Initial characterization of monoclonal antibodies against human monocytes.

Abstract

Three monoclonal antibodies against human monocytes have been produced by somatic cell fusion. Extensive specificity analysis suggests that these antibodies react with most if not all human peripheral blood monocytes and not with highly purified T or B cells. Initial chemical characterization of the monocyte antigen recognized by two of these antibodies is presented. The molecule is a single polypeptide chain with an apparent molecular weight of 200,000. These reagents should prove useful in the clinical definition of disorders of monocyte differentiation, in studies of monocyte function, and in the elucidation of the genetics and structure of monocyte cell surface antigens.It is apparent that the macrophage participates in virtually all phases of the immune response (1). To some extent the controversy about the many roles ascribed to cells of the macrophage-monocyte lineage derives from the difficulty encountered in attempts to define this cell morphologically, functionally, or antigenically (2-6). There is general agreement that a cell that is mononuclear, that phagocytoses via an Fc receptor, and that, at the same time, produces lysozyme is a macrophage. Although this definition is useful, it is difficult to utilize such criteria for either the isolation of monocytes or for the elimination of these cells from other cellular populations (7-9).Antigenic analysis of macrophages has been exceedingly complicated although much progress has been made. Heteroantisera to pooled populations of murine and human macrophages have been produced (10-12) and recently Springer et al. (13) described a murine macrophage differentiation antigen, termed Mac-i, by utilizing a monoclonal antibody. Similar studies in man have been limited, although Breard et al. (14) recently described a monoclonal antibody reactive with human peripheral blood monocytes.In this report, we describe the production of three monoclonal antibodies against human monocytes. Their initial characterization suggests their potential utility in defining both the structure of monocyte differentiation antigens and the relationship between monocytes and other cell populations.MATERIALS AND METHODS Immunization Eight-week-old BALB/c mice were immunized with human peripheral blood monocytes obtained from a single healthy donor. Mononuclear cells purified by centrifugation on Ficoll/Hypaque were plated in plastic dishes for 15 hr at 370C.The adherent cells were subsequently harvested and immediately injected into mice. Each mouse received two intraperitoneal injections of 107 cells, 3 weeks apart. Four days after the second injection, mice were sacrificed and their spleens were removed and used for fusion.The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. 6764Cell fusion and cloning of hybrids Two separate fusions were performed between spleen cells and the nonsecreting cell line Sp2/0-Agl4 (15) by...

Induction of monocytic differentiation and bone resorption by 1,25-dihydroxyvitamin D3.

“…The fact many of the latter cells are recognized by a reportedly monocyte-specific antibody suggests that induction by Me2SO leads to incomplete commitment to the granulocyte pathway. On the other hand, the observation that 63D3 recognizes a sizeable proportion of both 1,25(OH)2D3 and Me2SO-treated cells is consistent with the original-observation that this antibody reacts with both granulocytes and monocytes, although much more avidly with the latter than the former (24). This difference in avidity is clearly evident in fluorescence analysis of Me2SO-and 1,25(OH)2D3- (20) 100* 100* 87* GAP 8.3 Leukocytic (21) treated cells (Fig.…”

Delineation of four cell types comprising the giant cell tumor of bone. Expression of Ia and monocyte-macrophage lineage antigens.

“…The antimonocyte monoclonal antibody 63d3 was kindly provided by Dr. J. D. Capra, University of Texas, Southwestern Medical School, Dallas, TX. (7). The specificities of the antibodies directed to the mononuclear phagocyte lineage are summarized in Table I.…”

“…Recently, a number of monoclonal antibodies have been described that detect surface antigens exclusively or predominantly expressed on the mononuclear phagocyte lineage (7)(8)(9). The (10).…”

Evidence That HIV Budding in Primary Macrophages Occurs through the Exosome Release Pathway

“…The following murine monoclonal antibodies (mAbs) were used in flow cytometry, virus capture, and dot-blot assays: anti-MHC I (MHM.5), anti-CD45 (H5A5), and anti-MHC II (MHM.36 and H53) (28); anti-CD63 and biotinylated anti-CD63 (H5C6) (29); anti-CD14 (63D3) (30); anti-Lamp-1 (H4A3) and anti-Lamp-2 (H4B4) (31); anti-CD36 (produced as described previously) (32); anti-CD55, antimacrophage mannose receptor (MMR), and anti-CD81 (Pharmingen); and fluorescein isothiocyanate (FITC)-conjugated anti-CD36, fluorescein isothiocyanate-conjugated anti-CD45, and mouse IgG1 control myeloma (Coulter Immunotech, Hialeah, FL). Secondary antibodies (Texas Red-conjugated goat anti-mouse IgG (Fc␥ fragment-specific), fluorescein isothiocyanate-conjugated goat anti-mouse IgG (Fc␥ fragment-specific), horseradish peroxidase-conjugated goat anti-mouse IgG (heavy and light chain-specific), and rabbit anti-mouse IgG (Fc␥ fragmentspecific)) were from Jackson ImmunoResearch Laboratories, Inc. (West Grove, PA).…”