Abstract: The present disclosure provides a test strip for milk immunofluorescence assay (IFA) and use thereof, and relates to the technical field of test strip. The test strip of the present disclosure includes a sample pad, a conjugate pad, a nitrocellulose membrane, and a wicking pad assembled and pasted successively on a PVC backing card; fluorescent latex microsphere-labeled mixed antibodies are coated on the conjugate pad; anti-casein antibody (T1 line), anti-beta-lactoglobulin (BLG) antibody (T2 line), anti-alpha-lactalbumin (ALA) antibody (T3 line), anti-lactoferrin/anti-bovine serum albumin (BSA) antibody (T4 line), and rabbit anti-mouse IgG antibody (C line) are coated on the nitrocellulose membrane, where the T1, T2, T3, and T4 lines are test lines, and the C line is a control line. The test strip of the present disclosure accurately and quantitatively detects the content of casein, BLG, ALA, and lactoferrin/BSA in food, and features easy operation and high accuracy and sensitivity.