number

C7909-97

Size

1Kit

References

1. Itaka K et al (2010). Polyplex nanomicelle promotes hydrodynamic gene introduction to skeletal muscle. J Control Release. 143(1): 112-9.\n2. Steen C et al (2010). Reduced Creatine Kinase B Activity in Multiple Sclerosis Normal Appearing White Matter. PLoS One 5(5): e10811.\n3. Kwon S et al. (2010). ASB9 interacts with ubiquitous mitochondrial creatine kinase and inhibits mitochondrial function. BMC Biol. 8: 23.