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

Abstract

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.

Expression of BCR – ABL in M1 myeloid leukemia cells induces differentiation without arresting proliferation

“…M1 and M1.210 cells were labelled with biotinylated monoclonal antibodies recognizing F4/80 (Austyn and Gordon, 1981) and F c gRII (Unkeless, 1979) or an isotype control (Pharmingen, San Diego, CA, USA) as described previously (Smith et al, 1997). Speci®c binding was revealed with streptavidin-coupled phycoerythrin (Caltag Laboratories, West Grove, PA, USA).…”

Inability to Induce Tolerance Through Direct Antigen Presentation

“…The B7-1/2-/-mutation was screened using NEO sense and antisense primers in addition to the following primers: B7-1, 5ƒ-CTGTCCAAGTCAGTGAAAGAT-3ƒ (sense) and 5ƒ-GGACAACTTTAC-TAAAGCCA-3ƒ (antisense); and B7-2, 5ƒ-TATTTCAATGGGACTGCATATC-3ƒ (sense) and 5ƒ-CGATCACTGACAGTTCTGTTA-3ƒ (antisense) (Integrated DNA Technologies Inc). (52) were prepared in our laboratory. Low-toxicity rabbit complement was purchased from PelFreeze (Brown Deer, Wisconsin, WI).…”

Therapeutic implications of NK cell regulation of allogeneic CD8 T cell-mediated immune responses stimulated through the direct pathway of antigen presentation in transplantation

“…Fc receptors were blocked by pre-incubating cell suspensions with 2 μg/ml of blocking anti-FcγR mAb (2.4G2) was produced and purified in house and used to reduce nonspecific binding before adding the specified biotin- or fluorochrome-labeled mAbs. 53 Biotinylated antibodies were detected with SA-Brilliant Violet 421. Dead cells and debris were excluded from acquisition by propidium iodide (PI).…”