Dehne Michaela, Neidinger Simon Valentin, Stark Michael, Adamo Antonia Camilla, Kraus Xenia, Färber Nicolas, Westerhausen Christoph, Bahnemann Janina
Institute of Technical Chemistry, Leibniz University Hannover, Hannover 30167, Germany.
Chair Technical Biology, Institute of Physics, University of Augsburg, Augsburg 86159, Germany.
ACS Omega. 2024 May 3;9(19):21637-21646. doi: 10.1021/acsomega.4c02590. eCollection 2024 May 14.
For the process of transient transfection (TTF), DNA is often transported into the cells using polyplexes. The polyplex uptake and the subsequent transient expression of the gene of interest are of great importance for a successful transfection. In this study, we investigated a 3D-printed microfluidic system designed to facilitate direct TTF for suspension of CHO-K1 cells. The results demonstrate that this system achieves significantly better results than the manual approach. Furthermore, the effect of both post-transfection incubation time (t) and temperature (T) on polyplex uptake was explored in light of the membrane phase transitions. Attention was paid to obtaining the highest possible transfection efficiency (TFE), viability (V), and viable cell concentration (VCC). Our results show that transfection output measured as product of VCC and TFE is optimal for = 1 h at = 22 °C. Moreover, post-transfection incubation at = 22 °C with short periods of increased at = 40 °C were observed to further increase the output. Finally, we found that around = 19 °C, the TFE increases strongly. This is the membrane phase transition of CHO-K1 cells, and those results therefore suggest a correlation between membrane order and permeability (and in turn, TFE).
在瞬时转染（TTF）过程中，DNA通常利用多聚体转运到细胞中。多聚体摄取以及随后目的基因的瞬时表达对于成功转染至关重要。在本研究中，我们研究了一种3D打印微流控系统，该系统旨在促进对CHO-K1细胞悬浮液的直接TTF。结果表明，该系统比手动方法取得了显著更好的结果。此外，根据膜相变探讨了转染后孵育时间（t）和温度（T）对多聚体摄取的影响。重点关注获得尽可能高的转染效率（TFE）、活力（V）和活细胞浓度（VCC）。我们的结果表明，以VCC和TFE的乘积衡量的转染产出在22℃下孵育1小时时最佳。此外，观察到在22℃下转染后孵育，短时间在40℃下温度升高可进一步提高产出。最后，我们发现约19℃时，TFE大幅增加。这是CHO-K1细胞的膜相变，因此这些结果表明膜有序性与通透性（进而与TFE）之间存在相关性。
