A retrospective case-control study was conducted based on the records of 80 dogs with visceral haemangiosarcoma (HSA) and 200 dogs with various diseases that had clinical features similar to HSA. All dogs were more than 1 year old, had histologically confirmed disease, and had a complete blood count performed prior to the final diagnosis. A standard protocol was used to count acanthocytes on one blood film from each dog. Acanthocyte count had a maximum diagnostic sensitivity of 53.8% (and specificity of 61.5%) at a cutpoint of ≥1 acanthocyte/2,000 red blood cells. A diagnostic specificity of 100% (and sensitivity of 7.5%) was achieved at a cutpoint of >71 acanthocytes/2,000 red blood cells. The precision of acanthocyte count, within and between raters, varied from poor (unweighted kappa = 0.26) to good (weighted kappa = 0.71) due to the subjective nature of the identification of acanthocytes. Although dogs with acanthocytes were more likely to have HSA (P=0.02), and dogs with HSA had higher acanthocyte counts than controls (P=0.003), acanthocyte count had limited ability to distinguish between dogs with HSA and unaffected dogs with similar signs, as indicated by the receiver operator characteristic plot, which lay approximately along the diagonal. There was no level of acanthocytosis at which HSA could be ruled out, and although HSA could be ruled in at counts >71 acanthocytes/2,000 red blood cells, only six of the 80 dogs with HSA in the study could be identified by this cutpoint.