Quantitative Immunology of Immune Hemolytic Anemia

INTRODUCTION The commonest form of autoimmune hemolytic anemia is characterized by the presence of antibody of the IgG immunoglobulin class on the red cell (1). These antibodies characteristically react with red cell antigens at body temperature, and they fix complement relatively inefficiently since two antibody molecules in juxtaposition are required to initiate a complement sequence (2). The exact mechanism by which the red cells are destroyed in this syndrome is not completely understood. Recently it has been demonstrated that the presence of the IgG antibody on the surface of the red cell may cause it to adhere to monocytes and splenic macrophages (3, 4). In this reaction, a portion of the red cell memReceived for publication 20 July 1970 and in revised form 7 December 1970. brane may be removed, resulting in spherocytosis. Sequestration of the cells by elements of the reticuloendothelial system, especially the spleen, is probably responsible for the ultimate destruction of the cells. When complement is fixed, direct cytolysis may account for a small part of the cellular destruction. The rate of hemolysis in patients with hemolytic anemia due to warm-reacting antibody may vary greatly, and a relationship between the degree of anemia or the rate of hemolysis and the amount of antibody on the red cell surface has been suspected but has not been established (5, 6). The investigation of this relationship has been hampered by the difficulty in determining the amount of antibody on the cells and in the serum of patients with this disease. The amount of antibody on the cell has been estimated by elution (7), with radiolabeled antiglobulin (6) and, more recently, by an indirect estimate of the amount of anti-IgG which adheres to cells coated with IgG (8). In the present studies, we have undertaken to estimate the amount of antibody present on the red cell surface by measuring the fixation of the first component of complement, C1. Since IgG autoimmune antibodies fix complement only very poorly, if at all, we have reacted IgGcoated cells with rabbit anti-IgG. The combination of two anti-globulin molecules brings about the fixation of a C1 molecule (9). The concentration of anti-globulin can be adjusted so that the direct proportion exists between the number of Cl molecules fixed and the number of IgG autoimmune antibody molecules present on the red cell. By this method, we are able to derive a minimum but proportional estimate of the number of antibody molecules present on the surface of the red cell. The rate of hemolysis in these patients was measured by a modification' of the endogenous carbon monoxide 1Logue, G. L., W. F. Rosse, W. T. Smith, H. A. Salzman, and L. A. Gutterman. 1970. Endogenous carbon monoxide production measured by gas phase analysis: an estimation of heme catabolic rate. Submitted for publication. 734 The Journal of Clinical Investigation Volume 5