Paul Nguyen1, Paul Ventura2, Jeremy Lambert2, Eric Veien2, Vladislav Zarayskiy1, Jonathan O’Connell1
1FORMA Therapeutics, Watertown, MA 2Formulatrix, Inc., Bedford, MA
Cellular viability assays are widely used to investigate the potency or toxicity of biologically active molecules. One of the most widespread techniques to measure cellular viability is a luminescent CellTiter-Glo® (CTG) Assay which is a homogeneous method based on quantitation of the ATP present, an indicator of metabolically active cells. The "add-mix-measure" format results in generation of a luminescent signal proportional to the amount of ATP and, thus, proportional to the number of metabolically active cells present in culture. It is designed for use with multi-well formats, making it ideal for automated high-throughput screening, cell proliferation and cytotoxicity assays. However, in our hands, CTG has limitations for use on automatic platforms, due to the fact that the reliability of dispense degrades over extended run times, primarily as a result of liquid dispensers becoming clogged.
We have tested the use of CTG in two liquid dispensers: Multidrop® CombiLiquidDispenser (ThermoFisher) and Tempest® (Formulatrix). The Multidrop consistently provided accurate and precise dispense data during daily operations where many plates were dispensed in rapid succession. However, when runs had periods of inactivity (such as overnight, or longer pace-times), CTG would frequently clog the cartridge lines. Data obtained with Tempest has been found to be more reliable in extended run scenarios. Using the Tempest in a fully automated setting with recirculation turned on allows experiments to be performed in multiple time-based formats, including overnight runs and longer pace-times.
In long run and long pace-time cellular viability studies, the cell viability readout often occurs after a period of long inactivity. While using viscous reagents such as CTG, we have found that in these situations Multidrop Combi Liquid Dispenser (ThermoFisher) cartridge lines are clogged, sometimes beyond repair. Utilizing the Tempest (Formulatrix) with the recirculation feature turned on helps researchers to avoid clogging and perform CTG experiments with a high degree of precision and in a truly automated and time-independent manner.