bSLSBio LifeSpan BioSciences, Inc. Creatine Kinase (CK) Assay Kit (Colorimetric) LS-K311-100 (100 Tests) • Store at -20°C Introduction 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. Simple, direct and automation-ready procedures for measuring CK activity are very desirable. LSBio's 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. Key Features • Sensitive and accurate. Detection range: 5 to 300 U/L creatine kinase in 96-well plate assay. • Convenient. The procedure involves adding a single working reagent, and reading the optical density at 20 min and 40 min at room temperature or 37°C. • High-throughput. Can be readily automated as a high-throughput 96- well plate assay for thousands of samples per day. Applications • Direct Assays: CK in serum, plasma and other biological samples. • Pharmacology: effects of drugs on CK activity. Materials Not Supplied Pipetting (multi-channel) devices. Clear-bottom 96-well plates (e.g. Corning Costar) and plate reader. Storage The kit is shipped on ice. Store all kit components at -20 °C. FOR RESEARCH USE ONLY! Not for use in humans. LifeSpan BioSciences, Inc. • 2401 Fourth Avenue, Suite 900, Seattle, WA 98121 www.LSBio.com • (206) 464-1554 • [email protected]

Creatine Kinase (CK) Assay Kit (Colorimetric) LS-K311-100 (100 Tests) • Store at -20°C Assay Procedure Sample 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°C. Samples can be stored at -80°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 10 μL Substrate Solution, 100 μL Assay Buffer and 1 μL Enzyme Mix. Fresh reconstitution...

bSLSBio LifeSpan BioSciences, Inc. Creatine Kinase (CK) Assay Kit (Colorimetric) LS-K311-100 (100 Tests) • Store at -20°C Sample Data Samples were assayed in duplicate using the 96-well protocol. The CK activity (U/L) was 12.0 ± 0.9 for a rat serum sample, 11.0 ± 0.5 for human serum, 28 ± 1 for human plasma, 9.0 ± 0.8 for mouse serum and 49 ± 2 for bovine serum. FOR RESEARCH USE ONLY! Not for use in humans. LifeSpan BioSciences, Inc. • 2401 Fourth Avenue, Suite 900, Seattle, WA 98121 www.LSBio.com • (206) 464-1554 • [email protected]