Abstract Brain–computer interface (BCI) technology translates voluntary choices in active command using brain activity. In fact, brain electrical signals, particularly evoked related potentials (ERPs) produced some ms after cognitive tasks, are frequently used to activate commands, being the visual P300-based BCI system, the main interface used for communication and control purposes. In the P300-speller BCI, the user’s task consists in visualizing a matrix of 6 × 6 rows and columns and then focusing attention on a desired character (target, rare event). The aim of this study was to analyze the effect of three different speller sizes on participants’ usability while using a visual P300 BCI system based on the one developed by Farwell and Donchin. The study carried out in this experiment show how important it is to consider the ergonomics of the interface to obtain higher usability.