The invention discloses a bovine viral diarrhea virus differential detection test strip and a preparation method thereof, which are used for rapid and sensitive detection of BVDV antibodies. The test strip is composed of a support base plate, a coated adsorption material layer, a protein marker bearing material layer, a sample buffer material layer, a liquid absorbent material layer, and an outer protective material; the coated absorbent material is provided with a test line and a quality control line The protein label of the test strip is labeled with colloidal gold; the protein used for the protein label is the phage monoclonal combined with the BVDV antibody obtained by screening from the phage polypeptide library; the polypeptide epitope expressed by the phage monoclonal is LTPHKHHKHLHA. The preparation method comprises the preparation of gold-labeled phage monoclonal, the preparation of gold-labeled pads, the preparation of nitrocellulose membrane, and the assembly of various parts to obtain the finished product. The invention can obtain detection results within ten minutes, is fast and sensitive, has low cost, is simple to operate, and is easy to be popularized and applied in production and scientific research at the grassroots level.