Objective - To determine whether cats in the north-eastern United States develop serum antibodies against antigens of Borrelia burgdorferi and Anaplasma phagocytophilum and whether coinfection with the 2 organisms occurs. Sample population - Serum samples from 84 healthy cats and 9 cats with lameness, fever, anorexia, or fatigue. Procedure - Serum antibodies against B burgdorferi and A phagocytophilum were measured with an ELISA incorporating a whole-cell preparation or purified recombinant antigens, by means of western blot analysis, or indirect fluorescent antibody (IFA) staining. Results - ELISA results indicated that 44 of 93 (47%) sera contained antibodies against ≥ 3 B burgdorferi antigens, whereas 43 (46%) were reactive to whole-cell B burgdorferi. Serum reactivity to protein 35, VlsE, and outer surface proteins A and F was most common. Seropositivity to ≥ 3 antigens occurred at the same rate (5/9) in the 9 ill cats as in the 84 healthy cats (46% [39/84]). Of 13 sera reactive to recombinant antigens, 9 were seropositive as measured by western blot testing with whole-cell antigen. Seropositivity rates of 30% and 38% were detected for antibodies against A phagocytophilum via IFA and ELISA testing, respectively. Fifteen (16%) sera had antibodies against both pathogens. Conclusions and clinical relevance - Cats living in areas infested by Ixodes scapularis ticks are exposed to B burgdorferi and A phagocytophilum and, in some instances, may be coinfected. Most cats appeared healthy. An ELISA incorporating specific recombinant antigens may be used adjunctively with western blot and other assays to confirm B burgdorferi and A phagocytophilum infection in cats.