Printed Droplet Microfluidics for on demand dispensing of picoliter droplets and cells

Abstract

Although the elementary unit of biology is the cell, high throughput methods for the microscale manipulation of cells and reagents are limited. The existing options are either slow, lack single cell specificity, or utilize fluid volumes out of scale with those of cells. Here, we present Printed Droplet Microfluidics, a technology to dispense picoliter droplets and cells with deterministic control. The core technology is a fluorescence-activated droplet sorter coupled to a specialized substrate that together act as a picoliter droplet and single cell printer, enabling high throughput generation of intricate arrays of droplets, cells, and microparticles. Printed Droplet Microfluidics provides a programmable and robust technology to construct arrays of defined cell and reagent combinations and to integrate multiple measurement modalities together in a single assay.

Linked optical and gene expression profiling of single cells at high-throughput

“…Our single-cell analysis platform is based on Printed Droplet Microfluidics (PDM) [28,29], an approach that allows cells to be optically scanned and dispensed to custom nanoliter well plates (nanoplates) ( Fig. 1a).…”

“…Thus, to dispense it into the nanowell, electrodes positioned under the substrate emit an oscillating electric field. This field pulls the dispensed droplet into the nanowell by a dielectrophoretic force and is key to the speed of PDM, since it allows a droplet to travel the final few hundred microns from the printing nozzle to the nanowell in tens of milliseconds [28]. Moreover, because the trap extends above the substrate, the printer need not dispense the droplets with perfect accuracy into the nanowells, since any droplet within the electric field will, ultimately, be pulled into the nearest nanowell.…”

“…A strength of PDM for scRNA-seq is that it allows the nanowells to be controllably loaded with cells and beads; this contrasts with other high-throughput scRNA-seq methods which randomly load cells or beads and, thus, are less efficient. Moreover, PDM allows systematic variation of nanowell contents across the array, to choose conditions that maximize data quality [28]. For instance, minimizing the rate of doublets-when two cells are inadvertently sequenced as one-leads to better data quality and lower per-cell sequencing costs [31,32].…”

“…PDM chips are fabricated by poly(dimethylsiloxane) (PDMS) molding over a SU-8 master. Briefly, a three-layer SU-8 negative master is patterned to form 20, 80, and 150 μm tall features using previously described multi-layer SU-8 photolithography techniques [28]. Following casting of PDMS over the SU-8 and curing at 65°for 2 h, inlet holes are punched into devices using a 0.75-mm biopsy core.…”

“…Channels are then treated with AquaPel (AquaPel). Drop-making devices are fabricated as previously described [28]. Two devices with a T-junction cross-section of 80 μm × 45 μm and 80 μm × 80 μm are used.…”