L-amino acid oxidases (LAAOs) are an important class of oxidoreductases that participate in the metabolism of amino acids. Employing oxygen molecule as the electron acceptor, LAAOs catalyze the oxidative deamination of L-amino acids and produce corresponding alpha keto acids, ammonia (NH₃) and hydrogen peroxide (H₂O₂). LAAOs with high substrate specificity can recognize the preferred amino acid specially. Therefore, LAAOs with high substrate specificity are promising in the application of chiral amine compounds resolution, alpha keto acid biosynthesis, and especially, in the detection of certain amino acids in clinical and food samples. Therefore, specific LAAOs has attracted wide attention in recent years. In this review, we will focus on the recent progress in LAAOs researches. We will emphasize the substrate specificity, structure-function relationship, and family evolution of these enzymes. We will also discuss the applications of these enzymes in biocatalysis and amino acid detection. This review will provide guidance for molecular mechanism research and industrial applications of LAAOs.