Abstract: The present invention relates to a fluorescence immunochromatographic assay kit and an application method thereof, belonging to the technical field of in vitro diagnostic products for animal diseases. The kit comprises a plastic case, a test reagent card and a sample diluent. The case comprises a bottom case and an upper cover, wherein a test strip slot is formed in the bottom case, and a scan window and a sample loading hole are arranged on the upper cover; wherein the test reagent card consists of a sample pad, a binding pad, a nitrocellulose membrane and an absorbent paper which are sequentially adhered on a bottom board; the position of the scan window is matched with the position of the nitrocellulose membrane, and the position of the sample loading hole is matched with the position of the sample pad.