ATP Quantification

atp quantification


Follow these step-by-step instructions to perform an ATP quantification for 3D bioprinted constructs. This protocol uses a luminescent assay to determine the number of viable cells in culture.

Note: This is a luminescence-based assay and is unlikely to show signal with accuracy on a normal plate reader equipped for fluorescence.


  • CellTiter-Glo® 3D reagent 
  • Cell culture medium (stored in refrigerator)
  • Pipettes and Pipette tips
  • Black 96 well plates
  • At least 5 samples of all groups to be tested


  1. Remove CellTiter-Glo® 3D solution from freezer and allow to thaw to room temperature on counter. 
    1. Note: Do not thaw this solution in water bath – container may break.
  2. Mix with media at a 1:1 ratio.
  3. Add mixture of media and CellTiter-Glo® to samples (for samples in 96 well plate, add 100 µL). Allow to incubate for 1 hour and 20 min at 37˚C on a shaker. You will need at least 5 samples per group. Be sure to include negative control wells.
  4. After incubation, take 200 µL of solution from each sample and pipette into a black 96 well plate.
  5. Record luminescence following the CellTiter-Glo® protocol on microplate software.
  6. To quantify data, save each group in an excel sheet, and calculate the ATP as compaired to your negative control and positive control.

We hope this protocol helps you easily determine the number of viable cells in your 3D bioprinted construct using an ATP quantification method. For more assay protocols – click here.

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