CyQUANT™ XTT Cell Viability Assay

The CyQUANT XTT Cell Viability Assay is a complete, optimized assay that generates a consistent colorimetric detection of viable mammalian cells. The assay kit consists of two reagents, XTT Reagent and Electron Coupling Reagent. XTT Reagent is used to assess cell viability as a function of cellular redox potential, and the electron coupling reagent improves the dynamic range of the assay.

The assay is sufficient for ten 96-well plates and consists of 10 bottles of XTT Reagent and 10 tubes of Electron Coupling Reagent. One bottle of each (sufficient for one 96 well plate) are thawed, mixed together, added to cells, and incubated. The viability signal is detected using an absorbance-based microplate reader. In actively respiring viable mammalian cells, the water-soluble XTT reagent is reduced and converted to an orange-colored formazan product.

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Features of the CyQUANT XTT Cell Viability Assay include:
• Highly reproducible, absorbance-based viability assay
• Improved dynamic range and sensitivity compared to other colorimetric viability assays
• Simple mix-and-read format, configured for one 96-well plate at a time
• Non-toxic, continuous assay—multiple time points can be collected, no cell lysis

Measuring changes in cell viability is a fundamental method for assessing cell health, determining genotoxicity, and evaluating anti-cancer drugs. In addition to being cost effective, colorimetric viability assays generate consistent results with low well-to-well variability. Unlike other commercially available colorimetric mammalian cell viability assays, XTT-based assays display a high dynamic range and low variability. In addition to improved performance, the assay is a continuous assay and requires no cell lysis or solubilization.

The CyQUANT XTT Cell Viability Assay is a complete and optimized kit for the detection of mammalian cell viability. The assay kit includes XTT Reagent (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) and Electron Coupling Reagent. The XTT reagent, which is a tetrazolium-based compound, is sensitive to cellular redox potential and in the presence of actively respiring cells converts from a water-soluble compound to an orange-colored formazan product. The sensitivity and consistency of the assay is significantly increased when used with the electron coupling reagent.

Since no cell lysis or solubilization are required for detection, the XTT assay can be employed as a continuous assay and several time points can be analyzed. Additionally, the CyQUANT XTT assay generates viability results from various types of cell lines, including primary and suspension cells.

For each 96-well plate, one bottle of XTT Reagent and one tube of Electron Coupling Reagent are thawed, mixed to create a stock solution, and added to cells, with cell viability detected after a 2–4 hour incubation. The assay kit supplies all the materials needed for ten 96-well plates.

It is recommended that the XTT/Electron Coupling reagent stock solution be used soon after mixing. Assay performance is compromised and sensitivity is reduced if the stock solution is kept at room temperature for extended periods of time or subject to cycles of freeze/thaw. To ensure assay performance and reproducibility, the CyQUANT XTT Cell Viability Assay consists of separate XTT and Electron Coupling reagents that are mixed.