Characterization of Autoantibodies in Mixed-Type Autoimmune Hemolytic Anemia

Abstract

Three of 46 patients with autoimmune hemolytic anemia (AIHA) satisfied the diagnostic criteria for mixed-type AIHA in which both warm-type and cold-type autoantibodies against red blood cells (RBCs) are present. The specificities of these autoantibodies were analyzed. All of the warm-type autoantibodies were IgG-x, and the specificities were not serologically classified. The autoantibody of patient 1 reacted to the 41- and 80-kD peptides on immunoblotting, and the epitope corresponding to it was papain sensitive. Two warm-type autoantibodies from patients 2 and 3 resembled each other in serological analyses and reacted with a protease- and neuraminidase-resistant antigen. However, the antigen corresponding to the autoantibody of patient 3 was located on the 37-kD peptide by immunoprecipitation. All of the cold-type autoantibodies were IgM-x and showed high titer and high thermal amplitude. According to the reaction pattern with untreated and enzyme-treated RBCs, the cold-type autoantibodies of patients 2 and 3 were revealed to be anti-Om and anti-I, respectively. In patient 1, the cold-type autoantibody was characterized as having high affinity for autologous RBCs, but its specificity was unclassified. The antigens corresponding to the cold-type autoantibodies were not located by immunoblotting and immunoprécipitation. These serological and immunochemical approaches to autoantibodies in mixed-type AIHA revealed that the warm and cold components recognized the different antigens.

Systemic lupus erythematosus presenting with mixed‐type fulminant autoimmune hemolytic anemia

“…In addition, marked hypergammaglobulinemia and presence of monoclonal IgM-κ antibodies are also compatible with mixed AIHA features. 8,9 Cold autoantibodies, such as cold agglutinin and Donath-Landsteiner antibodies, were not detected in the present patient. Moreover, agglutinative reaction improved 1 week after the initiation of prednisolone therapy.…”

Mixed‐Type Auto‐Immune Haemolytic Anaemia in a Patient with HIV Infection

“…Cold autohaemoagglutinin titres and specificity testing were performed using heat-separated serum against group 0 normal adult and cord RBC as previously described [20]. Reduction of IgM was performed using 2-mercapto-ethanol(2-ME) [21]. Cold-reacting autoantibody was isolated from the heat-separated serum by affinity chroniatography using the lmmunopure IgM purification kit (Pierce Europe B.V. Oud-Beijerland, The Netherlands) according to the manufacturer's instructions.…”

“…Purified and concentrated IgM autoantibody (0.05 ml) was subsequently incubated for 2 h at 4°C with 0 test RBCs (0.05 ml of a 3% suspension) and group AB fresh serum from normal donors (0.05 ml). Agglutination was subsequently scored visually, while complcment fixation was assayed by washing RBC three times with saline and then performing a DAT using anti-C3d antiserum, as previously reported [21].…”

Mixed‐type autoimmune hemolytic anemia: differential diagnosis and a critical review of reported cases

“…Such cases are referred to as "combined warm and cold AIHA" or mixedtype (Mx) AIHA 6 and represent approximately 6 to 8 percent of all AIHA. [7][8][9] The serologic characteristics in these patients are the presence of a cold AB with a high thermal amplitude (>30°C) in association with a WAB. 6 In our experience, Mx AIHA is extremely rare, and there is evidence that some of the reported cases do not fulfill the criteria of this diagnosis.…”