- Thaw the Promega CellTiter-Glo Buffer and lyophilized CellTiter-Glo Substrate to room temperature prior to use. For six 384-well plates, we use one 100 ml vial of CellTiter-Glo® Buffer and one component vial of CellTiter-Glo® Substrate (lyophilized)
- Transfer contents of CellTiter-Glo Buffer into the glass bottle containing CellTiter-Glo Substrate. This reconstitutes the lyophilized enzyme/ substrate mixture and forms the CellTiter-Glo Reagent
- Mix gently by vortexing, swirling, or inverting the contents until a homogenous solution is obtained.
- Prepare 384- well opaque plates with cells in culture medium. We use 50 ul/ well of total cell and culture medium
- Prepare control wells containing medium without cells to obtain a value for background luminescence
- Add a volume of CellTiter-Glo Reagent equal to (or as low as 60%) the volume of cell culture medium present in each well. We dispense 27 ul/ well of CellTiter-Glo Reagent to 384-well plates using the WellMate
- Mix contents gently for 2 minutes on an orbital shaker to induce cell lysis
- Allow the plate to incubate at room temperature for 10 minutes to stabilize the luminescent signal
- Record luminescence (such as using a luminescence 'plate reader')