Mechanistic Insights Into Efficient Intracellular Delivery via a High‐Precision Piezoelectric‐Nanoneedle Microfluidic Platform

ABSTRACT Intracellular delivery is trending toward high‐throughput, efficient, and universal approaches, crucial for large‐scale applications such as biologics production and cell therapy. Combining nanoneedles with external vibrations holds potential for efficient delivery. However, an integrated system is urgently required to achieve precise control over vibrations and microfluids, and the mechanisms of cell‐nanoneedle contact processes remain unclear. In this work, we developed a nanoneedle‐based delivery platform, containing a miniaturized piezoelectric vibrator and patterned nanoneedle‐microfluidic chips (PN‐chip). The vibrator provided customized outputs and a user‐friendly interface for laboratory use. The PN‐chips were fabricated using pattern‐bonded OSTE (off‐stoichiometry thiol‐ene) microchannels and an improved self‐assembly hybrid etching method to ensure stable microchannel heights and effective cell membrane penetrations. Using this platform, we systematically investigated the effects of microchannel height, nanoneedle morphology, and vibration parameters on delivery outcomes. We further proposed a confined‐particle‐flow vibration model to analyze cell vibration dynamics and nanoneedle contact events, identifying an empirical cell–nanoneedle contact stress range for effective delivery. The optimized parameters achieved FITC‐dextran delivery rates of 83.1%∼94.6% in HFB, Jurkat, and K562 cells. The integrated vibration system and the rapid fabrication of the PN‐chip offered potential for a universal, high‐throughput delivery methodology.