Diagnostic evaluation of a flow cytometry-based direct antiglobulin test and demographic analysis in dogs with suspected immune thrombocytopenia.

Canine immune thrombocytopenia (ITP) is an autoimmune haematological disorder characterized by antibody-mediated platelet destruction, and diagnosis remains challenging because of limited definitive diagnostic tools. This study evaluated an in-house flow cytometry-based direct antiglobulin test (DAT) for the detection of platelet-bound IgG and investigated demographic factors associated with DAT positivity in thrombocytopenic dogs. A total of 100 healthy dogs and 1127 thrombocytopenic dogs were included. Platelets were stained with anti-CD41/61 and anti-canine IgG antibodies and analysed by flow cytometry. A diagnostic cutoff was established using healthy control dogs and subsequently applied to thrombocytopenic dogs. Breed, age, and sex were evaluated as potential risk factors for DAT positivity. A cutoff value of 10% IgG-bound platelets was established, and 39.3% of thrombocytopenic dogs were classified as DAT-positive. DAT-positive dogs had significantly lower platelet counts than DAT-negative dogs (median, 10,000/µL; mean, 23,665/µL vs. median, 39,000/µL; mean, 53,193/µL; P < 0.05). A weak but statistically significant inverse correlation was observed between the percentage of IgG-bound platelets and platelet count (r = -0.17, P = 0.0062). Mixed-breed and Shih Tzu dogs, young to middle-aged dogs, and intact females had increased odds of being DAT-positive. These findings suggest that integrating flow cytometry-based DAT results with platelet count and demographic characteristics may improve the diagnostic workup for canine ITP and support therapeutic decision-making.