A study was performed to develop hybridomas secreting antibodies reactive to human T lymphocytes as unlimited sources of reagents for T-cell monitoring and therapy for acute allograft rejection epidode.
Splenic lymphocytes from Balb/c mice immunized against Jurkat cells were fused with plasmacytoma cell lines, P 3x 63 Ag 8. V 653 by the use of 50%(w/v) polyethyleneglycol. The cell mixture. after hybridization was distributed into 362 wells of the 96-well culture plates, and allowed to grow in HAT media at 3TC under 5 % CO, tension for 2 weeks to select hybrid cells. Among 362 wells, 63 wells (22.5%) of which were positive for antibody activity determined by enzyme-linked immunosorbent assay (ELISA) against Jurkat cells. Culture fluid from the wells with the hybrids secreting antibodies reactive to the Jurkat cell lines were checked for cross reactivity with SNUMC- 2 cells(pre-B cell lines) by ELISA.
The supernatant from 3 wells, which were identified as Jurkat cell specific, were selected for cell cloning. The hydridomas obtained as such were named TC28, TC42, and TC44, respectively.
The titers of monoclonal antibody, TC28 (IgG2a) in culture supernatant and ascitic fluid from Balb/ c mice inoculated with the hybridoma were 1: 128 and 1:130,000, respectively. Monoclonal antibody, TC28 reacted strongly to the established T-cell lines (Jurkat, Molt 4, RPMI 8402, and HSB - 2) and fresh peripheral T-cells which were characterized by rosetting with sheep red cells. The reactions were determined by ELISA, immunoperoxidase staining, and microcytotoxicity test. But monoclonal antibody, TC 28 did not react with B-cell lines (SNUMC-2, SNUMC-3, SNUMC-5, HMC-BM 4, and Laz 474), fresh peripheral B-cells, and K562 cells.
The reactivity’s of monoclonal antibodies, TC28 and OKT3 to peripheral monouclear cells of normal persons by the immunofluorescence technique showed close similarity between them.
Monoclonal antibody TC28, which exclusively reacted with human T-lymphocyte seemed to be more specific for the surface antigens of immature T-cells than those of mature peripheral T-cells.
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