cell viability/proliferation kit (MTT-assay, XTT assay WST-1)

Introduction

Tetrazolium salts (e.g., MTT, XTT, WST-1) are particularly useful for cell viability & proliferation. They are cleaved to formazan by the 'succinate-tetrazolium reductase' system (EC 1.3.99.1), a component of the mitochondrial respiratory chain. This it is only active in metabolically intact cells.

MTT assay

Assay uses:

• quantitative determination of cellular proliferation & activation - response to growth factors or cytokines)
• quantitation of antiproliferative or cytotoxic effects
• quantitation of in vitro tumor cell chemosensitivity
• assessment of photoradiation therapy
• screening of anticancer compounds
• MTT dissolved in agarose overlays is used to improve visualization of virus-infected cells in plaque assays

Advantages

• no radioactive isotopes are used
• tetrazolium-salt conversion strongly correlates with metabolic acvtivity
• low cell numbers can be used
• the use of multi-well ELISA readers allows processing of large numbers of samples
• no washing steps or additional reagents are required

Principle

• based on the cleavage of the yellow tetrazolium salt MTT into purple formazan by metabolically active cells
• MTT formazan crystals are insoluble in aqueous solution, but may be solubilized (by addition of solution included in the kit & incubating overnight 37 C, 6.5% CO2)
• solubilized formazan product can be photometrically quantitated using an ELISA reader
• an increase in the number of living cells results in an increase of total metabolic activity which leads to more color formation

XTT assay

Assay uses:

• used for the same applications as the MTT assay

Assay principle:

• based on the conversion of the yellow tetrazolium salt XTT into an orange formazan dye by metabolically active cells
• in contrast to the purple formazan crystals in the MTT assay, the XTT formazan dye is soluble in aqueous solutions, thus be directly quantitated with an ELISA reader

WST-1 assay

Assay principle:

• WST-1 yields water-soluble cleavage products, like XTT, which can be measured directly
• see XTT assay

WST-1 advantages over XTT:

• WST-1 is more stable than XTT
• WST-1 can be used as a ready-to-use solution & can be stored at 4 C for several weeks without significant degradation
• WST-1 has a wider linear range & shows accelerated color development compared to XTT

More general terms

• biochemistry