What is Amplex Red assay?
The Amplex® Red reagent, in combination with horseradish peroxidase (HRP), has been used to detect H2O2 released from biological samples, including cells, or generated in enzyme-coupled reactions. Furthermore, Amplex® Red reagent can be used as an ultrasensitive assay for peroxidase activity when H2O2 is in excess.
What does Amplex Red measure?
Abstract. Reagents such as Amplex® Red have been developed for detecting hydrogen peroxide (H2O2) and are used to measure the release of H2O2 from biological samples such as mammalian leukocytes undergoing the oxidative burst.
How does Amplex Red work?
Amplex Red reagent is a colorless substrate that reacts with hydrogen peroxide (H2O2) with a 1:1 stoichiometry to produce highly fluorescent resorufin (excitation/emission maxima=570/585 nm) (Figure 1). In the scopoletin assay, HRP catalyzes conversion of the fluorescent scopoletin to a nonfluorescent product.
What is hydrogen peroxide assay?
The Hydrogen Peroxide Assay is for the quantitative determination of hydrogen peroxide in colorless tissue culture media and buffers. For research use only. DESCRIPTION. Hydrogen Peroxide (H202) is a reactive oxygen metabolic by-product that serves as a key regulator for a number of oxidative stress-related states.
Is Amplex Red Cell permeable?
The cells appeared fluorescent against a fluorescent background, but it should be noted that Amplex Red is cell-impermeable and is designed as an extracellular assay.
Is H2O2 a peroxide?
Hydrogen peroxide is a chemical compound with the formula H2O2. In its pure form, it is a very pale blue liquid, slightly more viscous than water.
Which indicator is used in assay of hydrogen peroxide?
H2O2 oxidizes iodide to iodine in the presence of acid and molybdate catalyst. The iodine formed is titrated with thiosulfate solution, incorporating a starch indicator.
How are cell ROS levels measured?
In cultured cells, an increase in cellular levels of reactive oxygen species (ROS) can be detected using multiple techniques including colorimetric assays, immunoblotting, and immunofluorescence.
Why is horseradish peroxidase important?
In addition to biomedical applications, horseradish peroxidase is one of the enzymes with important environmental applications. This enzyme is suitable for the removal of hydroxylated aromatic compounds (HACs) that are considered to be primary pollutants in a wide variety of industrial wastewater.
Which is better Amplex ultrared or Amplex Red?
Consequently, assays using Amplex Red as the substrate result in an increase in fluorescence, not a decrease—an inherently superior method for enzymatic assays. Amplex UltraRed reagent—the second-generation Amplex substrate—exhibits increased resistance to oxidation and works better in lower pH environments.
Which is the second generation Amplex Red reagent?
Amplex® Red reagent technology. Amplex® UltraRed reagent—the second-generation Amplex® substrate—exhibits increased resistance to oxidation and works better in lower pH environments. It is used in assays that have an acid intermediate resulting in a lower reaction pH (i.e., myeloperoxidase using the EnzChek® MPO Activity Assay Kit ).
Which is better Amplex Red or scopoletin based assays?
Other advantages of the Amplex Red reaction over scopoletin-based H 2 O 2 assays include high chemical stability of the Amplex Red reagent and its fluorescent product, resorufin, and the long-wavelength spectra of resorufin.
How is the Amplex Red monoamine oxidase assay performed?
The Amplex Red Assays are best performed with either purified enzymes or extracted H 2 O 2 in a defined buffer system, extracellular solutions or body fluids (media, serum, etc.) that do not exhibit high levels of endogenous protease or oxidase activity and do not contain antioxidants. No results found for your search criteria.