Lactic acid (C₃H₆O₃), also known as 2-hydroxypropionic acid, propanoic acid, is a type of hydroxy acid. It is an essential metabolite of human and microbial cells. In diagnosis and medical management, determination of lactate level in serum is greatly required, and it is also important to measure lactate in fermentative foods to access their quality. Therefore, how to detect lactic acid in different samples with high throughput has become the focus of different researches. The traditional lactic acid detection methods are complicated, time-consuming and laborious, or requires expensive detection equipments. However, the electrochemical enzymatic L-lactate biosensors combining the robustness of electrochemical techniques with the specificity of biological recognition processes showed great advantages over the conventional analytical techniques in size, cost, sensitivity, selectivity, response speed and sample pre-treatment, which show a broad application prospects. There are two main types of lactate biosensors based on L-lactate oxidase (L-LOD) and L-lactate dehydrogenase (L-LDH). Designing a successful enzyme-based L-lactate biosensor requires assembling the enzyme onto a solid carrier and selecting an appropriate transduction strategy between the enzyme and the electrode. Due to the restriction of enzyme molecular structures, reaction mechanism and electrode materials, the traditional lactate biosensors have some limitations in sensitivity, selectivity and stability. Therefore, an increased research was performed to improve the performance of lactate sensors according to the characteristic of the enzymes and the electron transfer type. In this paper, we provide an overview of the structural characteristics, origin and catalytic mechanism of L-LOD and L-LDH, and discuss three strategies, including electrode material modification, enzyme immobilization and enzyme engineering modification, to improve the performance of enzyme electrode based lactate biosensors. In addition, the lactate biosensors were compared and analyzed on the basis of different carriers including membrane, transparent gel matrix, hydrogel carrier, nano-particles, etc. Finally, we comprehensively described the merits and demerits of current commercial lactate sensors and preconceive how emerging new technologies may benefit to future lactate biosensor design.