Abstract
Intracellular delivery is a critical process in biology and medicine. During intracellular delivery, different constructs (e.g., functional macromolecules such as DNA, RNA, and protein, and various complexes) are delivered across the cell membrane and into the cytosol. Herein, we use a microfluidic post array to induce hydrodynamic conditions for cell membrane poration with microfluidic vortex shedding (µVS). μVS is then used for the intracellular delivery of mRNA to primary human pan T cells. The specific microfluidic device and experimental rig used in this study contains a 960 µm wide by 40 µm deep flow cell capable of processing more than 2 x 106 cells s-1 at volumes ranging from 100 µL to 1.5 mL. Furthermore, we demonstrate efficient enhanced green fluorescent protein (EGFP) mRNA expression (e.g., 57.4 ± 6.8% of viable, recovery cells, mean ± stdev) after mRNA delivery to human pan T cells with high cell viability (e.g., 83.7 ± 0.7% of recovered cells) and high cell recovery (e.g., 96.3 ± 1.1% of processed cells), resulting in net yield of 46.3 ± 5.6% viable, recovered, and GFP expressing human pan T cells at mRNA concentrations of 80 µg ml-1. We also demonstrate µVS does not alter human pan T cell growth nor activation. These results demonstrated that µVS is a rapid intracellular delivery platform with promising potential for cytosolic delivery of mRNA to human primary T cells for (1) clinical applications, where larger volumes of cells are required and demonstrated value for (2) research applications, where rapid screening and minimal reagent consumption is preferable.