Early-stage antibody discovery and engineering typically require the cloning, expression, and screening of large numbers of proteins. Normally, DNA fragments encoding proteins of interest are cloned into extra-chromosomal plasmids that are amplified in Escherichia coli. Following purification from the bacteria, the plasmids are introduced into appropriate cells, and the expressed recombinant proteins screened for desired binding or function in a high-throughput manner. Even in a 96-well plate format, plasmid purification from E. coli is typically a labor intensive and time-consuming process. To further accelerate our existing biotherapeutic discovery workflows we designed, qualified, and enabled a fully integrated high-throughput plasmid purification and quantification workstation which we have termed AMPS (Automated Miniprep Plasmid Station). Using components from a commercially available kit, AMPS can purify plasmid preparations from twenty 96-deep-well plates of E. coli cultures, measure DNA absorbance at 260 nm, calculate plasmid concentrations, and prepare 96-deep-well plates for mammalian expression in an operator-independent manner. Plasmid yields and concentrations are equivalent to those obtained off-line. Furthermore, the quality of the DNA purified on the AMPS is equivalent to that obtained off-line in terms of DNA topology, and absence of contaminating bacterial chromosomal DNA and RNA. Most importantly, plasmids purified on the AMPS provide similar antibody titers following transfection in CHO cells as plasmids purified off-line. The AMPS bridges high-throughput E. coli colony picking capabilities typically available in an automation lab with downstream CHO expression needs and will facilitate screening of large numbers of biotherapeutics in binding and cell assay screens.

中文翻译：

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用于产生哺乳动物细胞表达质量 DNA 的全自动高通量质粒纯化工作站。

早期抗体发现和工程通常需要克隆、表达和筛选大量蛋白质。通常，编码感兴趣的蛋白质的 DNA 片段被克隆到在大肠杆菌中扩增的染色体外质粒中。在从细菌中纯化后，将质粒导入适当的细胞中，并以高通量方式筛选表达的重组蛋白以获得所需的结合或功能。即使在 96 孔板中，从大肠杆菌中纯化质粒通常也是一个劳动密集型且耗时的过程。为了进一步加快我们现有的生物治疗发现工作流程，我们设计、认证并启用了一个完全集成的高通量质粒纯化和定量工作站，我们称之为 AMPS（自动小量制备质粒工作站）。使用市售试剂盒中的组件，AMPS 可以从 20 个大肠杆菌培养物的 96 孔深孔板中纯化质粒制剂，测量 260 nm 处的 DNA 吸光度，计算质粒浓度，并制备用于哺乳动物表达的 96 孔深孔板一种独立于运营商的方式。质粒产量和浓度与离线获得的相同。此外，在 AMPS 上纯化的 DNA 在 DNA 拓扑结构方面与离线获得的质量相当，并且没有污染细菌染色体 DNA 和 RNA。最重要的是，在 AMPS 上纯化的质粒在转染 CHO 细胞后提供与离线纯化的质粒相似的抗体滴度。AMPS 桥接高通量 E.