Design of transfections: Implementation of design of experiments for cell transfection fine tuning,Biotechnology and Bioengineering

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Design of transfections: Implementation of design of experiments for cell transfection fine tuning
Biotechnology and Bioengineering ( IF 3.5 ) Pub Date : 2021-08-18 , DOI: 10.1002/bit.27918
Sara Mancinelli ₁ , Andrea Turcato ₂ , Annamaria Kisslinger ₃ , Antonella Bongiovanni ₄ , Valeria Zazzu ₁ , Antonella Lanati ₂ , Giovanna Lucia Liguori ₁

Affiliation

Transfection is the process by which nucleic acids are introduced into eukaryotic cells. This is fundamental in basic research for studying gene function and modulation of gene expression as well as for many bioprocesses in the manufacturing of clinical-grade recombinant biologics from cells. Transfection efficiency is a critical parameter to increase biologics' productivity; the right protocol has to be identified to ensure high transfection efficiency and therefore high product yield. Design of experiments (DoE) is a mathematical method that has become a key tool in bioprocess development. Based on the DoE method, we developed an operational flow that we called “Design of Transfections” (DoT) for specific transfection modeling and identification of the optimal transfection conditions. As a proof of principle, we applied the DoT workflow to optimize a cell transfection chemical protocol for neural progenitors, using polyethyleneimine (PEI). We simultaneously varied key influencing factors, namely concentration and type of PEI, DNA concentration, and cell density. The transfection efficiency was measured by fluorescence imaging followed by automatic counting of the green fluorescent transfected cells. Taking advantage of the DoT workflow, we developed a new simple, efficient, and economically advantageous PEI transfection protocol through which we were able to obtain a transfection efficiency of 34%.

中文翻译：

转染设计：细胞转染微调实验设计的实施

转染是将核酸引入真核细胞的过程。这是研究基因功能和基因表达调节的基础研究以及从细胞制造临床级重组生物制剂的许多生物过程的基础。转染效率是提高生物制剂生产力的关键参数；必须确定正确的方案以确保高转染效率和高产率。实验设计 (DoE) 是一种数学方法，已成为生物工艺开发的关键工具。基于 DoE 方法，我们开发了一种称为“转染设计”(DoT) 的操作流程，用于特定转染建模和确定最佳转染条件。作为原则证明，我们应用 DoT 工作流程来优化神经祖细胞的细胞转染化学方案，使用聚乙烯亚胺 (PEI)。我们同时改变了关键影响因素，即 PEI 的浓度和类型、DNA 浓度和细胞密度。通过荧光成像测量转染效率，然后自动计数绿色荧光转染的细胞。利用 DoT 工作流程，我们开发了一种新的简单、高效且具有经济优势的 PEI 转染方案，通过该方案我们能够获得 34% 的转染效率。通过荧光成像测量转染效率，然后自动计数绿色荧光转染的细胞。利用 DoT 工作流程，我们开发了一种新的简单、高效且具有经济优势的 PEI 转染方案，通过该方案我们能够获得 34% 的转染效率。通过荧光成像测量转染效率，然后自动计数绿色荧光转染的细胞。利用 DoT 工作流程，我们开发了一种新的简单、高效且具有经济优势的 PEI 转染方案，通过该方案我们能够获得 34% 的转染效率。

更新日期：2021-10-13

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