Antibody affinity maturation through combining display of two-chain paired antibody and precision flow cytometric sorting
文献类型：期刊论文
| 作者 | Sun, Shuang1,2; Yang, Xiao3; Wang, Haifeng4; Zhao, Yun1; Lin, Yan1; Ye, Chen1; Fang, Xiangdong3; Hang, Haiying1,5 |
| 刊名 | Applied microbiology and biotechnology
 |
| 出版日期 | 2016-07-01 |
| 卷号 | 100期号:13页码:5977-5988 |
| 关键词 | Coiled coiled heterodimeric variable domain (ccfv) Anchored periplasmic expression (apex) Flow cytometry Affinity maturation Precision sorting |
| ISSN号 | 0175-7598 |
| DOI | 10.1007/s00253-016-7472-1 |
| 通讯作者 | Hang, haiying(hh91@ibp.ac.cn) |
| 英文摘要 | Recombination of antibody light and heavy chain libraries greatly increases the size of a two-chain paired antibody library, thus easing the construction of large antibody libraries. here, light and heavy chain variable domains paired by a coiled coil were applied to a bacterial inner membrane display system. however, the probability of the correct pairing of light and heavy chains through random recombination after each round of flow cytometric sorting and cloning was very low in the presence of mostly unmatched light and heavy chain genes, resulting in inefficient enrichment; a target antibody clone in the ratio of 1:100,000 negative control spheroplasts was unable to be enriched by six rounds of sorting and cloning by a conventional sorting strategy (sorting the top 1 %). by just sorting the top 0.000025 % of spheroplasts, we succeeded in enriching the target antibody clone mixed with negative control spheroplasts in a ratio of 1:10(8) by just one round of sorting and cloning. furthermore, using this gating strategy, we efficiently enriched for an antibody clone with an affinity slightly better than the parent antibody clone from mixed spheroplasts which were present in the ratio of 1 better affinity clone to 10 parent clones to 10(6) negative control clones after just two rounds of sorting and cloning, suggesting that this gating strategy is highly sensitive in distinguishing between clones with a small difference in affinity and also enriching for clones with a higher affinity. taken together, the combination of the display of a two-chain paired antibody library and the use of stringent gating has significantly increased the efficiency of the antibody maturation system. |
| WOS关键词 | FUNCTIONAL GABA(B) RECEPTOR ; YEAST-CELL-SURFACE ; ESCHERICHIA-COLI ; RIBOSOME DISPLAY ; MAMMALIAN-CELLS ; SCFV ANTIBODIES ; LIBRARY ; SELECTION ; DNA ; ELECTROTRANSFORMATION |
| WOS研究方向 | Biotechnology & Applied Microbiology |
| WOS类目 | Biotechnology & Applied Microbiology |
| 语种 | 英语 |
| WOS记录号 | WOS:000378725700027 |
| 出版者 | SPRINGER |
| URI标识 | http://www.irgrid.ac.cn/handle/1471x/2374639 |
| 专题 | 中国科学院大学 |
| 通讯作者 | Hang, Haiying |
| 作者单位 | 1.Chinese Acad Sci, Inst Biophys, Key Lab Prot & Peptide Pharmaceut, Natl Lab Biomacromol, Beijing 100101, Peoples R China 2.Univ Chinese Acad Sci, Beijing 100039, Peoples R China 3.Chinese Acad Sci, Beijing Inst Genom, CAS Key Lab Genome Sci & Informat, Beijing 100101, Peoples R China 4.North China Univ Sci & Technol, Coll Life Sci, Tangshan 063000, Hebei Province, Peoples R China 5.Chinese Acad Sci, Inst Biophys, Beijing 100101, Peoples R China |
| 推荐引用方式 GB/T 7714 | Sun, Shuang,Yang, Xiao,Wang, Haifeng,et al. Antibody affinity maturation through combining display of two-chain paired antibody and precision flow cytometric sorting[J]. Applied microbiology and biotechnology,2016,100(13):5977-5988. |
| APA | Sun, Shuang.,Yang, Xiao.,Wang, Haifeng.,Zhao, Yun.,Lin, Yan.,...&Hang, Haiying.(2016).Antibody affinity maturation through combining display of two-chain paired antibody and precision flow cytometric sorting.Applied microbiology and biotechnology,100(13),5977-5988. |
| MLA | Sun, Shuang,et al."Antibody affinity maturation through combining display of two-chain paired antibody and precision flow cytometric sorting".Applied microbiology and biotechnology 100.13(2016):5977-5988. |
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来源：中国科学院大学
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