Microdose jet injection reveals a tissue-dependent retention-expression window for carrier-free nucleic acid delivery

Efficient local nucleic acid delivery requires not only access to target cells but also retention of the administered payload. Yet whether increasing physical delivery input improves or compromises functional delivery remains poorly understood. Here, we show that microdose jet injection operates within a tissue-dependent retention-expression window that can be interpreted in terms of physical jet input. Using the Actranza Lab needle-free system, sub-microlitre to low-microlitre jet outputs were calibrated and characterised by high-speed imaging, defining a kinetic-energy range of approximately 0.38-18.5 mJ. Delivery outcomes were evaluated in mouse spleen, liver, and skin using dye-retention analysis and luciferase expression. In spleen and liver, low-volume jets were associated with local retention and high volume-normalised luciferase expression, whereas increasing jet input progressively increased retention failure and reduced expression efficiency despite larger administered DNA amounts. In contrast, skin maintained local retention over a substantially broader input range, and total luciferase expression showed a significant positive association with injection volume. At the same 1-µL splenic administration volume, Actranza Lab produced approximately 34-fold greater luciferase activity than a 30-G Hamilton microsyringe. Cross-tissue integration of retention-failure frequency and volume-normalised expression revealed narrower operational windows in spleen and liver and a broader window in skin. These findings show that successful carrier-free jet delivery is determined not by nominal dose alone, but by the interaction between physical jet input and tissue retention. This retention-expression framework provides a practical basis for tissue-specific optimisation of needle-free nucleic acid delivery.