Chimeric antibodies composed of mouse-derived variable regions and human-derived constant regions have been developed for clinical use. However, construction of chimeric mouse/human genes in expression vectors is time-consuming work. In this study, we developed convenient vectors for construction of chimeric mouse/human antibodies. The protocols are as follows: In mouse hybridomas and B cells, most active Vn and V, genes can be identified as rearranged bands by Southern hybridization of EcoRI-and HindHI-digested DNAs with Jn and J, probes, respectively, and such fragments can be isolated in I-EcoRI and I-Hind111 vectors, respectively. We constructed two plasmids: pSV2-HGlgpt contains human C;., and Ecogpt genes, and only one EcoRI site upstream of the C,, gene; pSV2-HC,neo contains human C, and neo genes, and only one Hind111 site upstream of the C, gene. An isolated EcoRI fragment containing a VuDnJu gene and a Hind111 fragment containing a V,J, gene are inserted into pSV2-HGlgpt and pSV2-HC,neo, respectively. Both resulting plasmid DNAs are co-transfected into SP2/0 cell, a non-Ig-secreting mouse myeloma. Transformants are selected by both mycophenolic acid and G418. With this procedure, it takes only 2 months to obtain chimeric antibodies.
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