Abstract
The factors contributing to RBC agglutination are complex. The RBC cytoskeleton's participation in and contribution to this phenomenon are difficult to separate from those of the plasma membrane. Immunoreactive, cytoskeleton-free, band 3-enriched microvesicles can be generated from normal RBCs. Band 3 has been defined as an important antigen in autoimmune hemolytic anemia (AIHA). RBC microvesicles devoid of major cytoskeletal proteins were generated and sensitized with eluates obtained from AIHA patients, DAT-positive blood donors, and antisera to common RBC antigens. Monoclonal anti-human IgG was added and agglutination was investigated. Autoantibody-specific binding was evaluated by employing (125)I protein A. RBC vesicle agglutination with a 4+ anti-human globulin score was obtained with 10 autoantibody eluates from AIHA patients and anti-D (3+), but not with eluates from 20 DAT-positive blood donors or antisera directed to eight other common RBC antigens. Microvesicles sensitized with AIHA eluates bound 67 to 167 times as much (125)I protein A radioactivity as did those incubated with buffered normal saline and 18 to 45 times more than vesicles incubated with normal serum. The major proteins of the RBC cytoskeleton are not required for supporting IgG immune-mediated agglutination of RBC microvesicles.
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