Application of a direct flow cytometric erythrocyte immunofluorescence assay in dogs with immune-mediated hemolytic anemia and comparison to the direct antiglobulin test

A direct flow cytometric erythrocyte immunofluorescence assay (FC) was deve loped and compared with the direct antiglobulin test (DAT) for detection of erythrocyte-bound immunoglobulin (IgG and IgM) and complement (C3) in dogs with immune-mediated hemolytic anemia (IMHA). Tests were performed on eryt hrocytes from 13 healthy nonanemic dogs and from 13 anemic dogs with IMHA. The FC: and DAT were negative for erythrocyte-bound immunoglobulin in all h ealthy dogs. The FC was negative for erythrocyte-bound C3 in 12 healthy dog s and positive in I healthy dog, and the DAT was negative for C3 in all hea lthy dogs. Of the 13 IMHA dogs tested for erythrocyte-bound I-G, 12 were po sitive using the FC: and 7 were positive using the DAT. Sensitivity for the detection of erythrocyte-bound IgG in the 26 dogs was 92% for FC and 53% f or DAT. Specificity for detection of erythrocyte bound IgG for FC and DAT w as 100%. The addition of IgM and/or C3 did not increase the sensitivity for FC or DAT. In this group of dogs, the FC provided a more rapid, cost-effec tive, sensitive, objective method to quantitate erythrocyte-bound immunoglo bulin and/or complement compared with the currently used DAT.