P2 - Sequential Capture and Pairing of Cells and Particles Using a Microfluidic Array

Introduction: : Microfluidic systems are indispensable for single cell studies, enabling precise cellular manipulation and stimulation. However, many existing designs often fall short in achieving robust particle/cell pairing in a high-throughput format without relying on complex fabrication techniques or flow manipulation schemes. Here, we present a novel bypass-channel-based microfluidic platform capable of high-efficiency, deterministic capturing and pairing both rigid particles and deformable cells under physiologically relevant flow conditions.

Materials and

Methods: : Each trap site comprises a U-shaped capture cup with a constricted trap channel at its base, which connects to the outlet channel, and two symmetric bypass channels flanking the capture cup. This configuration enables path-of-least-resistance trapping [1, 2], where the first particle enters the capture cup, followed by sequential recruitment of a second particle through residual flow in the integrated bypass channels. Once a pair is formed, the increased flow resistance through the capture cup diverts subsequent particles downstream, enabling sequential trapping. Unlike previous designs, our approach incorporates the bypass channels within the capture cup itself, enhancing pairing efficiency and spatial control [1, 3-6].
Devices were fabricated via standard soft lithography, with trap sites arranged in a linear array between inlet and outlet channels, Capture cup dimensions were tailored to match target particle sizes, resulting in widths of 17, 22, and 32 µm and depths of 32, 42, and 62 µm for 15, 20, and 30 µm particles, respectively. Colloidal solutions were introduced at flow rates (20 and 50 µL/hr) using syringe pumps. Particle and cell capture events were monitored via time-lapse microscopy. Trapping and pairing kinetics were analyzed across various flow rates and particle concentrations.

Results, Conclusions, and Discussions:: Our microfluidic device achieved consistent pairing across all tested particle sizes (Fig. 1). Time-lapse imaging confirmed deterministic, sequential occupation of trap sites. Trapping times were strongly dependent on flow rate and particle concentrations. At 20 µL/hr, average total times to trap and pair particles in a five-trap array were 60s (15 µm), 45s (20 µm), and 36s (30 µm). These times dropped to 24s, 18s, and 14.4s, respectively at 50 µL/hr. Particle arrival followed Poisson kinetics, where the total trapping times can be represented by a gamma (Erlang) distribution (Fig. 2).
The same trap design was implemented to trap and pair U87 and HEK cells (~12-15 µm diameter) (Fig. 3), demonstrating compatibility with soft biological samples. Captured cells remained viable, and deformation during trapping was minimal, underscoring the importance of optimizing trap geometry for heterogeneous soft particles. Our results underscore the critical role of trap design in achieving reliable particle/cell pairing. This microfluidic platform offers a robust and scalable solution for high-throughput studies of single-cell pairs, enabling the application of precise chemical and mechanical cues. The versatility of our approach supports a wide range of investigations in mechanotransduction, intercellular force transmission, and cell signaling.

Acknowledgements and/or References (Optional):: REFERENCES

[1] S. Kobel, A. Valero, J. Latt, P. Renaud, and M. Lutolf, "Optimization of microfluidic single cell trapping for long-term on-chip culture," Lab on a Chip, vol. 10, no. 7, pp. 857-863, 2010.
[2] W.-H. Tan and S. Takeuchi, "A trap-and-release integrated microfluidic system for dynamic microarray applications," Proceedings of the National Academy of Sciences, vol. 104, no. 4, pp. 1146-1151, 2007/01/23 2007, doi: 10.1073/pnas.0606625104.
[3] J. Chung, Y.-J. Kim, and E. Yoon, "Highly-efficient single-cell capture in microfluidic array chips using differential hydrodynamic guiding structures," Applied Physics Letters, vol. 98, no. 12, p. 123701, 2011, doi: 10.1063/1.3565236.
[4] J.-P. Frimat et al., "A microfluidic array with cellular valving for single cell co-culture," Lab on a Chip, 10.1039/C0LC00172D vol. 11, no. 2, pp. 231-237, 2011, doi: 10.1039/C0LC00172D.
[5] D. Jin et al., "A microfluidic device enabling high-efficiency single cell trapping," Biomicrofluidics, vol. 9, no. 1, p. 014101, 2015, doi: 10.1063/1.4905428.
[6] A. Guan, A. Shenoy, R. Smith, and Z. Li, "Streamline based design guideline for deterministic microfluidic hydrodynamic single cell traps," Biomicrofluidics, vol. 9, no. 2, p. 024103, 2015, doi: 10.1063/1.4914469.