One of the most enigmatic features of humoral immunity is the prevalent presence of circulating autoantibodies against IgG. These autoantibodies consist of several subsets, including rheumatoid factors, anti-Fab/anti-F(ab')2-autoantibodies, and anti-idiotypic antibodies. Anti-IgG autoantibodies can impair the safety and efficacy of therapeutic antibodies and interfere with immunogenicity tests in clinical trials. They can also cross-react with allospecific IgG, presenting as heterophilic antibodies that interfere with diagnostic immunoassays. Owing to these factors, recent years have seen a resurgent interest in anti-IgG autoantibodies, but their underlying clinical significance, as well as biological roles and origins, remain opaque. Increased knowledge about canine anti-IgG autoantibodies could facilitate the development of canine immunotherapies and help in understanding and counteracting immunoassay interference. This study investigated the clinical significance and interconnection of heterophilic antibodies, anti-Fab, and anti-F(ab')2-autoantibodies in dogs. We performed a 2-year prospective follow-up of dogs with heterophilic antibodies and analyzed serum for anti-Fab and anti-F(ab')2-autoantibodies. Canine heterophilic antibodies can persist for at least 2 years in serum. A widespread occurrence of anti-Fab and anti-F(ab')2-autoantibodies was found, with reactivity to cryptic epitopes in the IgG hinge region and sporadic cross-reactivity with mouse IgG. Canine anti-Fab and anti-F(ab')2-autoantibodies are thus potential sources of clinical immunogenicity and immunoassay interference.