HI,
I´m trying to figure out the best method to measure cell proliferation. The WST-8 based cell “counting” method has been used in our lab for years but I’m aware of the fact that this method measures cell metabolism rather than cell number and that the results can be influenced by other factors than just number of cell in a sample.
I would thus like to ask whether there is any added value in using luciferase/ATP based method that also measures cell metabolism and what are the main pros and cons of these methods compared to BrdU incorporation or direct cell ceounting.
Thank you
The main advantages of the ATP/luciferase assay are very high sensitivity (10-100 times more sensitive), speed, and stability of the signal. Unlike WST (and other tetrazolium assays) there is no extended incubation time to convert the reagent into the species that is detected. If you're using 384 well (or more) plates, for sure ATP/luciferase will be preferred.
I recommend looking at the following chapter as a useful reference:
Chapter Cell Viability Assays
Both WST and ATP/luciferase are viability assays, correlated with the number of metabolically active cells. BrdU on the other hand is a proliferation assay: the BrdU is incorporated during DNA synthesis, when the cell is replicating. So it tells you a different answer.
Which assay you use really depends on what you are studying, what question/hypothesis you have. Microscopy—which enables direct counting—is always good to ensure that the assay results make sense: sometimes the assay reagents can react with a compound, confounding the results.
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