Self-synchronization of reinjected droplets for highefficiency droplet pairing and merging

Droplet merging serves as a powerful tool to add reagents to moving droplets for biological and chemical reactions.However,unsynchronized droplet pairing impedes high-effciency merging.Here,we develop a microfluidic design for the self-synchronization of reinjected droplets.A periodic increase in the hydrodynamic resistance caused by droplet blocking a T-junction enables automatic pairing of droplets.After inducing spacing,the paired droplets merge downstream under an electric field.The blockage-based design can achieve a 100%synchronization efficiency even when the mismatch rate of droplet frequencies reaches 10%.Over 98%of the droplets can still be synchronized at nonuniform droplet sizes and fluctuating reinjection flow rates.Moreover,the droplet pairing ratio can be adjusted flexibly for on-demand sample addition.Using this system,we merge two groups of droplets encapsulating enzyme/substrate,demonstrating its capacity to conduct multi-step reactions.We also combine droplet sorting and merging to coencapsulate single cells and single beads,providing a basis for high-efficiency single-cell sequencing.We expect that this system can be integrated with other droplet manipulation systems for a broad range of chemical and biological applications.

Design and aligner-assisted fast fabrication of a microfluidic platform for quasi-3D cell studies on an elastic polymer

While most studies of mechanical stimulation of cells are focused on two-dimensional(2D)and three-dimensional(3D)systems,it is rare to study the effects of cyclic stretching on cells under a quasi-3D microenvironment as a linkage between 2D and 3D.Herein,we report a new method to prepare an elastic membrane with topographic microstructures and integrate the membrane into a microfluidic chip.The fabrication difficulty lay not only in the preparation of microstructures but also in the alignment and bonding of the patterned membrane to other layers.To resolve the problem,we designed and assembled a fast aligner that is cost-effective and convenient to operate.To enable quasi-3D microenvironment of cells,we fabricated polydimethylsiloxane(PDMS)microwell arrays(formed by micropillars of a few microns in diameter)with the microwell diameters close to the cell sizes.An appropriate plasma treatment was found to afford a coating-free approach to enable cell adhesion on PDMS.We examined three types of cells in 2D,quasi-3D,and 3D microenvironments;the cell adhesion results showed that quasi-3D cells behaved between 2D and 3D cells.We also constructed transgenic human mesenchymal stem cells(hMSCs);under cyclic stretching,the visualizable live hMSCs in microwells were found to orientate differently from in a 3D Matrigel matrix and migrate differently from on a 2D flat plate.This study not only provides valuable tools for microfabrication of a microfluidic device for cell studies,but also inspires further studies of the topological effects of biomaterials on cells.