Catalog No. ABIN1000316
Quick Overview for Invertase Assay Kit (ABIN1000316)
Target
Invertase
Application
Activity Assay (AcA)
Sample Type
Soil
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Specificity
- 0.007 U/L
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Characteristics
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Safe. Non-radioactive assay.
Sensitive and accurate. As low as 0.007 U/L invertase activity can be quantified.
Homogeneous and convenient. Mix-incubate-measure type assay. No wash and reagent transfer steps are involved.
Robust and amenable to HTS: can be readily automated on HTS liquid handling systems for processing thousands of samples per day. -
Components
- 10x Reaction Buffer: 12 mL (pH 4.5). 10x Sucrose: 1.5 mL. Assay Buffer: 10 mL. Enzyme Mix: 120 µL. Glucose Standard: 1 mL. Dye Reagent: 120 µL.
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Material not included
- Pipetting devices, centrifuge tubes, clear or black flat bottom 96-well plate (e.g. Corning Costar).
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Application Notes
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Invertase and sucrase activity determination in biological and environmental (e.g. soil) samples.
Evaluation and screening for invertase inhibitors. -
Protocol
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Interference: thiols (beta-mercaptoethanol, dithioerythritol etc) at > 10 µMinterfere with this assay and should be avoided. Glucose, if present in the sample, should be removed by dialysis or membrane filtration.
1. Assay Preparation. Prior to assay, bring all components to room temperature, briefly centrifuge tubes before opening. Dilute the provided 10x Reaction Buffer and 10x Sucrose to 1-fold by mixing 1 vol of the reagent with 9 vol of dH2O. Use the diluted reagents for all assays. For glucose standard curve, mix 5 µL Glucose Standard with 828 µL dH2O (final 100 µM). Dilute and transfer 40 µL standards to separate wells in a clear flat-bottom 96-well plate. Sample: transfer 40 µL sample to separate wells of the plate. As a sample control, use 40 µL diluted Reaction Buffer.
2. Enzyme Reaction. Add 5 µL of the diluted Sucrose to each well. Tap plate to mix. Incubate 20 min at desired temperature (e.g. 30°C).
3. Glucose Determination. Prepare enough Working Reagent in bulk. For each well, mix 95 µL Assay Buffer, 1 µL Enzyme Mix, 1 µL Dye Reagent. Add 90 µL Working Reagent to each well. Immediately tap plate to mix. Incubate for 20 min in the dark. Read OD570nm. Note: the procedure for fluorimetric assays is the same except that (1) a black flat-bottom 96-well plate is used, (2) glucose standards should be at 20, 12, 6 and 0 µMand that fluorescence intensity at em/ex = 585/530nm is measured. -
Restrictions
- For Research Use only
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Storage
- 4 °C
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- Invertase
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Background
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Quantitative determination of invertase/sucrase activity in biological and environmental (e.g. soil) samples by colorimetric (570nm) or fluorimetric (530/585nm) method.
Procedure: 40 min.
Invertase (beta-fructofuranosidase, EC 3.2.1.26) is an enzyme that catalyzes the hydrolysis of sucrose to fructose and glucose. Invertases cleave at the O-C(fructose) bond, whereas a related enzyme sucrase (EC 3.2.1.48) cleaves at the O-C(glucose) bond. A wide range of microorganisms produce invertase and can, thus, utilize sucrose as a nutrient. Invertase assay finds wide applications in environmental (e.g. soil), agricultural and food (confectionery) industry. This Invertase Assay Kit provides a convenient and ultra-sensitive colorimetric and fluorimetric means to measure invertase activity. In the assay, invertase cleaves sucrose, resulting in the formation of fructose and glucose, which is determined by a colorimetric (570nm) or fluorimetric method (gamma em/ex = 585/530nm). The assay is simple, sensitive, stable and high-throughput adaptable.
Target
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