Objectives Ovarian remnant syndrome (ORS) is suspected when heat signs occur in spayed individuals, but further diagnostic procedures are necessary to exclude other possible oestrogen sources, such as the adrenal gland or exogenous supplementation. Anti-Müllerian hormone (AMH), secreted by granulosa cells or Sertoli cells, serves to differentiate sexually intact from gonadectomised animals and has been described in dogs as a tool for diagnosing ORS. The aim of this study was to evaluate if AMH determination can be used to diagnose ORS in cats. Methods AMH was measured with a chemiluminescence immunoassay in serum samples of 15 sexually intact, 9 spayed and 16 cats with a history of heat signs after spaying. Abdominal ultrasound (n = 13), vaginal smears (n = 7), progesterone measurement (n = 5) and laparotomy (n = 14) were used to determine the presence of ovarian tissue. After surgery, a histological examination of the obtained tissue was performed in the cats with suspected ORS. Results In 15 cats with ORS the AMH serum concentrations were significantly higher than in spayed cats (n = 10; P = 0.025) and significantly lower than in sexually intact cats (n = 15; P = 0.001). Among the cats with ORS, the highest AMH serum concentrations were measured in the queens with cystic ovarian alterations and in one cat from which a whole ovary was obtained. The cat with the lowest AMH serum concentration had a simultaneous high progesterone serum concentration. Cats with ORS did not show any heat signs after surgical removal of the ovarian tissue. Conclusions and relevance A single determination of AMH in blood serum is a useful diagnostic tool for the diagnosis of ORS in cats, regardless of the hormonal activity of the remnant ovarian tissue.