title: Expression vectors for human-mouse chimeric antibodies.
abstract: The production of recombinant antibodies has been generally recognized as time-consuming and labor-intensive. The aim of our study is to construct mammalian expression vectors containing the cDNA encoding the human constant regions and murine variable regions to massively and cost-effectively produce full-length chimeric antibodies. Unique restriction sites flanking the Ig variable region were designed to allow for the replacement of variable regions generated by PCR. Western blot analysis of the chimeric antibodies revealed that the expressed products were of the predicted size, structure and specificity. The usefulness of the vectors was confirmed by construction of human-mouse chimeric antibody-HCAb which secretes murine antibody against the human colorectal cancer. Selected in medium containing gradually increasing methotrexate (MTX), clones with increased expression of the product gene can be efficiently generated. The secretion of recombinant chimeric antibody-HCAb yielded 30 pg cell(-1) day(-1) at 10(-6 )M MTX. With this high-level expression from pools, the convenient and rapid production of over 100 milligram amounts per liter of recombinant antibodies may be achieved, which indicates the significant roles of pYR-GCEVH and pYR-GCEVL in the production of chimeric antibodies.
authors.affiliation: Cell Engineering Research Center, State Key Laboratory of Cancer Biology, Fourth Military Medical University, Xi'an City 710032, China.
authors.forename: Hua
authors.initials: H
authors.lastname: Xiong
authors.forename: Yuliang
authors.initials: Y
authors.lastname: Ran
authors.forename: Jinliang
authors.initials: J
authors.lastname: Xing
authors.forename: Xiangmin
authors.initials: X
authors.lastname: Yang
authors.forename: Yu
authors.initials: Y
authors.lastname: Li
authors.forename: Zhinan
authors.initials: Z
authors.lastname: Chen
mesh_terms: D000818:Animals; D000912:Antibodies, Neoplasm / Q000235:genetics* / Q000276:immunology; D016466:CHO Cells; D015179:Colorectal Neoplasms / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006224:Cricetinae; D005822:Genetic Vectors; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000378:metabolism; D008727:Methotrexate / Q000494:pharmacology; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: D000912:Antibodies, Neoplasm; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D008727:Methotrexate