Controllable manipulation of fluid flow is crucial for efficient particle separation, which is associated with plenty of biomedical and industrial applications. Microfluidic technologies have achieved promising progress in particle positioning depending on inertial force with or without the help of the Dean effect. Herein, we describe an inertial microfluidic system containing a spiral microchannel for various highly efficient particle separations. We demonstrated that Dean-like secondary flow can be regulated by geometric confinement in the microchannel. On the introduction of a library of micro-obstacles into the spiral microchannels, the resulting linear acceleration of secondary flow can be applied to remarkably enhance particle focusing in time and space. Further, multiple separating and sorting manipulations of particles including polymeric particles, circulating tumor cells, and blood cells, can be successfully accomplished in the dimension-confined spiral channels in a sheathless, high-throughput (typically 3 ml min⁻¹), long-term (at least 4 h), and highly-efficient (up to 99.8% focusing) manner. The methodological achievement pointing to ease-of-use, effective, and high-throughput particle manipulations is useful for both laboratory and commercial developments of microfluidic systems in life and material sciences.

The presence and quantity of rare cells in the bloodstream of cancer patients provide a potentially accessible source for the early detection of invasive cancer and for monitoring the treatment of advanced diseases. The separation of rare cells from peripheral blood, as a “virtual and real-time liquid biopsy”, is expected to replace conventional tissue biopsies of metastatic tumors for therapy guidance. However, technical obstacles, similar to looking for a needle in a haystack, have hindered the broad clinical utility of this method. In this study, we developed a multistage microfluidic device for continuous label-free separation and enrichment of rare cells from blood samples based on cell size and deformability. We successfully separated tumor cells (MCF-7 and HeLa cells) and leukemic (K562) cells spiked in diluted whole blood using a unique complementary combination of inertial microfluidics and steric hindrance in a microfluidic system. The processing parameters of the inertial focusing and steric hindrance regions were optimized to achieve high-throughput and high-efficiency separation, significant advantages compared with existing rare cell isolation technologies. The results from experiments with rare cells spiked in 1% hematocrit blood indicated >90% cell recovery at a throughput of 2.24 × 10⁷ cells min⁻¹. The enrichment of rare cells was >2.02 × 10⁵-fold. Thus, this microfluidic system driven by purely hydrodynamic forces has practical potential to be applied either alone or as a sample preparation platform for fundamental studies and clinical applications.

Single-cell encapsulation in droplet microfluidics is commonly hindered by the tradeoff between cell suspension density and on-chip focusing performance. In this study, we introduce a novel droplet microfluidic chip to overcome this challenge. The chip comprises a double spiral focusing unit, a flow resistance-based sample enrichment module with fine-tunable outlets, and a crossflow droplet generation unit. Utilizing a low-density cell/bead suspension (2 × 10⁶ objects/mL), cells/beads are focused into a near-equidistant linear arrangement within the double spiral microchannel. The excess water phase is diverted while cells/beads remain focused and sequentially encapsulated in individual droplets. Focusing performance was assessed through numerical simulations and experiments at three flow rates (40, 60, 80 μL/min), demonstrating successful focusing at 40 and 80 μL/min for beads and cells, respectively. In addition, both simulation and experimental results revealed that the flow resistance at the sample enrichment module is adjustable by punching different outlets, allowing over 50% of the aqueous phase to be removed. YOLOv8n-based droplet detection algorithms realized the counting of cells/beads in droplets, statistically demonstrating single-cell and bead encapsulation rates of 72.2% and 79.2%, respectively. All the results indicate that this on-chip sample enrichment approach can be further developed and employed as a critical component in single-cell encapsulation in water-in-oil droplets.

Isolation and manipulation of single cells play a crucial role in drug screening. However, previously reported single-cell drug screening lacked multiple-dose concentration gradient studies, which limits their ability to predict drug performance accurately. To solve this problem, we constructed a multiconcentration gradient generator in which a Tai Chi-spiral mixer can accelerate solution mixing in a short time and produce a linear concentration gradient. Later, a gradient generator combined with a single-cell capture array was adopted to investigate the effects of single or combined doses of 5-fluorouracil and cisplatin on human hepatoma cells and human breast carcinoma cells (at the single-cell level). The results showed that both drugs were effective in inhibiting the growth of cancer cells, and the combination was more effective for human hepatoma cells. In addition, the relationship between the biomechanical heterogeneity (e.g., deformability and size) of tumor cells and potential drug resistance at the single-cell level was investigated, indicating that small and/or deformable cells were more resistant than large and/or less deformable cells. The device provides a simple and reliable platform for studying the optimal dosage of different drug candidates at the single-cell level and effectively screening single-agent chemotherapy regimens and combination therapies.