To investigate the antigenic relationship between the macrophage and lymphocyte Fc receptors (FcR), a monoclonal antibody capable of blocking mouse macrophage Fc receptors was selected. Hybrids were formed by fusing the P3U1 mouse myeloma and spleen cells from a rat immunized with the mouse macrophage-like cell lines J774 and P388D1. The Fab fragment of the monoclonal IgG secreted by clone 2.4G2, inhibited the trypsin-resistant Fc receptor II (FcRII), which is specific for immune aggregates of mouse IgG1 and IgG2b, but had no inhibitory effect on the trypsin-sensitive Fc receptor I (FcRI), which binds monomeric IgG2a and erythrocytes coated with IgG2a. Thus, the monoclonal 2.4G2 IgG appeared to be specific for macrophage FcRII. Further evidence that the 2.4G2 IgG was directed against FcRII came from binding studies of the monoclonal antibody to J774 cells and a series of independently isolated variants which do not express FcRII. These variants of J774 bound 5% as much of the monoclonal antibody as the parent line, which bound 600,000 molecules of 2.4G2 IgG per cell. The antigenic relatedness of mouse lymphocyte FcR to mouse macrophage FcRII was demonstrated by the binding of 2.4G2 IgG to FcR-bearing lymphoid cell lines and the inhibition of the lymphocyte FcR by the monoclonal antibody. Preincubation of spleen cells and peritioneal cells with 2.3G2 IgG likewise inhibited rosette formation with ox erythrocytes coated with rabbit IgG. The ability of the hybridoma IgG to inhibit mouse FcRII was independent of the major histocompatibility complex. The 2.4G2 IgG antigenic determinant was not present on rat, guinea pig, rabbit, or human FcR-bearing cells.

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Characterization of a monoclonal antibody directed against mouse macrophage and lymphocyte Fc receptors.

Identification of a monoclonal antibody that specifically recognizes corneal and skeletal keratan sulfate. Monoclonal antibodies to cartilage proteoglycan

Structures resembling nuclei but depleted of protein may be released by gently lysing cells in solutions containing non-ionic detergents and high concentrations of salt. These nucleoids sediment in gradients containing intercalating agents in a manner characteristic of DNA that is intact, supercoiled and circular. The concentration of salt present during isolation of human nucleoids affects their protein content. When made in I-95 M NaCl they lack histones and most of the proteins characteristic of chromatin; in 1-0 M NaCl they contain variable amounts of histones. The effects of various treatments on nucleoid integrity were investigated.

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Characterization of nuclear structures containing superhelical DNA.

The binding properties of surface receptors of immunoglobulins on mouse macrophages were studied with mouse myeloma proteins and normal peritoneal macrophages, thioglycollate-stimulated macrophages, and a macrophage cell line, P388D1. Primary cultures of mouse embryo fibroblasts served as controls. IgG2a proteins were bound strongly;IgG2b was bound weakly (one-twentieth as well as IgG2a);IgM, IgA, and IgG1 were not bound significantly. The number of binding sites per cell for IgG2a was 4 X 10(5) for thioglycollate-stimulated cells and 1 X 10(5) for normal and P388D1 cells. Binding was exothermal: with decreasing temperature the equilibrium (association) constants increased and dissociation rate constants decreased (at 37degreesC the respective values were 2 X 10(7) M-1 and 0.26 min-1, the latter value corresponds to a half time for dissociation of 2.6 min). From the rapidity of association and dissociation, it appears that the surface of the macrophage is in a dynamic equilibrium with IgG2a molecules in the cell's immediate microenvironment. The receptors for IgG2a are clearly specific for determinants in the immunoglobulin constant domain: two IgG2a proteins with greatly different isoelectric points (determined by isoelectric focusing) were bound with the same affinity to the same receptors; moreover, the Fc fragment was bound and Fab fragments were not. The Fc receptors for IgG2a proteins were readily eliminated by exposing macrophages briefly to trypsin. The receptors were regenerated during subsequent cultivation in serum-free medium; regeneration was inhibited totally by cycloheximide and partially by actinomycin D.

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Shunro Sonoda

Papers

Studies of 125I trace labeling of immunoglobulin G by chloramine-T.

Localization of iodine in trace-labeled immunoglobulin G.