Identification of an anti‐monocyte monoclonal antibody that is specific for membrane complement receptor type one (CR1)

Abstract

A monoclonal antibody (E11) was produced by immunization of mice with intact human cells of monocyte lineage. Despite the finding that E11 did not inhibit rosettes with C3b-coated sheep erythrocytes (EC3b), several lines of evidence indicated that E11 was specific for complement receptor type one (CR₁). All monocytes, neutrophils, lymphocytes and erythrocytes that reacted with E11 formed EC3b rosettes. The E11 antigen on these cells was shown to be a molecule of 222 × 10 kDa. Treatment of lymphocytes, monocytes, and neutrophils with E11 followed by fluorescein-coupled F(ab′)₂ anti-mouse-IgG at 37 °C in buffer lacking sodium azide, led to capping or apparent endocytosis of the E11 antigen and a diminution in CR₁ activity of 88%, 59% and 25%, respectively. This same treatment had no detectable effect on monocyte or neutrophil CR₃ activity (EC3bi rosettes). Furthermore, with E11-capped lymphocytes, the residual EC3b rosetting was capped directly over the E11-fluorescence cap, whereas EC3d, g rosetting (CR₂ specific) was undiminished and distributed evenly around the circumference of cells containing E11-fluorescence caps. Finally, the binding of E11 to cells was inhibited by the prior treatment of these cells with a well characterized rabbit polyclonal anti-CR₁. These data indicated that E11 was specific for a site in CR₁ that was distal from the C3b-binding site, so that E11 was unable to block CR₁ activity. E11 proved to be useful for identifying CR₁ on various cells in tissue sections, and for quantitating CR₁ on erythrocytes and neutrophils. Erythrocytes and neutrophils from normal individuals were found to bind an average of 610 and 4.6 × 10^{4 125}I-labeled E11 molecules per cell. When E11 was visualized in tissues by immunoperoxidase staining, the cells that apparently contained the greatest amounts of CR₁ were dendritic reticulum cells and kidney podocytes. The E11 reactive dentritic reticulum cells were characteristic of both follicular and diffuse follicular center cell tumors. Lymphocytes from patients with chronic lymphocytic leukemia (CLL) characteristically expressed little E11, confirming earlier studies that CLL cells lacked CR₁ activity detected by EACI-3b rosette formation. Because normal B cells have been shown to express CR₁ at a very early stage of maturation, the absence of CR₁ on CLL cells is discordant with the immature nature of CLL cells defined by immunoglobulin expression.