TY - JOUR AU - Raes, Laurens AU - Stremersch, Stephan AU - Fraire, Juan C. AU - Brans, Toon AU - Goetgeluk, Glenn AU - Munter, Stijn De AU - Hoecke, Lien Van AU - Verbeke, Rein AU - Hoeck, Jelter Van AU - Xiong, Ranhua AU - Saelens, Xavier AU - Vandekerckhove, Bart AU - Smedt, Stefaan De AU - Raemdonck, Koen AU - Braeckmans, Kevin PY - 2020/09/27 Y2 - 2024/03/28 TI - Intracellular Delivery of mRNA in Adherent and Suspension Cells by Vapor Nanobubble Photoporation JF - Nano-Micro Letters JA - Nano-Micro Lett VL - 12 IS - SE - Articles DO - 10.1007/s40820-020-00523-0 UR - https://nmlett.org/index.php/nml/article/view/130 SP - 185 AB - <p>Efficient and safe cell engineering by transfection of nucleic acids remains one of the long-standing hurdles for fundamental biomedical research and many new therapeutic applications, such as CAR T cell-based therapies. mRNA has recently gained increasing attention as a more safe and versatile alternative tool over viral- or DNA transposon-based approaches for the generation of adoptive T cells. However, limitations associated with existing nonviral mRNA delivery approaches hamper progress on genetic engineering of these hard-to-transfect immune cells. In this study, we demonstrate that gold nanoparticle-mediated vapor nanobubble (VNB) photoporation is a promising upcoming physical transfection method capable of delivering mRNA in both adherent and suspension cells. Initial transfection experiments on HeLa cells showed the importance of transfection buffer and cargo concentration, while the technology was furthermore shown to be effective for mRNA delivery in Jurkat T cells with transfection efficiencies up to 45%. Importantly, compared to electroporation, which is the reference technology for nonviral transfection of T cells, a fivefold increase in the number of transfected viable Jurkat T cells was observed. Altogether, our results point toward the use of VNB photoporation as a more gentle and efficient technology for intracellular mRNA delivery in adherent and suspension cells, with promising potential for the future engineering of cells in therapeutic and fundamental research applications.</p><p>Highlights:</p><p>1 Vapor nanobubble (VNB) photoporation represents a promising physical technique for mRNA transfection of adherent and suspension cells.<br>2 A multitude of parameters related to the VNB photoporation procedure were optimized to enable efficient mRNA transfection.<br>3 VNB photoporation was found to yield five times more living, transfected Jurkat T cells as compared to electroporation, i.e., currently the standard nonviral transfection technique for T cells.</p> ER -