Monoclonal antibody FC-5.01, directed against CD63 antigen, is internalized into cytoplasmic vesicles in the IIB-BR-G human breast cancer cell line.
Monoclonal antibody (MAb) FC-5.01, raised against the undifferentiated breast cancer cell line IIB-BR-G, has been recently shown to react with CD63. The antigen (Ag) recognized by MAb FC-5.01 is expressed in plasma membranes of IIB-BR-G and other neoplastic cells, as well as in activated platelets and endothelial cells, as detected by indirect immunofluorescence performed at 4 degrees C on live cells. In permeabilized cells, MAb FC-5.01 colocalizes with acridine orange in acidic vesicles (lysosomal/endosomal compartment). Scatchard plot analysis performed on IB-BR-G cells demonstrated a 1.4+/-0.4 x 10(7) M(-1) affinity constant and 2.1 x 10(6) antigenic sites per cell. MAb FC-5.01 is not able to mediate C fixation or ADCC toward CD63+ cells, but the FC-5.01-CD63 complex is efficiently internalized into cytoplasmic vesicles, as shown by an acid wash immunofluorescence assay. Cellular catabolism of the antibody bound by IIB-BR-G cells was studied using [125I]-FC-5.01. At 18 h, >70% of the radioactivity was present in the supernatant as degraded fragments (TCA-soluble). After internalization, rapid Ag re-expression could be demonstrated in IIB-BR-G cells. MAb FC-5.01 diminished migration of CD63+ cells in a Boyden chamber assay. Some of the above-mentioned properties would enable the use of MAb FC-5.01 as a vehicle to target different compounds inside CD63+ cells.
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Instituto de Investigaciones Bioquímicas Fundación Campomar, Capital Federal, Argentina.
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Mordoh
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D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000648:ultrastructure*; D000937:Antigen-Antibody Reactions; D015703:Antigens, CD / Q000276:immunology*; D001692:Biological Transport / Q000276:immunology; D001943:Breast Neoplasms / Q000276:immunology* / Q000473:pathology; D003594:Cytoplasmic Granules / Q000276:immunology*; D005260:Female; D006801:Humans; D010980:Platelet Membrane Glycoproteins / Q000276:immunology*; D060149:Tetraspanin 30; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; C554586:CD63 protein, human; D010980:Platelet Membrane Glycoproteins; D060149:Tetraspanin 30