Intracellular Photothermal Delivery for Suspension Cells Using Sharp Nanoscale Tips in Microwells.

Efficient intracellular delivery of biomolecules into cells that grow in suspension is of great interest for biomedical research, such as for applications in cancer immunotherapy. Although tremendous effort has been expended, it remains challenging for existing transfer platforms to deliver materials efficiently into suspension cells. Here, we demonstrate a high-efficiency photothermal delivery approach for suspension cells using sharp nanoscale metal-coated tips positioned at the edge of microwells, which provide controllable membrane disruption for each cell in an array. Self-aligned microfabrication generates a uniform microwell array with three-dimensional nanoscale metallic sharp-tip structures. Suspension cells self-position by gravity within each microwell in direct contact with 8 sharp tips, where laser-induced cavitation bubbles generate transient pores in the cell membrane to facilitate intracellular delivery of extracellular cargo. A range of cargo sizes were tested on this platform using Ramos suspension B cells with an efficiency >84% for Calcein green (0.6 kDa) and >45% for FITC-dextran (2,000 kDa), with retained viability >96%, and a throughput >100,000 cells delivered per minute. The bacterial enzyme -lactamase (29 kDa) was delivered into Ramos B cells and retained its biological activity, whereas a green fluorescence protein (GFP) expression plasmid was delivered into Ramos B cells with a transfection efficiency of >58% and viability of >89% achieved.