The invention discloses an immune chromatography test paper strip for the quantitative determination of clenbuterol based on up-conversion fluorescent nanoparticle label and a preparation method thereof. The test paper strip comprises a supporting layer, an adsorption layer, and a protection layer, wherein the adsorption layer comprises an adsorption fibrous layer, a fluorescent antibody fibrous layer, a cellulose film and an absorbent material layer at the handle end, the cellulose film is provided with detection blots printed by using a CL coupling carrier protein solution and control blots printed by using goat anti-mouse IgG antibodies, and the fluorescent antibody is an NaYF4:Yb:Er nanoparticle labelled CL monoclonal antibody or polyclonal antibody. According to the invention, the application of immune chromatography based on the up-conversion fluorescent nanoparticle label in the quantitative determination of CL residues is realized, so that the detection of the CL residues has no background interference; and the test paper strip has the advantages of strong specificity, high sensitivity, simple, and accurate detection, low cost, wide application scope, and easiness in popularization and application.