Existing commercial vaccines against babesiosis contain soluble parasite antigens (SPAs) from in vitro culture supernatants of Babesia canis. This study identified and characterized the specific antigen in SPA serum that confers immunity in vaccinated dogs (ie, canine Babesia antigen [CBA]) and sequenced the gene that encodes it. The gene was then cloned and expressed in Escherichia coli. The recombinant CBA (rCBA) was found to protect against challenge infection in rCBA-vaccinated dogs. The rCBA antigen could replace existing SPA vaccines, thereby eliminating the need for dog blood and serum for production of vaccine.