The present invention discloses a fluorescent quantitative test strip for the porcine epidemic diarrhea virus (PEDV), and a preparation method and application thereof. The fluorescent quantitative test strip comprises a sample pad, a fluorescent label binding pad sprayed with a fluorescent label probe, a nitrocellulose membrane sprayed with a detection line T and a quality control line C, and a piece of absorbent paper that are arranged sequentially in the order of connection. The fluorescent quantitative test strip further comprises a plastic bottom liner arranged in the lower part. The plastic bottom liner is configured to provide an assembly platform; the fluorescent label binding pad is sprayed with a PEDV monoclonal antibody fluorescent microsphere label and a goat-anti-rabbit IgG fluorescent microsphere label; and the nitrocellulose membrane is sprayed with a detection line T formed by the PEDV monoclonal antibody and a quality control line C formed by the goat-anti-rabbit IgG polyclonal antibody. Experiments have proved that by using the test strip provided by the present invention, the sensibility is at least 10 times higher than that by using the existing RT-PCR and colloidal gold test strip detection method, the test sample is initially quantified, and the test result is digitized, so that the human subjective bias is avoided, and a technical means is provided for early diagnosis of the PEDV.