The application of methanol sensor-less control in a direct methanol fuel cell (DMFC) system eliminates most of the problems encountered when using a methanol sensor and is one of the major solutions currently used in commercial DMFCs. This study focuses on analyzing the effect of the operating characteristics of a DMFC system on its performance under the methanol sensor-less control as developed by Institute of Nuclear Energy Research (INER). Notably, the influence of the dispersion of the methanol injected on the behavior of the system is investigated systematically. In addition, the mechanism of the methanol sensor-less control is investigated by varying factors such as the timing of the injection of methanol, the cathode flow rate, and the anode inlet temperature. These results not only provide insight into the mechanism of methanol sensor-less control but can also aid in the improvement and application of DMFC systems in portable and low-power transportation.