Bioluminescence atp determination kit

Cells were treated for up to 48 h as indicated in the Results section. Total cellular ATP levels were measured in cell extracts using a bioluminescence ATP determination Kit (#A22066, ThermoFisher Scientific) per the manufacturer's recommendations.

ATP quantification was determined in live cultures using a bio luminescence ATP determination kit (Thermo Fisher Scientific, A22066). The assay employs luciferin that reacts with ATP in the presence of luciferase, which in turn is converted to oxyluciferin. Light is emitted in proportion to the amount of ATP in the cells.^{2 (link)} The assay begins with preparing the standard reaction solution by mixing the components together: 20X TE buffer, ultrapure H₂O, DTT, D-luciferin and firefly luciferase in appropriate quantities as stated in the company’s protocol. ATP standard solutions were prepared by diluting supplied ATP in ultrapure H₂O and the assay standard reaction solution. Thereafter, 500 μl of the standard reaction solution is added to each sample in a clear 24 well-plate and luminescence values were recorded (Tecan Infinite M200). Finally, the absolute ATP values of the unknown samples were found by comparison with known values from the standard curve.