## 4 Discussion and Conclusions
The developed device has the advantage of rapidly dissolving continuously injected blood cells by applying an electric field without any chemical reagents or fluid control. Its simple structure is expected to be easily integrated into other pre-treatment systems. However, certain limitations of this study must be addressed. Firstly, diluted blood is required, which drastically reduces the expected throughput of blood lysis. This is a common problem with many microfluidic systems that handle blood samples and needs to be solved in practical applications. For example, in applications that require the immediate use of collected blood, such as point-of-care devices, it will be necessary to combine a microfluidic mixing system that allows dilute solutions to be mixed inside the device or to develop a robust system that can use whole blood as-is. To overcome this limitation, future work will incorporate an on-chip blood dilution module upstream of the lysis region. Microfluidic passive mixers such as herringbone structures or serpentine channels have been extensively studied and can be readily integrated with our device without introducing active components or disturbing the constant flow rate, owing to the valve-free nature of our system [39; 40]. This integration would enable the direct handling of whole blood samples while maintaining the advantages of our platform, such as low-voltage operation and rapid lysis. Furthermore, the modular design of our device allows for straightforward coupling with additional pre-processing units, opening the possibility for a fully integrated blood preparation platform suitable for point-of-care applications. Additionally, we acknowledge that the integrity of intracellular biomolecules, such as nucleic acids and proteins, was not evaluated in the present study. Importantly, confirming the preservation of these biomolecules is essential for ensuring compatibility with down stream molecular diagnostic workflows, as ICP may induce ionic gradients and strong local electric fields that could affect their stability. Future studies will therefore include dedicated assays, such as RNA integrity analysis (e.g., electrophoresis and RT-PCR) and protein activity measurements, to assess both the structural and functional preservation of cell contents. Nevertheless, this proof-of-concept work establishes a foundation for non-chemical, low-voltage, high-speed lysis, which can be readily extended toward integrated diagnostic systems in future developments. Next, a continuous accumulation of blood cell lysates can cause clogging or lysate leakage at the outlet. Because the drag force and electrophoretic force are balanced in the concentrated region, an excessive number of particles can break the force balance and cause lysate leakage. If a stronger electric field is applied to balance the force, a vortex is generated owing to the stronger electric field, and the concentrated particles stick to the microchannel due to the strong impact. While the current study demonstrates high lysis efficiency under relatively low-voltage conditions (75 V), the long-term robustness of the system requires further investigation. In particular, biofouling and lysate accumulation near the Nafion membrane could potentially interfere with electric field formation and flow stability over extended operation. To partially address this, we conducted a supplementary experiment at a higher voltage condition (100 V at 1 \(\mu\)L/min), which resulted in the visible adhesion of blood cells near the Nafion region, as shown in Figure S2. This observation suggests that operating the device beyond the optimized conditions may exacerbate fouling and reduce device performance. Future work will involve a quantitative evaluation of membrane fouling and strategies to minimize surface adhesion, such as surface coatings or pulse field operation modes, to improve long-term usability.