A rapid screening method for cell lines producing singly-tagged recombinant proteins using the "TARGET tag" system

作者：Tabata Sanae; Nampo Maiko; Mihara Emiko; Tamura Kawakami Keiko; Fujii Ikuo; Takagi Junichi^*

来源：Journal of Proteomics, 2010, 73(9): 1777-1785.

DOI：10.1016/j.jprot.2010.05.012

• Affinity tag
• Monoclonal antibody
• Surface plasmon resonance
• Phage display
• Stable cell line
• Mammalian cell expression

Recombinant production of extracellular glycoproteins in stable mammalian cell lines is an ideal technique for obtaining a large quantity of high-quality proteins. In most cases, however, current methodologies do not allow for sufficiently rapid cell line development and protein purification. Here, we describe a 21-residue peptide tag (designated as TARGET tag) and its use for rapid stable cell line development and purification. The ability of the anti-tag antibody P20.1 to withstand repetitive regeneration cycles has enabled the development of a sensitive surface plasmon resonance-based screening format that requires only 20 l of cell culture supernatants. Direct and semi-quantitative screening at the 96-well culture scale eliminated the need for a second screening, re-cloning, or sorting, thereby minimizing culture pre-production time. Using this system, "high producer" cell lines were established in less than a month, and milligram quantities of target proteins could be purified with a standardized protocol.

• 出版日期2010-8-5