Easy-to-Use Osmosis-Based Microfluidic Chip for Protein Crystallization: Application to a Monoclonal Antibody

Morais Sandy; Clisson Gerald; Mastropietro Teresa Fina; Briuglia Maria L.; ter Horst Joop H.; Leng Jacques; Salmon Jean-Baptiste

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Easy-to-Use Osmosis-Based Microfluidic Chip for Protein Crystallization: Application to a Monoclonal Antibody

机译：易于使用的基于渗透性的微流体芯片用于蛋白质结晶：应用于单克隆抗体

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We developed a disposable microfluidic chip which mimics the vapor diffusion method for exploring protein crystallization conditions but at the nanoliter scale. This device exploits the permeation of water through a thin poly(dimethylsiloxane) (PDMS) layer separating droplets stored in the chip and containing a mixture of proteins and precipitants from an open microfluidic reservoir. The water chemical activity fixed by the reservoir makes it possible to modify the volume of the droplets in a controlled way (reduction or increase). Because PDMS is only permeable to water, the imposed water activity therefore increases or decreases the concentration of the solutes present in the droplet and consequently the supersaturation of the solution. The specificity of our approach, in addition to the low volumes ensuring controlled mass transport conditions, is that the concentration of all the solutes is known at any time, thus allowing the extraction of quantitative information on the crystallization process. We exploit these chips on a protein of therapeutic interest: the full-length monoclonal antibody anti-CD20. Our experiments allow us in particular to estimate both the solubility of this protein and the width of the metastable zone with Na2SO4 and PEG400 as crystallizing agents. We also show that the fine-tuning of the permeation rate makes it possible to perform crystallization/dissolution cycles to selectively dissolve small crystals and increase the mean size of the remaining anti-CD20 crystals.

机译：我们开发了一种一次性微流控芯片，模拟蒸汽扩散法探索蛋白质结晶条件，但规模为纳升。该装置利用水通过薄聚二甲基硅氧烷（PDMS）层的渗透，将存储在芯片中的液滴分离，并包含来自开放微流控储存器的蛋白质和沉淀剂的混合物。储液罐固定的水化学活性使以受控方式（减少或增加）修改液滴体积成为可能。由于PDMS仅对水具有渗透性，因此施加的水活度会增加或降低液滴中存在的溶质浓度，从而增加或降低溶液的过饱和度。除了确保受控质量传输条件的低体积外，我们方法的特殊性在于，所有溶质的浓度在任何时候都是已知的，因此可以提取结晶过程的定量信息。我们利用这些芯片对一种具有治疗意义的蛋白质进行研究：全长单克隆抗体抗CD20。我们的实验特别允许我们用Na2SO4和PEG400作为结晶剂来估计这种蛋白质的溶解度和亚稳区的宽度。我们还表明，渗透速率的微调使执行结晶/溶解循环成为可能，以选择性地溶解小晶体，并增加剩余抗CD20晶体的平均尺寸。

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《Crystal growth & design》 |2021年第6期|共8页
作者
Morais Sandy; Clisson Gerald; Mastropietro Teresa Fina; Briuglia Maria L.; ter Horst Joop H.; Leng Jacques; Salmon Jean-Baptiste;
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作者单位

Univ Bordeaux LOF UMR 5258 CNRS Solvay F-33600 Pessac France;

Univ Bordeaux LOF UMR 5258 CNRS Solvay F-33600 Pessac France;

CNR Ist Tecnol Membrane I-87036 Arcavacata Di Rende Italy;

Univ Strathclyde EPSRC Ctr Innovat Mfg Continuous Mfg &

Crystallis Strathclyde Inst Pharm &

Biomed Sci Glasgow G1 1RD Lanark Scotland;

Univ Strathclyde EPSRC Ctr Innovat Mfg Continuous Mfg &

Crystallis Strathclyde Inst Pharm &

Biomed Sci Glasgow G1 1RD Lanark Scotland;

Univ Bordeaux LOF UMR 5258 CNRS Solvay F-33600 Pessac France;

Univ Bordeaux LOF UMR 5258 CNRS Solvay F-33600 Pessac France;

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正文语种 eng
中图分类晶体学;
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4. Evaluation of cell culture in microfluidic chips for application in monoclonal antibody production [J] . Penaherrera A., Payes C., Sierra-Rodero M., Microelectronic Engineering . 2016,第Juna期

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机译：具有和不具有结合半抗原的单克隆抗自旋标记抗体的Fab片段的结晶。

Easy-to-Use Osmosis-Based Microfluidic Chip for Protein Crystallization: Application to a Monoclonal Antibody

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