Invitrogen™ Catalase Colorimetric Activity Kit

Description

The Catalase Activity research-use-only kit is a colorimetric activity assay designed for the quantification and detection of catalase activity in serum, plasma, cells, tissues and erythrocyte lysates.

This complete, ready-to-use kit includes clear 96 well plate(s), catalase standard (100 Unit/mL), catalase substrate, and other components to perform the assay. A 96-well microplate reader capable of reading optical density at 560 nm (acceptable range: 540-580 nm) is required for use of this kit.

Performance characteristics
• Assay type: colorimetric activity kit
• Sample types: serum, plasma, cells, tissues, and erythrocyte lysates.
• Sensitivity: 0.052 U/mL
• Standard curve range: 0.15–5.0 U/mL
• Reactivity: species independent

Background
Hydrogen peroxide is one of the most frequently occurring reactive oxygen species. It is formed either in the environment or as a by-product of aerobic metabolism, superoxide formation and dismutation, or as a product of oxidase activity. Both excessive hydrogen peroxide and its decomposition product hydroxyl radical, formed in a Fenton-type reaction, are harmful for most cell components. Its rapid removal is essential for all aerobically living prokaryotic and eukaryotic cells. However, hydrogen peroxide can act as a second messenger in signal transduction pathways, in immune cell activation, inflammation processes, cell proliferation, and apoptosis.

One of the most efficient ways of removing peroxide is through the enzyme catalase, which is encoded by a single gene and is highly conserved among species. Mammals, including humans and mice, express catalase in all tissues, and a high concentration of catalase can be found in the liver, kidneys, and erythrocytes. The expression is regulated at transcription, post-transcription, and post-translation levels. High catalase activity is detected in peroxisomes. More recently, short wavelength UV radiation has been shown to produce reactive oxygen species (ROS) through the action of catalase. This response is thought to act as a mechanism to protect DNA by converting damaging UV radiation into ROS species that can be metabolized and detoxified by cellular antioxidant enzymes. This assay has been validated for serum, EDTA and heparin plasmas from a variety of species.

Assay principle
The Catalase Activity kit is designed to quantitatively measure catalase activity in a variety of samples. A bovine catalase standard is provided to generate a standard curve for the assay and all samples should be read off of the standard curve. Samples are diluted in the provided Assay Buffer and added to the wells of a half area clear plate. Hydrogen peroxide is added to each well and the plate incubated at room temperature for 30 minutes. The supplied Colorimetric Detection Reagent is added, followed by diluted horseradish peroxidase, and incubated at room temperature for 15 minutes. The HRP reacts with the substrate in the presence of hydrogen peroxide to convert the colorless substrate into a pink-colored product. The colored product is read at 560 nm. Increasing levels of catalase in the samples causes a decrease in hydrogen peroxide concentration and a reduction in pink product.

Catalase unit definition
One unit of Catalase decomposes one micromole of hydrogen peroxide per minute at 25°C and pH 7.0.