Cytotoxicity study of anticancer compounds using Alamar blue on Cal27 cell lines
Hello,
Dead cells should not interfere with Alamar Blue assay. Alamar Blue is an important redox indicator used to evaluate metabolic function and cellular health. This assay relies on an indicator Alamar Blue that is reduced in living cells. Viable cells continuously convert resazurin to resorufin thereby generating a quantitative measure of viability.
When added to cell culture, the oxidized form of Alamar Blue enters the cytosol and is converted to the reduced form by mitochondrial enzyme activity by accepting electrons from NADPH, FADH, FMNH, NADH and cytochromes. This redox reaction is accompanied by a shift in colour of the culture medium from indigo blue to fluorescent pink. So, dead cells in no way could cause any interference in the assay.
Best Wishes.
Hello,
Janhavi Bhadwalkar, The Alamar Blue assay may not be affected by dead cells. The Alamar Blue Cell Viability Assay reagent assesses the proliferation of mammalian cell lines, bacteria, and fungi in a quantitative manner. The dye contains an oxidation-reduction (REDOX) indicator that fluoresces and changes color as the growth media is chemically reduced due to cell growth. Viable cells convert resazurin to resorufin on a continuous basis, yielding a quantitative measure of viability.
The oxidized version of the Alamar Blue (resazurin) dye is blue and non-fluorescent. Growing cells produce a chemical decrease of the Alamar Blue dye from non-fluorescent blue to red fluorescence in the Alamar Blue assay. A reducing environment (fluorescent, red) is maintained by the ongoing development of viable cells, whereas an oxidizing environment (non-fluorescent, blue) is maintained by the inhibition of growth, which can be detected with a fluorescence or absorbance detector. If your drug works successfully on the cell line (Cal27) you're using, you'll see a concentration gradient dependant fluorescence absorbance reading. As a result, dead cells could not cause any interference in the assay.
Best wishes.
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