Single entity electrochemistry experiments are typically motivated by the need to reveal how heterogeneity affects performance within inherently diverse nanoparticle populations. Here we show that a commonly used supporting electrode, tin‐doped indium oxide (ITO), can also play a significant role in creating heterogeneity in nanoparticle electrochemical responses. To investigate the impact of the substrate, we optically monitored the electrodissolution kinetics of gold nanoparticles on ITO thin films with similar resistivity from two different suppliers. The ITO from the two suppliers showed marked differences in the gold electrodissolution kinetics, with ITO from one of the suppliers even producing poor sample‐to‐sample reproducibility across substrates within the same lot number. The role of nanoparticle size and surface effects were accounted for in our analysis to validate that the observed heterogeneity is dominated by the ITO electrodes. The results show that the role of the supporting electrode cannot be ignored when performing single entity structure‐function studies.