Materials for Improving Immune Cell Transfection

Purpose: Immune-cell engineering is central to cell therapies, but primary immune cells are difficult to transfect without compromising viability, phenotype or function. Viral vectors offer strong expression but raise concerns about manufacturing, immunogenicity and insertional risk, while bulk electroporation can cause substantial cell stress. The review surveys material-enabled alternatives that could make immune-cell transfection more precise and scalable.

Hypothesis: No formal testable hypothesis is proposed. The central premise is that combining material design with controlled physical delivery can overcome the efficiency, viability and scale limitations of conventional immune-cell transfection.

Aims: - Compare viral vectors and non-viral transfection approaches for immune-cell engineering. - Review microfluidic, nanoparticle and high-aspect-ratio nanostructure platforms. - Assess delivery of DNA, mRNA, siRNA, proteins and CRISPR components into T cells, NK cells, dendritic cells and other immune cells. - Identify the performance, manufacturing and regulatory barriers that remain before clinical translation.

Delivery system: The reviewed platforms include lentiviral and retroviral vectors; bulk electroporation; microfluidic cell squeezing, micro-electroporation, acoustic and nanopore systems; lipid, polymeric and inorganic nanoparticles; and silicon nanowires, nanostraws, magnetic nanospikes and three-dimensional nanochannels. Payloads include plasmid DNA, mRNA, siRNA, proteins and CRISPR/Cas9 ribonucleoproteins.

Approach: This is a narrative review rather than a primary experimental study. It synthesises published work across primary T cells, NK cells, dendritic cells, macrophages and model cell lines, and discusses both ex vivo manufacturing and in vivo delivery considerations.

Key methods: The cited studies use flow cytometry, fluorescence and confocal microscopy, viability and proliferation assays, cytokine and cytotoxicity measurements, gene-expression and sequencing analyses, genome-editing readouts, biodistribution studies and animal models.

Key results: - Viral vectors can achieve high expression but require complex manufacturing and have limited cargo flexibility. - Bulk electroporation can deliver mRNA and genome-editing cargo efficiently, but electrical stress, Joule heating and high cargo exposure can reduce viability and alter cell state. - Microfluidic and nano-electroporation methods reduce reagent use and can improve control over transient membrane permeabilisation, although primary-cell performance and throughput remain platform-dependent. - Lipid and polymer nanoparticles have enabled delivery of siRNA, mRNA and genome-editing components to immune cells, but intracellular release, toxicity and reproducible formulation remain challenges. - High-aspect-ratio structures can provide high viability and efficient cytosolic access by creating temporary membrane interfaces, but fabrication, throughput and device integration complicate scale-up.

Interpretation: The authors argue that no single platform satisfies all requirements for immune-cell engineering. The most promising direction is to match material chemistry and physical delivery conditions to the cell type, payload and intended duration of expression, while measuring both transfection and preserved cellular function. Translation will depend on reproducible, closed and scalable processes rather than peak efficiency alone.

Limitations: - The review compares heterogeneous studies that use different cell sources, payloads, assays and definitions of efficiency. - Many results are obtained in model cell lines or small-scale ex vivo experiments rather than clinical manufacturing workflows. - Long-term effects on immune phenotype, proliferation and function are not consistently measured. - Nanoparticle toxicity, device clogging, fabrication cost, process control and GMP compatibility remain incompletely resolved. - Most approaches still require optimisation for each cell type and cargo, limiting general-purpose use.