Intracellular Delivery Techniques for Biomolecular Applications

Intracellular delivery underpins advances in gene therapy, cell engineering, drug screening and molecular diagnostics. Techniques range from viral vectors, which rely on natural infection pathways, to non-viral methods that employ physical, chemical or mechanical means to breach the plasma membrane. Physical methods include electroporation, where electric pulses induce transient pores; photoporation, which uses laser energy to disrupt the membrane locally; and microinjection, in which fine glass needles deposit cargo directly into the cytosol. Mechanical strategies such as cell squeezing generate transient membrane openings through constrictive microchannels, preserving cell function in many cases. Nanostructured tools—nanoneedles, nanotubes and hollow nanoelectrodes—offer subcellular precision and can modulate endocytic pathways to enhance uptake. Together, these approaches enable delivery of nucleic acids, proteins, nanoparticles and small molecules into a broad range of cell types, balancing efficiency, throughput and viability for research and clinical applications.

Research from Nature Portfolio

Recent foundational work has refined single-cell microinjection by automating a micropipette platform that pre-patterns cells in arrays and precisely controls injection volume via pressure and timing. This system delivers defined amounts of synthetic mRNA or plasmid DNA into individual cells, correlating the fluorescent output directly with the injected dose and enabling targeted genetic modification with minimal perturbation. A parallel advance employs microfluidic squeezing of B-cells through narrow channels to create transient pores and load whole proteins intracellularly. Antigen-loaded B-cells processed and presented tumour antigens to CD8+ T-cells both in vitro and in vivo, establishing mechano-poration as a scalable vaccine strategy. Complementing these, a soft-electroporation platform uses gold-coated hollow nanoelectrodes to achieve low-voltage, high-precision delivery to adherent cells. Cells form a tight seal around three-dimensional nanochannels, permitting controlled uptake of membrane-impermeable molecules while maintaining viability and enabling spatially selective dosing.

Intracellular Delivery Techniques for Biomolecular Applications publication trend

The graph below shows the total number of articles in intracellular delivery techniques for biomolecular applications across all publications each year (not limited to Nature Index journals).

Technical terms

Electroporation: Use of brief electrical pulses to induce transient membrane pores for cargo entry.

Microinjection: Direct delivery of molecules into the cytosol using a micrometre-scale glass pipette.

Mechano-poration (cell squeezing): Creation of transient membrane openings by forcing cells through narrow constrictions.

Photoporation: Localised membrane permeabilisation induced by focused laser light, often with sensitising nanoparticles.

Nanoneedle array: High-aspect-ratio nanostructures that interface with cell membranes to facilitate endocytic uptake and cargo delivery.

References

1. Engineering Efficient CAR‐T Cells via Electroactive Nanoinjection. Advanced Materials (2023).
2. Cell engineering with microfluidic squeezing preserves functionality of primary immune cells in vivo. Proceedings of the National Academy of Sciences of the United States of America (2018).
3. Laser-assisted photoporation: fundamentals, technological advances and applications. Advances in Physics X (2016).
4. Porous Silicon Nanoneedles Modulate Endocytosis to Deliver Biological Payloads. Advanced Materials (2019).
5. Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes. Scientific Reports (2016).
6. Advanced physical techniques for gene delivery based on membrane perforation. Drug Delivery (2018).
7. Microfluidic squeezing for intracellular antigen loading in polyclonal B-cells as cellular vaccines. Scientific Reports (2015).
8. Soft electroporation for delivering molecules into tightly adherent mammalian cells through 3D hollow nanoelectrodes. Scientific Reports (2017).