An EDTA-associated anti-B agglutinin: the role of ionized calcium

It is believed that EDTA-dependent panagglutination is associated with free carboxylic acids that support reactions of rare autoagglutinins.

An ABO typing discrepancy occurred in an 88-year-old patient. The specificity of his autoagglutinin was demonstrated by panel cell study and absorption tests using normal donors' red cells or immunoadsorbents coated with A, B, or O substances. Inhibition assays were performed to determine whether the autoagglutinin was inhibited by ionized calcium or carboxylic acids. The autoagglutinin had anti-B specificity when tested in the presence of EDTA. It was neutralized by group B secretor saliva and adsorbed by crystalline silica coated with simple B substances with or without EDTA, although it was absorbed by group B red cells only in the presence of EDTA. The agglutinating activity was stronger at 25 degrees C (titer 64) than at 37 degrees C (titer 16) and was destroyed by treatment of the serum with dithiothreitol, which suggests that the autoagglutinin is IgM. This activity also appeared in the patient's serum after dialysis and in an eluate obtained after adsorption with simple B substances, and it was inhibited by the addition of CaCl2 at 0.5 mM or higher concentrations. This suggests that the agglutination is not dependent on EDTA but, rather, on the concentration of ionized calcium. The autoagglutinin failed to react with group B red cells treated with glutaraldehyde for 10 minutes.

An anti-B autoagglutinin was shown to have caused an ABO typing discrepancy in the presence of EDTA. These results suggest that autoagglutination requires an environment with low levels of ionized calcium, but not the presence of carboxyl groups.