ATP measurements on single living cells: a dynamic approach

ATP is a good indicator of cellular viability, due to its important role of energy supplier in many biochemical reactions. Contractile phenomena are directly related to its local concentration, and to its mitochondrial rate of production. For these reasons, the developement of a reliable technique to measure the intracellular free ATP concentration on isolated or cultured single cells presents an actual physiological and pharmacological interest. This work describes a method to determine at the cellular scale the intracellular ATP concentration. The luminescence of the firefly Luciferin-Luciferase couple has been already widely used for in vitro and tissular [ATP] measurements, but problems occur in isolated cells to calibrate the light intensity signal-nor the reagents concentration neither the cell volume can be evaluated precisely inside a given cell-the high ATP concentration leads to a fast light intensity decay, with a time constant depending on [ATP], and also on environmental factors such as the local accessibility of oxygen molecules. This latter point will be sought for in the present work