Electrogenerated chemiluminescence (ECL) is a light emission phenomenon caused by electrochemically generated radical anions (R•-) and cations (R•+), in which the ion annihilation results in the formation of a pair of excited (R*) and ground state (R) of a luminescent molecule. Here, the ECL properties of pyrene (Py) and 2,7-di-tert-butylpyrene (di-t-BuPy) are reported. It was found that at a commonly employed concentration (1 mM), the ECL spectra were time-dependent because of increasing the oligomer emission and increasing the concentration of R near R*, leading to an enhancement of the excimer emission. At a low concentration range (20-30 μM), the shape of the ECL spectra containing the monomer and excimer emission was determined by isolated pairs of R* and R, which were generated through ion annihilation of R•- and R•+. It was found that in the ECL of Py and di-t-BuPy originated from the isolated pairs of R•- and R•+, 58 and 48% of the excited states were the excimer states, respectively. Diffusion equation analysis indicates that the lower excimer formation in the case of di-t-BuPy is because of a farther initial separation distance between R* and R, i.e., a longer electron transfer distance between the radical ions. The Marcus model for the electron transfer kinetics suggests that the farther electron transfer distance is mainly caused by the larger molecular size, which resulted in a smaller reorganization energy of the solvent acetonitrile molecule. Taking advantage of the photophysical and electrochemical properties of Py and di-t-Bu Py, the monomer and excimer emission in ECL is discussed.