Urease Assay Kit, BioAssay(TM)

Urease (Amidohydrolase, EC 3.5.1.5) is an enzyme that catalyzes the hydrolysis of urea into carbon dioxide and ammonia.\n(NH2)2CO+H2O ? CO2+2NH3\nMany gastrointestinal or urinary tract pathogens produce urease. Thus its activity is a useful diagnostic parameter for the presence of pathogens such as Helicobacter pylori. Urease is found in bacteria, yeast, and higher plants. Urease activity is commonly determined in anaerobes of the bovine rumen, human feces and environmental samples such as soils and phytoplanktons.\nUrease Assay Kit provides a very sensitive and convenient means to measure urease activity in a variety of samples including soil. In the assay, urease reacts with urea, resulting in the formation of ammonia, which is determined by the Berthelot method at 670nm. The assay is simple, sensitive, stable and high-throughput adaptable.\n\nKey Features:\nSafe. Non-radioactive assay.\nSensitive and accurate. As low as 0.003 U/L urease activity can be quantified.\nHomogeneous and convenient. "Mix-incubate-measure" type assay. No wash and reagent transfer steps are involved.\nRobust and amenable to HTS: can be readily automated on HTS liquid handling systems for processing thousands of samples per day.\n\nApplications:\nUrease activity determination in biological and environmental samples.\nEvaluation and screening for urease inhibitors.\n\nKit Contents:\nAssay Buffer: 20ml (pH 7.0) Reagent A: 12ml\nUrea: 1.5ml Reagent B: 6ml\nNH4Cl: 100ul 50mM\nStorage conditions: store all reagent at 4 degrees C. This product is shipped at room temperature. Shelf life of 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\nAssay Procedure:\nInterference: ammonia is known to interfere with this assay and prior to assay, should be removed by dialysis or filtration.\n1. Assay Preparation. Prior to assay, bring all components to room temperature. For calibration curve, prepare a 500uM premix by mixing 5ul 50mM NH4Cl and 495ul Buffer. Dilute NH4Cl as follows\nNo Premix+Buffer Vol (ul) NH4\n+ (uM)\n1 100ul+0ul 100 500\n2 80ul+20ul 100 400\n3 60ul+40ul 100 300\n4 40ul+60ul 100 200\n5 30ul+70ul 100 150\n6 20ul+80ul 100 100\n7 10ul+90ul 100 50\n8 0ul+100ul 100 0\nTransfer 90ul into separate wells of a clear flat-bottom 96-well plate. Samples. Dilute sample in Assay Buffer (Note: it is prudent to test different dilutions to ensure urease activity is within the detection range). Transfer 90ul sample into separate wells. Use 90ul enzyme buffer as a Sample Blank.\n2. Enzyme Reaction. Add 10ul Urea to each well. Incubate at desired temperature for 10 min.\n3. Detection. Add 100ul Reagent A to each well. Tap plate to mix. Then add 50ul Reagent B to each well. Tap plate to mix again. Note: addition of Reagent A terminates the urease reaction. Incubate for 30 min in the dark. Read optical intensity at 670nm (630-700nm).\n\nCalculation:\nPlot NH4Cl calibration curve and determine its Slope (uM-1). Urease enzyme activity in the sample is calculated as\nUrease Activity =\nODSAMPLE

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SPECIFICATIONS

Catalog Number

U2015-02

Size

1Kit

References

1. Kandeler, E., Gerber, H., 1988. Short-term assay of soil urease activity using colorimetric determination of ammonium. Biology and Fertility of Soils 16: 249-254.\n2. Bremner, J.M., Mulvaney, R.L., 1978. Urease activity in soils. In: Burns, R.G. (Ed.). Soil Enzymes, Academic Press, New York, pp. 149- 197.\n3. Sinsabaugh, R.L. et al (2000). Rapid assay for amidohydrolase (urease) activity in environmental samples. Soil Biol & Biochem. 32: 2095-97