Glycerol Assay Kit, BioAssay(TM)

Glycerol [Glycerin or Glycerine, C3H5(OH)3] is widely used in foods, beverages and pharmaceutical formulations. It is also a main by- product of biodiesel production. Simple, direct and automation-ready procedures for measuring glycerol concentrations find wide Applications:. Glycerol assay uses a single Working Reagent that combines glycerol kinase, glycerol phosphate oxidase and color reactions in one step. The color intensity of the reaction product at 570nm or fluorescence intensity at lem/ex=585/530nm is directly proportional to glycerol concentration in the sample.\n\nKey Features:\nSensitive and accurate. Use as little as 10ul samples. Linear detection range in 96-well plate: 10 to 1000uM (92 ug/dL to 9.2 mg/dL) glycerol for Colorimetric Assay:s and 2 to 50uM for Fluorimetric Assay:s.\nSimple and convenient. The procedure involves addition of a single working reagent and incubation for 20 min at room temperature, compatible for HTS assays.\nImproved reagent stability. The optimized formulation has greatly enhanced the reagent and signal stability.\n\nApplications:\nDirect Assays: glycerol in biological samples (e.g. serum and plasma).\nDrug Discovery/Pharmacology: effects of drugs on glycerol metabolism.\nFood and Beverages: glycerol in food, beverages, pharmaceutical formulations etc.\n\nKit Contents:\nAssay Buffer: 24ml Enzyme Mix: 500ul ATP: 250ul\nDye Reagent: 220ul Standard: 100ul 100mM Glycerol\nStorage conditions. The kit is shipped on dry ice. Store Assay Buffer at 4 degrees C and other reagents at -20 degrees C. 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\nColorimetric Procedure:\nNote: SH-group containing reagents (e.g. mercaptoethanol, DTT) may interfere with this assay and should be avoided in sample preparation.\n1. Equilibrate all components to room temperature. Keep thawed Enzyme Mix in a refrigerator or on ice. Dilute standard in distilled water as follows (diluted standards can be used for future assays when stored refrigerated).\nNo STD+H2O Vol (ul) Glycerol (mM)\n1 10ul+990ul 1000 1.0\n2 6ul+994ul 1000 0.6\n3 3ul+997ul 1000 0.3\n4 0ul+1000ul 1000 0\nTransfer 10ul standards and 10ul samples into separate wells of a clear 96-well plate.\n2. For each reaction well, mix 100ul Assay Buffer, 2ul Enzyme Mix, 1ul ATP and 1ul Dye Reagent in a clean tube. This Working Reagent should be used on the same day of preparation. Transfer 100ul Working Reagent into each reaction well. Tap plate to mix.\n3. Incubate 20 min at room temperature. Read optical density at 570nm (550-585nm). Note: if the Sample OD is higher than the Standard OD at 1.0mM, dilute sample in water and repeat the assay. Multiply result by the dilution factor.\n\nCalculation:\nSubtract blank OD (water, #4) from the standard OD values and plot the OD against standard concentrations. Determine the slope using linear regression fitting. The glycerol concentration of Sample is calculated as\n[Glycerol]=ODSAMPLE

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SPECIFICATIONS

Catalog Number

G8145-10

Size

1Kit

References

1. Duncan RE, et al. (2007). Regulation of lipolysis in adipocytes. Annu Rev Nutr. 27: 79-101.\n2. Moller F, Roomi MW. (1974). An enzymatic, spectrophotometric glycerol assay with increased basic sensitivity. Anal Biochem. 59(1): 248-58.\n3. MacRae AR. (1977). A semi-automated enzymatic assay for free glycerol and triglycerides in serum or plasma. Clin Biochem. 10(1): 16-9.