High‐throughput deterministic single‐cell encapsulation and droplet pairing, fusion, and shrinkage in a single microfluidic device

Abstract

In this article, we present a microfluidic device capable of successive high-yield single-cell encapsulation in droplets, with additional droplet pairing, fusion, and shrinkage. Deterministic single-cell encapsulation is realized using Dean-coupled inertial ordering of cells in a Yin-Yang-shaped curved microchannel using a double T-junction, with a frequency over 2000 Hz, followed by controlled droplet pairing with a 100% success rate. Subsequently, droplet fusion is realized using electrical actuation resulting in electro-coalescence of two droplets, each containing a single HL60 cell, with 95% efficiency. Finally, volume reduction of the fused droplet up to 75% is achieved by a triple pitchfork structure. This droplet volume reduction is necessary to obtain close cell-cell membrane contact necessary for final cell electrofusion, leading to hybridoma formation, which is the ultimate aim of this research.

Droplet Manipulations in Two Phase Flow Microfluidics

“…The thin oil film is squeezed out until the droplets merge. Applying a high frequency (kHz) electric field perturbs the interface between the droplets increasing the efficacy of merging [107]. In the same manner for EWOD and electrostatic potential wells two droplets have to be brought towards neighboring electrodes and the AC frequency of the applied voltage helps to destabilize the interface and speeds up the coalescence of droplets [13].…”

Electrowetting as a tool for two phase flow microfluidic operations

“…The step can also be a gradual change in height where the gradient influences the eventual droplet size. 34 The step emulsification method has the advantage that the drop size is mainly controlled by the geometry and the resulting change in Laplace pressure, and less influenced by Required manipulations pressure differences between the liquid phases. This allows to parallelize the step geometry and thus generate a high throughput of reasonably uniform droplets.…”

“…4-5 also shows the ability to coalesce 2, 3, 4 or 5 droplets at the trapping electrodes. Compared to other methods for electric field induced merging of droplets, 34,35 our technique has two advantages: the number of coalescing droplets is flexible and can be controlled via the voltage, and the droplets do not need to reach the electrodes simultaneously: the first droplet is simply trapped and waits for the consecutive droplet(s) to merge, after which the final drop is released automatically (See Appendix 4D and Movie 4-1).…”

“…34,35 We still incorporate this system in our microchannel, but also add a new feature: we exploit electrostatic potential wells to manipulate drops in a microchannel flow. Several operations can be implemented, such as on-demand trapping and release, which is crucial for the storage of drops that need to be analyzed in situ for some time.…”

“…These allow to steer a droplet in flow along these rails, 33 or to fuse two droplets by using two converging rails. 34 Other researchers used similar rails for the fusion and sorting of droplets. 35 As is the case with other passive techniques: it is difficult to release the drop once trapped, or to select which way to steer a drop.…”