iBLSBio LifeSpan BioSciences, Inc. Alkaline Phosphatase (ALP) Assay Kit (Fluorometric) LS-K301-100 (100 Tests) • Store at -20°C Introduction Alkaline phosphatase (ALP) catalyzes the hydrolysis of phosphate esters in an alkaline environment, resulting in the formation of an organic radical and inorganic phosphate. In mammals, this enzyme is found mainly in the liver and bones. Marked increase in serum ALP levels, a disease known as hyperalkalinephosphatasemia, has been associated with malignant biliary obstruction, primary biliary cirrhosis, primary sclerosing cholangitis, hepatic lymphoma and sarcoidosis. Simple, direct and automation-ready procedures for measuring ALP activity in serum are becoming popular in Research and Drug Discovery. This improved Method utilizes 4-methylumbelliferyl phosphate that is hydrolyzed by ALP into a highly fluorescent product 4-methylumbelliferone. The rate of the fluorescence increase is directly proportional to the enzyme activity. ' i 4-Methylumbelliferyl phosphate Alkaline Phosphatase 4-Methylumbelliferone + phosphate fluorescent (360/450 nm) _’_/ Key Features • High sensitivity and wide linear range. Use 10 pL sample. Detection limit of 0.02 U/L (20 min reaction). • Homogeneous and simple procedure. Simple "mix-and-measure" procedure allows reliable quantitation of ALP activity within 20 minutes. • Robust and amenable to HTS. All reagents are compatible with high-throughput liquid handling instruments. Applications • Direct Assays: ALP activity in serum, plasma and other sources. • Characterization and Quality Control for ALP production. • Drug Discovery: high-throughput screen for ALP modulators. Materials Not Supplied Pipetting devices and accessories (e.g. multi-channel pipettor). Black flat-bottom 96-well plates (e.g. Corning Costar) and plate reader. 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]

iBLSBio LifeSpan BioSciences, Inc. Alkaline Phosphatase (ALP) Assay Kit (Fluorometric) LS-K301-100 (100 Tests) • Store at -20°C Storage The kit is shipped at room temperature. Store at -20°C. Shelf life of 2 years after receipt. Assay Procedure Thaw reagents to room temperature prior to use. This assay is based on a kinetic reaction. Use of a multi-channel pipettor is recommended. Addition of Reagent to samples should be quick and mixing should be brief but thorough. Assays can be executed at room temperature or 37°C. Sample preparation ALP is stable for 48 hours at 4°C and 2 months at -20°C....

Alkaline Phosphatase (ALP) Assay Kit (Fluorometric) LS-K301-100 (100 Tests) • Store at -20°C Sample Data Standard Curve (20 min incubation) 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 • TechnicalSupport@LSB