Adoptive cell therapy has emerged as a promising strategy to treat cancer. It relies on patient-derived cells, such as T cells and mesenchymal stromal cells (MSCs), which are genetically engineered to become better equipped to fight cancer cells. While ex vivo genetic modification of T cells and MSCs has traditionally been performed with viral vectors, they come with concerns about safety, sustainable production and high development costs. Electroporation is a non-viral alternative transfection technology, but can lead to significant gene expression changes, phenotypic alterations, and decreased therapeutic potency. Recently, photoporation with electrospun photothermal nanofibers (PEN photoporation) was demonstrated to provide a safer alternative with minimal impact on the cell’s functionality and phenotype. The technology makes use of photothermal nanofibers which, upon stimulation with laser light, can transiently permeabilize cells to allow gene-modifying effector molecules to enter the cells. Having been thoroughly demonstrated and validated in a research setting (TRL4), this project aims to bring the PEN photoporation technology to TLR6 by developing hard- and software for automated high-throughput transfections of T cells (>1B cells/h) and MSCs (>10M cells/h). The technology will be extensively tested and validated in the cGMP compliant laboratories of the project partners for the genetic engineering of T cells and MSCs. At the same time, steps will be taken to prepare for commercialization and market deployment. By the end of the project a fully automated and validated high-throughput prototype system will be available for installation at centralized cell production facilities or ready for integration in point-of-care cell manufacturing equipment. This project aligns with the Micro-Nano-Bio challenge as it combines nanotechnology with microfluidics to enhance genetically engineered cell therapy products.