The invention belongs to the technical field of biology, and particularly relates to a full-swine-origin African swine fever virus monoclonal antibody as well as a preparation method and application thereof. The monoclonal antibody comprises heavy chain constant regions, heavy chain variable regions, light chain constant regions and light chain variable regions, the amino acid sequences of the heavy chain variable regions are as shown in SEQ ID NO: 1 or SEQ ID NO: 5, and the amino acid sequences of the light chain variable regions are as shown in SEQ ID NO: 2 or SEQ ID NO: 6. The full-swine-origin monoclonal antibody specifically aiming at the African swine fever virus retains natural pairing of light chain and heavy chain variable regions, has the advantages of good gene diversity, full swine origin, good antibody affinity, strong specificity and the like, and can be efficiently expressed in a eukaryotic expression system. According to the invention, a novel raw material is provided for developing a diagnostic kit for African swine fever virus and a new technical support is also provided for developing a preventive and therapeutic drug of a full-swine-origin monoclonal antibody.