用于抗体工程的哺乳动物细胞展示
Mammalian cell display for antibody engineering.
作者信息
Ho Mitchell, Pastan Ira
机构信息
National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
出版信息
Methods Mol Biol. 2009;525:337-52, xiv. doi: 10.1007/978-1-59745-554-1_18.
Abstract
Antibody engineering has generally been carried out by displaying mouse or human antibodies or antibody fragments on the surface of microorganisms (phage, bacteria, and yeast). We have shown that mammalian cells can be used to display single-chain antibody fragments (scFvs) for affinity maturation. Using mammalian cell display one can isolate and engineer scFvs, Fabs, or whole IgGs for increased affinity and other specific biological functions. Here, we describe a mammalian cell display strategy to isolate high-affinity scFvs specific for CD22. Our strategy uses flow cytometry and human embryonic kidney 293T (HEK-293T) cells that are widely used for transient protein expression. Flow cytometry enhances the screen's sensitivity thereby allowing us to isolate high-affinity antibodies.
摘要
抗体工程通常是通过在微生物（噬菌体、细菌和酵母）表面展示小鼠或人抗体或抗体片段来进行的。我们已经表明，哺乳动物细胞可用于展示单链抗体片段（scFv）以进行亲和力成熟。利用哺乳动物细胞展示技术，可以分离和改造scFv、Fab或完整的IgG，以提高亲和力和其他特定的生物学功能。在此，我们描述一种用于分离针对CD22的高亲和力scFv的哺乳动物细胞展示策略。我们的策略使用流式细胞术和广泛用于瞬时蛋白表达的人胚肾293T（HEK-293T）细胞。流式细胞术提高了筛选的灵敏度，从而使我们能够分离出高亲和力抗体。
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