Cell assay, total ATP

  1. 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)
  2. 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
  3. Mix gently by vortexing, swirling, or inverting the contents until a homogenous solution is obtained.
  4. Prepare 384- well opaque plates with cells in culture medium. We use 50 ul/ well of total cell and culture medium
  5. Prepare control wells containing medium without cells to obtain a value for background luminescence
  6. 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
  7. Mix contents gently for 2 minutes on an orbital shaker to induce cell lysis
  8. Allow the plate to incubate at room temperature for 10 minutes to stabilize the luminescent signal
  9. Record luminescence (such as using a luminescence 'plate reader')