The Indirect Antiglobulin (Coombs’) Test Is Specific but Less Sensitive Than the Direct Antiglobulin Test for Detecting Anti-Erythrocytic Autoantibodies and Thereby Immune-Mediated Hemolytic Anemia in Dogs

The immunodiagnostic assessment of dogs suspected of having immune-mediated hemolytic anemia (IMHA) is based on persistent autoagglutination of erythrocytes (after three saline washes), marked spherocytosis, and a positive direct antiglobulin (Coombs’) test (DAT). However, the value of using the indirect antiglobulin test (IAT) for the detection of anti-erythrocytic autoantibodies in serum from dogs suspected of having IMHA is unclear. To evaluate the IAT, leftover serum samples from a large cohort of 94 dogs suspected of having IMHA and for which DAT results were known were incubated with DAT− erythrocytes, and five IAT techniques were performed (in part with different reagents and temperatures): microtiter plate (MICRO), microcapillary, laboratory gel column, gel minitube kit (GEL KIT), and immunochromatographic strip kit. Two IAT techniques (MICRO at 37 °C and GEL KIT with rabbit anti-dog polyvalent reagent) detected autoantibodies against erythrocytes in serum from 53% and 57% of DAT+ dogs, respectively, while other IATs performed less well. Moreover, while the analytic specificity of the IAT methods compared to the DAT ranged from 96–100%, the sensitivity range was only 9–57%. Thus, we still recommend DAT for diagnosis and monitoring of IMHA in dogs but conclude that a positive IAT result may aid diagnostically when serum is available, but fresh red blood cells are not.