Biofuel cells use chemical reactions and biological catalysts (enzymes or microorganisms) to produce electrical energy, providing clean and renewable energy. Enzymatic biofuel cells (EBFCs) have promising characteristics and potential applications as an alternative energy source for low-power electronic devices. Over the last decade, researchers have focused on enhancing the electrocatalytic activity of biosystems and on increasing energy generation and electronic conductivity. Self-powered biosensors can use EBFCs while eliminating the need for an external power source. This review details improvements in EBFC and catalyst arrangements that will help to achieve complete substrate oxidation and to increase the number of collected electrons. It also describes how analytical techniques can be employed to follow the intermediates between the enzymes within the enzymatic cascade. We aim to demonstrate how a high-performance self-powered sensor design based on EBFCs developed for ethanol detection can be adapted and implemented in power devices for biosensing applications.