title: Tools for coproducing multiple proteins in mammalian cells.
abstract: Structural and functional studies of many mammalian systems are critically dependent on abundant supplies of recombinant multiprotein complexes. Mammalian cells are often the most ideal, if not the only suitable host for such experiments. This is due to their intrinsic capability to generate functional mammalian proteins. This advantage is frequently countered by problems with yields in expression, time required to generate overexpressing lines, and elevated costs. Coexpression of multiple proteins adds another level of complexity to these experiments, as cells need to be screened and selected for expression of suitable levels of each component. Here, we present an efficient fluorescence marking procedure for establishing stable cell lines that overexpress two proteins in coordination, and we validate the method in the production of recombinant monoclonal antibody Fab fragments. This procedure may readily be expanded to systems of greater complexity, comprising more than two components.
authors: Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, USA.
authors: Zahra
authors: Z
authors: Assur
authors: Wayne A
authors: WA
authors: Hendrickson
authors: Filippo
authors: F
authors: Mancia
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D018929:Cell Culture Techniques; D003001:Cloning, Molecular; D005434:Flow Cytometry / Q000379:methods*; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000032:analysis* / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D014162:Transfection
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D011994:Recombinant Proteins