Droplet microfluidics
Encapsulate single cells or reactions in picolitre droplets.
Our ruleSimulations prepare you for the bench but don't replace hands-on practice.
1Millions of identical picolitre test tubes
Droplet microfluidics turns a stream of water into a train of uniform droplets carried in an immiscible oil. At a T-junction or a flow-focusing nozzle the oil pinches the aqueous thread off at a fixed point, so every droplet has the same volume, typically a few picolitres to a few nanolitres, and they are produced hundreds to thousands per second.
Break-up at a junction
Size is set by the nozzle width, the ratio of the two flow rates and the capillary number of the continuous phase.
Surfactant
A fluorosurfactant coats each new interface, so droplets packed together in a tube do not fuse.
Poisson statistics
Cells or beads arrive at random, so single occupancy is bought by leaving most droplets empty.
Incubate, sort, read
Droplets can be collected, re-injected, merged with reagent, incubated and sorted by fluorescence.
2What sets the droplet size
In the dripping regime the droplet diameter scales with the nozzle width, grows roughly with the cube root of the ratio of dispersed to continuous flow, and shrinks as the continuous phase runs faster, because the governing number is the capillary number, the ratio of viscous shear to interfacial tension. Push the aqueous phase too hard, or run at a high capillary number, and break-up moves off the nozzle: the thread becomes a jet that breaks downstream at random and the emulsion turns polydisperse.
3Why most droplets have to be empty
Cells and beads are not metered into droplets; they arrive at random, so the number in a droplet is Poisson distributed with mean λ equal to the concentration times the droplet volume. At λ = 1, about 63% of droplets are occupied but 42% of those hold more than one cell.
4How runs go wrong
- Droplets of every size: jetting from too high a flow ratio, or a pulsating pump and compliant tubing.
- Droplets fusing in the collection tube: not enough surfactant, or the wrong oil and surfactant pairing.
- No droplets at all: hydrophilic channels, so the aqueous phase wets the walls.
- Occupancy that drifts downwards during the run: cells sedimenting in the syringe.
- A blocked nozzle: debris or clumps bigger than the constriction.
- Satellite droplets: small daughters formed at break-up, which confuse sizing and sorting.
Key terms
Practise these as flashcards →The phase that becomes droplets (usually aqueous) and the phase that carries them (usually a fluorinated oil).
A geometry where the continuous phase enters from both sides and squeezes the dispersed stream through a narrow nozzle.
Viscous shear over interfacial tension, μu/γ; the main control on droplet size and break-up regime.
Break-up at the nozzle, giving uniform droplets, versus break-up of an extended thread downstream, giving a broad distribution.
A PEG–perfluoropolyether block surfactant that stabilises water-in-fluorinated-oil droplets and is biocompatible.
Random occupancy of droplets by discrete particles, with mean λ = concentration × droplet volume.
Standard deviation of droplet diameter over its mean; a good junction gives a few per cent or less.
Adding a controlled small volume of reagent into droplets as they pass an electrified nozzle.
When to use it
- Single-cell assays and single-cell sequencing, where each cell's contents must stay separate.
- Digital, partitioned reactions such as droplet digital PCR, where counting positive partitions gives absolute quantification.
- Screening very large libraries of cells, enzymes or antibodies one variant at a time, with sorting of the hits.
- Making uniform particles, gel beads or capsules, where the droplet is a template rather than a reactor.
Compared with alternatives
The partitioned counting assay most often run in droplets; choose it when absolute nucleic-acid quantification is the goal.
The main application of droplet encapsulation, with barcoded beads co-loaded with the cells.
Sorts single cells without compartments; choose droplets when a secreted product or a reaction must stay with its cell.
How the junction is made, and why its surface chemistry has to be fluorophilic.