Seo Hidetaka, Hashimoto Shu-ichi, Tsuchiya Kyoko, Lin Waka, Shibata Takehiko, Ohta Kunihiro
Chiome Bioscience Inc., RIKEN, 2-1 Hirosawa, Wako-shi, Saitama-ken 351-0198, Japan.
Nat Protoc. 2006;1(3):1502-6. doi: 10.1038/nprot.2006.248.
Here, we describe a protocol for using the ADLib (Autonomously Diversifying Library) system to rapidly generate specific monoclonal antibodies using DT40, a chicken B-cell line that undergoes constitutive gene conversion at both light- and heavy-chain immunoglobulin loci. We previously developed the ADLib system on the basis of our finding that gene conversion in DT40 cells was enhanced by treatment of the cells with a histone deacetylase inhibitor, trichostatin A (TSA). TSA treatment evolves a diversified library of DT40 cells (ADLib), in which each cell has different surface IgM specificity. Antigen-specific DT40 cells are selected from ADLib using antigen-conjugated magnetic beads, and their specificity can be examined by various immunological assays, using culture supernatant containing secreted IgM. The whole process from selection to screening can be completed in about 1 week. Thus, the ADLib system will accelerate biological studies, including drug discovery and design.
在此，我们描述了一种使用ADLib（自主多样化文库）系统的方案，该方案利用DT40（一种鸡B细胞系，其轻链和重链免疫球蛋白基因座均发生组成型基因转换）快速生成特异性单克隆抗体。我们之前基于以下发现开发了ADLib系统：用组蛋白脱乙酰酶抑制剂曲古抑菌素A（TSA）处理DT40细胞可增强其基因转换。TSA处理可产生一个DT40细胞的多样化文库（ADLib），其中每个细胞都具有不同的表面IgM特异性。使用抗原偶联磁珠从ADLib中筛选出抗原特异性DT40细胞，其特异性可通过各种免疫测定法，利用含有分泌型IgM的培养上清液进行检测。从筛选到筛选的整个过程大约可在1周内完成。因此，ADLib系统将加速包括药物发现和设计在内的生物学研究。
