) expressed inprokaryotic cells was washed by a gradient of urea concentrations and then purified by Ni-NTA agarose affinitychromatography. The target protein appeared as a single peak when eluted by 300 mmol/L imidazole in affinitychromatography. SDS-PAGE analysis and gel-filtration HPLC on a TSK-GEL G2000SWxl column revealed thatrecombinant CAT L was highly purified, and the molecular weight was about 28 kD with purity greater than 95%.The activity assay with Z-Phe-Arg-MCA as a substrate indicated that the recombinant CAT L could combine withits endogenesis inhibitor of Cystatin at a 11 ratio, and thus took on the biological activity of cysteine protease.Balb/C mice were immunized by the purified protein to obtain antiserum, and ELISA showed that the titer of CATL antiserum was higher than 1512000. Western blotting showed that the CAT L polyclonal antibody was highlyspecific for recognizing recombinant CAT L protein expressed in prokaryotic cells. Immunohistochemistry analysisindicated that this antibody also recognized endogenous CAT L protein expressed in the hepatopancreas, muscle,small intestine, heart, and spleen of Jian carp. Based on these results, the polyclonal antibody obtained in thisstudy could be used to detect CAT L expression and distribution in different tissues of fish based on protein level.