After the cleavage of disulfide bonds of macroglobulin isolated from channel catfish (Ictalurus punctatus), an electrophoretically fast-moving polypeptide, which resembled human J chain, was released. On a Sephadex G-200 column equilibrated in 5 M guanidine, the elution position of the J chain overlapped with the descending part of the L chain peak. Further purification was achieved by DEAE ion-exchange chromatography. The isolated polypeptide, which had a molecular weight of 14,800 ± 500, as determined ultracentrifugally by sedimentation equilibrium in 5 M guanidine, contained 7% carbohydrate with one residue of fucose, two of mannose, one of galactose, two of glucosamine, and one of sialic acid per chain. A comparison of catfish and human J chain amino acid analyses showed the former to have a higher content of serine, glycine, and phenylalanine and a lower content of aspartic acid, isoleucine, and arginine. Tryptic peptide maps of catfish and human J chains revealed very few common peptides. Rabbit and guinea pig antisera to human J chain did not cross-react with catfish J chain. Untreated, resuced and alkylated, S-sulfonated, or cyanogen bromide cleaved macroglobulin from the gar (Lepisosteus osseus) contained no polypeptide analogous to either catfish or human J chain by the criteria employed in this study.