柠檬酸铁铵对悬浮 HEK293 细胞转染的影响机制探究.

Suspended HEK293（human embryonic kidney）cell transfection based on PEI（polyethyleneimine）is a prospective technology. However, the components of the medium often affect the transfection efficiency, which needs to be changed with transfection medium by centrifugation. This process is cumbersome and not only has the risk of contamination, but also affects the cell state. In order to fundamentally solve this problem, the influence mechanism of key components that inhibit the efficiency of transfection on the transition process need to be explored. First we explored that the key composition that have potential effects on transfection efficiency in the medium is ferric ammonium citrate（FAC）, and determined the its inhibitory effect. Then we analyzed its mechanism for inhibiting transit efficiency, including the effects of ferric ammonium citrate on the cell state, PEI-DNA complex formation, and destination gene expression. Results showed that certain amount of FAC had significant inhibitory effect on the transfection that was also gradually enhanced with concentration increasing. Further studies found that the transfection was inhibited only when citrate and iron ions were present at the same time. Over high concentration of FAC increased the particle size of the PEI-DNA complex, making it more difficult to enter the cell. This led to a reduction in the number of complexes entering the cells, and ultimately resulted in a significant drop in cell transfection efficiency. FAC restricted DNA into cells by affecting the size of the PEI-DNA complex, thereby inhibiting PEI-mediated suspended HEK293 cell transfusion process. The inhibitory effect can be eliminated when the FAC concentration was < 20 μmol/L. Through this study, the effect of FAC on the transfection process of PEI-mediated suspended HEK293 cells and its mechanism were deeply understood, providing a solid reference for the development and rational design of the transfection medium for suspended HEK293 cells. [ABSTRACT FROM AUTHOR]