Discussion

Recently developed assay technologies make it possible to use multi-well plate readers to measure the number of live or dead cells in culture in real time over a period of days. Live cells are measured in real time by adding a reagent containing a shrimp-derived luciferase and a pro-substrate directly to the culture medium. Only viable cells can convert the pro-substrate into a luciferase substrate and generate light. Dead cells are measured by adding a novel non-toxic, non-permeable DNA-binding dye. Viable cells exclude the dye while dead cells take up the dye, which becomes fluorescent upon binding to DNA. Both assays are non-toxic to cells, so viable cells remain in the sample well following measurement of the live or dead signals. In addition to providing real time kinetic measurements that are valuable for assay development and characterization activities, multiplexing with other assays (e.g. apoptosis, oxidative stress markers or RNA extraction) provides a time saving approach and statistical advantage inherent in taking measurements from the same sample of cells.