Tuberc Respir Dis.  1984 Sep;31(3):83-90. 10.4046/trd.1984.31.3.83.

Production and Study on the Specificity of Monoclonal Antibodies to Pressate Extract Antigen of Mycobacterium tuberculosis

• KMID: 2528766
• DOI: http://doi.org/10.4046/trd.1984.31.3.83

Abstract

Pressate extract of M. tuberculosis was used as antigen for immunization of Balb/c mouse to obtain spleen cells producing antibodies to M. tuberculosis. Antibody producing spleen cells were prepared 3 days after the last . challenge and then fused with HGPRT deficient, nonsecreting mouse myeloma cell line, P3-X 63-Ag 8.653. Primary screening by enzyme linked immunosorbent assay(ELISA) was performed 2 weeks after fusion to determine antibody secreting hybridom cells in 96-we11 culture plates, and among 576 wells, 124 wells (21. 5%) were positive. These hybridom cells were cloned by limiting dilution method and 5 celllines (TB-1 , TB-2, TB-3, TB-4 and TB-5) , were expanded to obtain culture supernatants containing antibodies reacting to M. tuberculosis antigen. These 5 moüoclonal antibodies were tested for their specificity toM. tuberculosis extract antigen by antibody absorption tests. All showed below than 50% of absorption when mixed with each extract antigen from M. bovis, M. aviltm, and M. fortuitum and no reactivities on ELl SA plates coated with these three antigens. Antibodies produced by TB-2 and TB-3 cell lines were determined to be more specific to TE antigen than those from TB-l , TB-4, and TB-5 hybridom, cells, These hybridoma cells were injected into the peritoneal cavity of Balb/c mice piletreated with pristane to induce tumor. 7 days after injection of the hybridoma cells, the antibody titer of ascitic fluid were determined by ELISA. All showed 2 or 3 times higher optical density in ELISA than those of hybridoma cell culture fluid.