Abstract Intracellular drug delivery is at the heart of many diagnosis procedures and a key step in gene therapy. Research has been conducted to bypass cell barriers for controlled intracellular drug release and made consistent progress. However, state-of-the-art techniques based on non-viral carriers or physical methods suffer several drawbacks, including limited delivery yield, low throughput or low viability, which are key parameters in therapeutics, diagnostics and drug delivery. Nevertheless, gold nanoparticle (AuNP) mediated photoporation has stood out as a promising approach to permeabilize cell membranes through laser induced Vapour NanoBubble (VNB) generation, allowing the influx of external cargo molecules into cells. However, its use as a transfection technology for the genetic manipulation of therapeutic cells is hindered by the presence of non-degradable gold nanoparticles. Here, we report a new optofluidic method bringing gold nanoparticles in close proximity to cells for photoporation, while avoiding direct contact with cells by taking advantage of hydrodynamic focusing in a multi-flow device. Cells were successfully photoporated with $$\sim {70}{\%}$$ ∼ 70 % efficiency with no significant reduction in cell viability at a throughput ranging from $$10^3$$ 10 3 to $$10^4~\text {cells}~{\hbox {min}^{-1}}$$ 10 4 cells min - 1 . This optofluidic approach provides prospects of translating photoporation from an R &D setting to clinical use for producing genetically engineered therapeutic cells.