AbstractAutoimmune hemolytic anemia (AIHA) is a potentially severe disease in which red blood cells (RBC) are destroyed by IgG anti-RBC autoantibodies which can lead to hemolysis. We recently found IgG Fc-glycosylation towards platelet and RBC alloantigens to be skewed towards decreased fucosylation, increased galactosylation and sialylation. The lowered core-fucosylation increases the affinity of the pathogenic alloantibodies to FcγRIIIa/b, and hence RBC destruction. It is known that in autoimmune diseases plasma IgG1 galactosylation and sialylation are lowered, but Fc-glycosylation of RBC-specific autoantibodies has never been thoroughly analyzed. We investigated by mass spectrometry the N-linked RBC autoantibody and plasma IgG1 Fc-glycosylation in relation to occurrence of hemolysis for 103 patients with a positive direct antiglobulin test (DAT). We observed that total IgG1 purified from plasma of patients with RBC-bound antibodies showed significantly decreased galactosylation and sialylation levels compared to healthy controls, similar to what previously has been shown for other autoimmune diseases. The anti-RBC- autoantibodies showed a profile with even lower galactosylation, but higher sialylation and lower bisection levels. In contrast to alloantibodies against RBCs, RBC-bound IgG1 Fc-fucosylation was not different between healthy controls and patients. Analysis of anti-RBC Fc-glycoprofiles suggested that lower bisection and higher galactosylation associate with lower Hb levels.