The authors describe a new kind of adenosine triphosphate (ATP) assay. It is based on the use of gold nanoparticles (AuNPs) coated with 4-mercaptophenylboronic acid (MPBA) as the recognition element for ATP. MPBA has a specific affinity for AuNPs through Au-S interaction, and three boronic acid groups undergo condensation to form a boroxine ring. This induces the aggregation of AuNPs and a visible color change from red to blue. However, in the presence of ATP, the boronic acid group of MPBA preferentially binds to the 2’, 3’-hydroxy group of ATP to form a stable boronate ester. Hence, the aggregation of AuNPs is progressively decreased as the concentration of ATP increases, and the color change is increasingly reversed. The ratio of absorbance at 520 and 683 nm increases linearly in the 8 to 100 μM ATP concentration range, with a 0.12 μM limit of detection (at an S/N ratio of 3 σ). The colorimetric assay was successfully applied to the determination of ATP in T47D breast cancer cells and in cultured cells with added anticarcinogen. Graphical abstract Schematic of a colorimetric assay for the visualization and sensitive and selective detection of adenosine triphosphate (ATP) based on the use of gold nanoparticles (AuNPs) coated with 4-mercaptophenylboronic acid (MPBA). The assay was applied to the determination of ATP in T47D breast cancer cells and in cultured cells with added anticarcinogen.