All products have special prices for bulk purchase, please contact us for more details if required.

Cat. No.: XOISKW8-100 (for 100T)

Description

Xanthine Oxidase Inhibitor Screening Kit with WST-8, also known as the XO Inhibitor Screening Kit with WST-8 or XOD Inhibitor Screening Kit with WST-8, is a reagent kit designed for high-throughput screening of xanthine oxidase (XO) inhibitors. It utilizes a WST-8-based colorimetric reaction for simple, rapid, and sensitive detection through absorbance measurements.

Xanthine oxidase (XO or XOD) is an important enzyme in nucleic acid metabolism within the body. It catalyzes the oxidation of hypoxanthine to xanthine and further oxidizes xanthine to uric acid, while generating hydrogen peroxide and superoxide anion through a series of oxidation reactions. This enzyme plays a significant role in purine catabolism across various species, including humans. In humans and other primates, XO is typically present in serum, lungs, liver, and intestinal mucosa. In rodents, XO is expressed in most tissues. The activity of XO in blood is usually low, but during influenza A infection, XO activity in serum and lungs increases. When liver function is impaired, XO is released into the bloodstream in large amounts, making blood XO levels a sensitive indicator for assessing severe liver damage (e.g., jaundice). XO may also be related to the pathogenesis of gout, as xanthine oxidase is involved in the metabolic pathway of uric acid formation. Therefore, xanthine oxidase inhibitors such as allopurinol are used to treat gout. Additionally, studies have shown that inhibiting XO activity can alleviate cardiovascular diseases and chronic obstructive pulmonary disease.

Components

• XO Assay Buffer
• XO (20X)
• XO Indicator (10X)
• XO Substrate (10X)
• Febuxostat (10µM)

Principle

Under the catalysis of XO, the substrate generates superoxide anions (O2.-), which then react with WST-based indicators to produce a water-soluble formazan dye. The formazan dye has a maximum absorption peak around 450 nm. By measuring the absorbance at 450 nm, xanthine oxidase activity can be detected with high sensitivity. If a xanthine oxidase inhibitor is added during the reaction, the formation of the formazan dye will be inhibited. The amount of formazan generated is inversely proportional to the inhibitory effect of the inhibitor, allowing for the detection of the inhibitor's effectiveness.

Features

• This kit includes a positive control for the inhibitor, making it convenient to use. The kit provides XO Assay Buffer, xanthine oxidase (XO), XO Substrate, the positive control febuxostat, and an indicator (XO Indicator) that can be catalyzed by XO to produce fluorescence. Febuxostat, the positive control, is a selective non-purine XO inhibitor commonly used in clinical settings for the treatment of gout and hyperuricemia.
• This kit offers rapid detection with high sensitivity. The entire detection process can be completed in about 40 minutes. The amounts of XO and XO Substrate used in the kit have been optimized to detect inhibitors with very low IC50 values as well as those with higher IC50 values.
• We also provide Amplex Red Xanthine Oxidase Inhibitor Screening Kit.

Storage

Store at -20ºC for one year. XO Indicator should be stored protected from light.

Precautions

• The XO Assay Buffer needs to be completely thawed and equilibrated to room temperature before use, as this may affect the test results. XO should be handled on ice. After use, all reagents should be stored according to the conditions specified in the kit.
• Solvents used to prepare the test inhibitors may interfere with the assay. It is recommended to use XO Assay Buffer, Milli-Q grade water, or DMSO to prepare and dilute the test inhibitors, and add an equal volume of solvent to the control wells to exclude interference. If other solvents are used, ensure that the solvent concentration in the final reaction system does not exceed 5%. If it exceeds 5%, set up a solvent control group to test the solvent's effect on enzyme activity and exclude interference.
• The quantities of XO, XO Indicator, XO Substrate, and other reagents are small, so please centrifuge briefly before use and mix them thoroughly.
• This product is intended for scientific research use by professionals only and is not for clinical diagnosis or treatment, food, or drugs, nor should it be stored in a regular household.
• For your safety and health, please wear a lab coat and disposable gloves while handling.

Instruction: Protocol

Related: Amplex Red Xanthine Oxidase Inhibitor Screening Kit