Abstract: The present invention provides an A-FABP neutralizing monoclonal antibody or antigen-binding fragment thereof and a preparation method and use thereof, which comprises: complementarity determining regions 1 to 3 of a heavy chain variable region, the amino acid sequences of which are set forth in SEQ ID NOs:1-3, respectively, and/or complementarity determining regions 1-3 of the light chain variable region, the amino acid sequences of which are set forth in SEQ ID NOs:4-6. This monoclonal antibody is highly specific for A-FABP and has no cross-reactivity with human and mouse epidermal FABP and heart FABP; it has a good binding affinity for human A-FABP and mouse A-FABP; and its treatment alleviates MCAO-induced ischemic stroke injury in mice, which is associated with alleviated BBB disruption and reduced activation of pro-inflammatory JNK signaling; treatment of healthy mice for 21 days does not change physiological parameters, indicating its safety.