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Hydrogen Peroxide Colorimetric Detection Kit


ICT’s Hydrogen Peroxide Colorimetric Detection Kit allows you to quantitatively measure H2O2 in a variety of samples. This kit is validated for use in fresh urine, buffers, and tissue culture media. This kit is species independent.

Catalog # Size Quantity Price
9132 2 96-well plates $399.00
Catalog #: 9132 Category:

In biological systems, the one electron (incomplete) reduction of molecular O2 during respiration produces the highly reactive superoxide anion radical (O2-), which is known to be the source of most reactive oxygen species (ROS). Further reduction of the reactive O2– radical by the enzyme superoxide dismutase (SOD) results in the creation of H2O2. This is a relatively stable, (non-radical-electron-state) oxygen form which unlike its more oxidized precursor, is capable of readily penetrating cell membrane barriers and influencing intracellular metabolic activities. Many cells produce low levels of O2– and H2O2 in response to a variety of extracellular stimuli, such as cytokines or peptide growth factors. The addition of exogenous H2O2 or the intracellular production in response to receptor stimulation affects the function of various proteins, including protein kinases, protein phosphatases, transcription factors, phospholipases, ion channels, and G proteins. H2O2 and O2 may participate in the production of singlet oxygen and peroxynitrite. Generation of these species may be concurrent with reactions involving iron, and under some circumstances, they might be important contributors to H2O2 toxicity.

ICT’s Hydrogen Peroxide (H2O2) Colorimetric Detection Kit is designed to quantitatively measure H2O2 in a variety of samples. A hydrogen peroxide standard is provided to generate a standard curve. Samples are mixed with the Colorimetric H2O2 Detection Substrate and the reaction is initiated by addition of horseradish peroxidase (HRP). HRP reacts with the substrate in the presence of hydrogen peroxide to convert the colorless substrate into a pink colored product, which is read at 560 nm. Increasing concentration levels of H2O2 cause a corresponding linear increase in color.

1. Prepare Assay Buffer; dilute Assay Buffer Concentrate 1:5 in diH2O.
2. Prepare samples; dilute > 1:10 in Assay Buffer.
3. Prepare standard curve and zero (blank) standard.
4. Add 50 µL blanks, standards, and samples to plate.
5. Add 25 µL Colorimetric H2O2 Detection Substrate to plate.
6. Prepare HRP, dilute Horseradish Peroxidase Concentrate 1:50 in Assay Buffer.
7. Add 25 µL HRP Preparation to plate.
8. Incubate 15 minutes at RT.
9. Read absorbance of the plate at 560 nm.

Typical Standard Curve
9132 Standard Curve

• Target: Hydrogen Peroxide
• Method of Analysis: Absorbance Plate Reader
• Types of Samples: fresh urine, buffers, and tissue culture media
• Storage: 2-8°C
• Shipping: Ships overnight (domestic), International Priority Shipping

• 2 Clear Half-Area 96-well Microwell Plates, #267
• Hydrogen Peroxide Standard (Hydrogen Peroxide at 1,000 µM in a special stabilizing solution), 200 µL, #6604
• 5X Assay Buffer Concentrate (a buffer concentrate containing detergents and stabilizers), 25 mL, #6605
• Colorimetric H2O2 Detection Substrate (a solution of the substrate in a special stabilizing buffer), 5 mL, #6608
• 50X Horseradish Peroxidase Concentrate (a concentrated solution of HRP in a special stabilizing solution), 120 µL, #6609
• Kit Manual