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The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lym-phoma has led to new treatment options for this group of patients. Production of monoclonal antibodies by the traditionalhybridoma technology is costly, and the resulting murine antibodies often have the disadvantage of triggering humananti-mouse antibody (HAMA) response. Therefore recombinant Fab antibodies generated by the phage display technol-ogy can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cellsof BALB/c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the κ lightchains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successivelyinserted into the phagemid vector pComb3H-SS to construct an immunized Fab phage display library. The diversity ofthe constructed library was approximately 1.94×10~7. Following five rounds of biopanning, soluble Fab antibodies wereproduced from positive clones identified by ELISA. From eight positive clones, FabC06, FabC21, FabC43 and FabC59were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains (V_H) and variablelight domains (V_L) were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murinegermline genes respectively. Furthermore, reactivity with membrane proteins of the B cell lymphoma was demonstratedby immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for furtherstudy of B cell lymphoma and could also contribute to the improvement of disease therapy.
The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lym-phoma has led to new treatment options for this group of patients. Production of monoclonal antibodies by the traditional hybridoma technology is costly, and the resulting murine antibodies often have this disadvantage of triggering human anti-mouse antibody (HAMA) response. Thus recombinant Fab antibodies generated by the phage display technol- ogy can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cells of BALB / c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the κ light chains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successively inserted into the phagemid vector pComb3H-SS to construct an immunized The diversity of the constructed library was approximately 1.94 × 10 ~ 7. Following five rounds of From eight positive clones, FabC06, FabC21, FabC43 and FabC59were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains (V_H) and variablelight domains (V_L ), were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murine germline genes respectively. These, immunoglobulin molecules of the B cell lymphoma were demonstrated by immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for furtherstudy of B cell lymphoma and could also contribute to the improvement of disease therapy.