Lymphocyte activation gene‐3 (LAG‐3) is a type I transmembrane protein with structural similarities to CD4. Overexpression of LAG‐3 enables cancer cells to escape immune surveillance, while its blockade reinvigorates exhausted T cells and strengthens anti‐infection immunity. Blockade of LAG‐3 may have antitumor effects. Here, we generated a novel anti‐LAG‐3 chimeric antibody, 405B8H3(D‐E), through hybridoma technology from monoclonal antibodies produced in mice. The heavy‐chain variable region of the selected mouse antibody was grafted onto a human IgG4 scaffold, while a modified light‐chain variable region was coupled to the human kappa light‐chain constant region. 405B8H3(D‐E) could effectively bind LAG‐3‐expressing HEK293 cells. Moreover, it could bind cynomolgus monkey (cyno) LAG‐3 expressed on HEK293 cells with a higher affinity than the reference anti‐LAG‐3 antibody BMS‐986016. Furthermore, 405B8H3(D‐E) promoted interleukin‐2 secretion and was able to block the interactions of LAG‐3 with liver sinusoidal endothelial cell lectin and major histocompatibility complex II molecules. Finally, 405B8H3(D‐E) combined with anti‐mPD‐1‐antibody showed effective therapeutic potential in the MC38 tumor mouse model. Therefore, 405B8H3(D‐E) is likely to be a promising candidate therapeutic antibody for immunotherapy.