Efficient method for production of high yields of Fab fragments in Drosophila S2 cells
Efficient method for production of high yields of Fab fragments in Drosophila S2 cells
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DOI：
10.1093/protein/gzp088
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发表时间：
2010-04-01
影响因子：
2.4
通讯作者：
Rey, Felix A.
中科院分区：
文献类型：
--
作者：
Backovic, Marija;Johansson, Daniel X.;Rey, Felix A.
Fab分子被用作治疗剂，并且是结构生物学中极有价值的工具。我们在此报道一种在果蝇S2细胞中生产用于结构生物学的重组Fab的方法。在数周内创建了稳定转染表达Fab的S2细胞系。重组Fab被分泌出来，经过亲和层析和尺寸排阻色谱后，从一升细胞培养物中获得了16毫克纯蛋白。该Fab具有功能，并与其同源抗原形成复合物，这通过共沉淀和尺寸排阻色谱得以证明。生化特性表明，来自S2细胞的Fab比通过消化杂交瘤细胞产生的抗体所获得的Fab糖基化程度更低，这一特征对于结晶形成可能是有利的。综上所述，与单克隆抗体的酶消化相比，从S2细胞获得重组Fab是一种更快且成本效益高得多的方法。
Fab molecules are used as therapeutic agents, and are invaluable tools in structural biology. We report here a method for production of recombinant Fab in Drosophila S2 cells for use in structural biology. Stably transfected S2 cell lines expressing the Fab were created within weeks. The recombinant Fab was secreted, and after affinity and size exclusion chromatography, 16 mg of pure protein were obtained from a liter of cell culture. The Fab was functional and formed a complex with its cognate antigen as demonstrated by co-precipitation and size exclusion chromatography. Biochemical characterization indicated that the Fab from S2 cells is less extensively glycosylated than the Fab obtained by digestion of antibody produced in hybridoma cells, a feature that may be advantageous for the purposes of crystallogenesis. Taken together, obtaining recombinant Fab from the S2 cells has been a faster and considerably more cost-effective method compared with the enzymatic digestion of the monoclonal antibody.