Rapid generation of specific antibodies by enhanced homologous recombination
Rapid generation of specific antibodies by enhanced homologous recombination
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DOI：
10.1038/nbt1092
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发表时间：
2005-06-01
影响因子：
46.9
通讯作者：
Ohta, K
中科院分区：
文献类型：
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作者：
Seo, H;Masuoka, M;Ohta, K
在鸡免疫系统中，基因转换，一种同源重组，主要有助于免疫球蛋白基因的多样化（1，2）。在这里，我们报告使用鸡DT 40 B细胞系进行基因转换的快速生成特异性单克隆抗体。我们发现，通过用组蛋白脱乙酰酶抑制剂，曲马斯他汀A（TSA）（3）处理DT 40细胞，免疫球蛋白基因座的基因转换频率增加，从而在大多数处理的细胞中产生免疫球蛋白基因座的多样性。这表明TSA处理加速了DT 40细胞上表面IgM的自主多样化。我们利用这种效应，选择DT 40细胞产生特异性抗体与抗原结合的磁珠。这种自主多样化文库（ADLib）选择系统能够快速建立（类似于从多样化文库开始1周）产生具有足够特异性和亲和力的单克隆IgM的各种克隆，用于免疫测定，并且适用于包括合理蛋白质设计的各种生物技术。
In the chicken immune system, gene conversion, a type of homologous recombination, primarily contributes to diversification of the immunoglobulin gene(1,2). Here, we report on the rapid generation of specific monoclonal antibodies using the chicken DT40 B-cell line undergoing gene conversion. We discovered that the gene conversion frequency at the immunoglobulin locus is increased by treating DT40 cells with a histone deacetylase inhibitor, trichostatin A (TSA)(3), thereby generating diversity at the immunoglobulin locus in the majority of treated cells. This indicates that TSA treatment accelerates the autonomous diversification of surface IgMs on DT40 cells. We took advantage of this effect to select DT40 cells producing specific antibodies with antigen-conjugated magnetic beads. This autonomously diversifying library (ADLib) selection system enables the quick establishment (similar to 1 week from a diversifying library) of various clones producing monoclonal IgMs with enough specificity and affinity for immunological assays, and is applicable to various biotechnologies including rational protein design.