ABSTRACT Intracellular delivery into suspension cells, particularly hard‐to‐transfect immune cells such as T‐ and B‐lymphocytes, remains challenging. Membrane disruption‐based microfluidic methods offer a carrier‐free alternative but often depend on high‐viscosity buffers, compromising viability and scalability. Here, we introduce a viscoelastic mechanoporation platform using a hyperbolic microfluidic channel and low‐viscosity λDNA buffer for the efficient delivery of mRNA and small molecules. The system harnesses extensional strain to transiently deform cell membranes, enabling high‐throughput cytosolic uptake with minimal cellular stress. Our platform achieved up to ∼17‐fold enhanced mRNA delivery while maintaining >85% viability across multiple suspension cell lines. Mechanistic insights from Laurdan spectral analysis, ice incubation, and metabolic profiling revealed how membrane dynamics govern delivery outcomes. We further modulated efficiency through osmotic and cytoskeletal perturbations, demonstrating a tunable strategy for safe and effective delivery into fragile immune cells.
Park et al. (Sat,) studied this question.