WST-1 Cell Proliferation Assay Kit From Roche Applied Science

The WST-1 Cell Proliferation Assay from Roche is a colorimetric assay that is based on the cleavage of a tetrazolium salt, MTS, by mitochondrial dehydrogenases to form formazan in viable cells. The greater the number of viable, metabolically active cells, the greater the amount of formazan product produced following the addition of WST-1. By detection of the formazan level in the cells, we can quantify the cell number. The assay can be used not only for quantifying cell proliferation, but cytotoxicity can also be measured. WST-1 can be used to assay either adherent or suspension cells. The WST-1 assay is very sensitive; it can detect 500 to 50,000 cells in a single well of a 96-well plate. Another advantage of the WST-1 assay is that there is no need to wash or harvest cells for the assay. You do not need to remove culture media or wash with PBS for this assay. It is simple, just add 10 ul of WST-1 reagent to 100 ul of media in a 96-well plate. The reduced handling required with WST-1 means that there is a reduced chance of errors due to pipetting/handling.

I typically use adherent cells, such as HeLa and HEK293T cells. I use a hemocytometer to count cells and seed 5000 cells/well. Those seeded cells are incubated overnight at 37°C with 5% CO₂ in a humidified atmosphere. The next day, cells are treated with apoptotic inducers, cell proliferation triggers, or cytotoxic reagents and then 10 ul/well of WST-1 is added and incubation is continued for an additional 1−2 hrs at 37°C with 5% CO₂ in a humidified atmosphere. Here is one of another advantage of the WST-1 assay: the color change (to yellow) is visible to the naked eye. After incubation, I thoroughly shake the plate for 1 min. Absorbance is then measured at 450 nm versus a 650 nm reference by using a plate reader to compare my experimental samples to controls. Typically, I have used the Spectra MAX 190 microplate reader and SoftMax Pro software from Molecular Devices. I analyze my results by exporting the data to MS Excel and expressing the mean ± SD for each sample. I generally run my samples in triplicate and did not get much deviation between samples. In my experience, this assay is easy to use and generate reliable results.