LE2E9- and mAb 2A9-like anti-C1 IgG contribute to patient’s total anti-C1 IgG population. Purified native HSA-hC1 domain and its variants E2066D and F2068H were immobilized on microtiter plates, and antibody binding from patient plasma was detected with an HRP-conjugated goat anti-human IgG. Antibody binding to immobilized HSA alone (background) was subtracted. The loss of IgG binding to the C1 domain variants E2066D (black bars) and F2068H (gray bars) compared with HSA-hC1 was calculated in percentage, and the results are depicted for the 9 of 30 patients with AHA and 1 of 20 patients with HA and inhibitors for which binding was reduced ≥15%. The experiments were repeated 2 times with similar results.