Convenient plasmid vectors for construction of chimeric mouse/human antibodies

Koh zoh Kameyama, Kenji Imai, Toshihiro Itoh, Masaru Taniguchi, Keiji Miura, Yoshikazu Kurosawa

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14 Citations (Scopus)

Abstract

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 VH and Vκ{script} genes can be identified as rearranged bands by Southern hybridization of EcoRI- and HindIII-digested DNAs with JH and Jκ{script} probes, respectively, and such fragments can be isolated in λ-EcoRI and λ-HindIII vectors, respectively. We constructed two plasmids: pSV2-HG1gpt contains human Cγ1 and Ecogpt genes, and only one EcoRI site upstream of the Cγ1 gene; pSV2-HCκ{script}neo contains human Cκ{script} and neo genes, and only one HindIII site upstream of the Cκ{script} gene. An isolated EcoRI fragment containing a VHDHJH gene and a HindIII fragment containing a Vκ{script}Jκ{script} gene are inserted into pSV2-HG1gpt and pSV2-HCκ{script}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.

Original languageEnglish
Pages (from-to)301-306
Number of pages6
JournalFEBS Letters
Volume244
Issue number2
DOIs
Publication statusPublished - 27-02-1989

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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