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Usage of the new Trinder ’s reagent DAOS
Application: Hydrogen peroxide detection, colorimetric method
Features: As a member of a new Trinder's reagent, Daos is a stable aniline analogue with high water solubility compared with other chromogenic substrates. The pH range of chromogenic process and oxidation reaction is very wide, which is widely used in diagnosis and biochemical test. In the colorimetric determination of the activity of hydrogen peroxide, it has some advantages over conventional reagents. The new Trinder's reagent is stable enough to be used in both solution and test line detection system.
Colorimetric determination of hydrogen peroxide
Testing principle: In the presence of hydrogen peroxide and peroxidase, the new Trinder's reagent forms a very stable purple or blue dye in the process of oxidative coupling reaction with 4-aminoantipyrine (4-AA) or 3-methylbenzothiazolylsulfonhydrazone (MBTH). The molar absorbance of the dye coupled with MBTH is 1.5-2 times higher than that of the dye coupled with 4-AA. The substrate is oxidised by its oxidase to produce hydrogen peroxide. The concentration of hydrogen peroxide corresponds to the substrate concentration. Therefore, the amount of substrate can be determined by the development of oxidative coupling reaction. For a specific substrate, it is necessary to test different kinds of new Trinder's reagent to develop the best detection system.
Preparation of detection solution-DAOS (Configuration methods of other substrate are basically the same):
1. Dissolve 23mg of DAOS in 10 ml of PBS buffer to prepare a 6.6 mM DAOS solution. (The specific solubility is adjusted as needed)
2. Dissolve 14 mg 4-aminoantipyrine (4-AA) in 10 ml PBS buffer to prepare a 6.6 mM 4-AA solution.
3. Prepare 2U/ ml horseradish peroxidase solution in PBS.
4. Mix equal volumes of each solution to prepare a test solution. Store the test solution in the dark at 4 ℃.
1. Prepare a sample solution for the enzymatic oxidation reaction. The pH value of the buffer solution should be 5.5-9.5.
2. Use the same buffer to prepare a standard solution containing a known amount of substrate.
3. Add appropriate units of oxidase to the sample solution, followed by an equal volume of detection solution.
4. Incubate the mixture at room temperature or 37 ° C for 30 minutes to 1 hour.
5. Determine the O.D. value at 593 nm.
6. Prepare a standard curve and determine the substrate concentration in the sample solution.
Desheng Biochemical's research on new Trinder ’s reagents is very effective. The produced DAOS, TOOS, TOPS, MAOS, etc. have high purity, good experimental stability, and are widely recognized by the industry. Welcome to call us for consultation. Tel: +86 15071057538!