Light-activated nanocomposite thin sheet for high throughput contactless biomolecular delivery into hard-to-transfect cells

High throughput intracellular delivery of biological macromolecules is crucial for cell engineering, gene expression, therapeutics, diagnostics, and clinical studies; however, most existing techniques are either contact-based or have throughput limitations. Herein, we report a light-activated, contactless, high throughput photoporation method for highly efficient and viable cell transfection of more than a million cells within a minute. We fabricated reduced graphene oxide (rGO) nanoflakes that was mixed with a polydimethylsiloxane (PDMS) nanocomposite thin sheet with an area of 3 cm² and a thickness of ∼600 μm. Upon infrared (980 nm) nanosecond pulse laser exposure, the rGO nanoflakes induced heat and created photothermal bubbles, leading to cell membrane deformation and biomolecular delivery. Using this platform, we achieved delivery of small to large size molecules, such as propidium iodide (PI) dye (668 Da), dextran (3000 Da), siRNA (20-24 bp), EGFP (6159 bp) and enzymes (465 kDa), in L929, N2a, and HeLa cells as well as in hard-to-transfect NiH3T3 and HuH7 cells. The best results were achieved for enzymes with ∼97% transfection efficiency and 98% cell viability in Huh7 cells. This highly efficient cargo delivery tool is simple and easy to use, and its dimensions can be varied according to the user requirements. Moreover, this safe and successful method has applicability in diagnostics and cell therapy.