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Droplet generation studies have been growing considerably in the past few years mainly due to the significant advantages which such methods can bring, particularly for high-throughput single cell analysis: millions of single-cell reaction droplets in one eppendorf tube is the equivalent of >10,000 96-well plates with a single cell per well.  

Applications for droplet generation are far-reaching and go beyond drug discovery and diagnostics into such areas as food and cosmetics production; and industrial applications such as paints.  Microfluidic droplet generation can offer significant cost savings compared to more traditional production techniques and it's an exciting field which is under continuous development.   

Droplet Generation with DropChip & Cellix pumps


Droplets are generated or formed by precisely controlling immiscible liquids (usually aqueous and oil-based) with microfluidic pumps in microfluidic chips of a precise geometry.    This is known as "passive" droplet generation ("active" uses electric, magnetic or centrifugal means) and there are three types of geometries which are generally described:

Cross-flow:  T-junction or Y-junction geometry in microfluidic chip. For water-in-oil droplets; the oil sample (continuous phase) flows in one direction.  The aqueous sample (dispersed phase) flows into the oil sample at the T- or Y-junction.  As the aqueous sample joins the oil sample, the shear forces of the oil continuously flowing breaks the aqueous sample into a droplet.  In this case, the size of the droplet is determined by flow rate ratio of the oil and aqueous samples flowing in the channels of the microfluidic chip and the viscosity, velocity and interfacial tension of the oil sample.  The flow rate of the continuous phase (oil in this example) is usually higher than the dispersed phase (water).  To achieve oil-in-water droplets, the liquids are reversed.

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Cross-Flow Droplet Generation:

Water-in-oil droplets

Flow focusing:  "+" or X-junction geometry in microfluidic chip.

For water-in-oil droplets; the water sample (dispersed phase) meets the oil sample (continuous phase) at the junction, where there is usually a narrowing of the channels at the junction point.  Similar to cross-flow, the flow rate of the continuous phase (oil in this example) is usually higher than the flow rate of the dispersed phase (water).  In this case, it is possible to increase the size of the droplets by decreasing the flow rate of the continuous phase.  To achieve oil-in-water droplets, the liquids are reversed.

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Flow Focusing Droplet Generation:

Water-in-oil droplets

Co-Flow focusing: the dispersed phase channel is enclosed inside a continuous phase channel.  As the dispersed fluid enters the continuous phase fluid, it experiences the shear forces of the continuous phase fluid until it eventually breaks and forms a droplet by dripping or jetting.

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Co-Flow Focusing Droplet Generation:

Water-in-oil droplets

How does it work?


We can provide you with a complete set-up (Droplet Generation Kit) or just the components you need.  Click here to see some example experimental set-ups.

In general, as a minimum, you will need the following to execute droplet generation experiments:

  • 2 x Microfluidic pumps (or 2x channels on one pump): these are essential for flow control of the continuous (oil) phase and dispersed (water) phase. 

    • Depending on your application, we recommend the 4U pressure pump (4 channels to choose from) or 2x ExiGo microfluidic syringe pumps. 

    • 4U pressure pump pressurises the reservoirs in which the sample is held; this effectively pushes the fluid into the tubing which is connected to the microfluidic chip.  

    • ExiGo microfluidic syringe pumps use the syringes to hold the sample (oil or water phase).  If you are using the optional manifold for automatic refilling of the syringe, a reservoir to hold the sample can also be provided.

  • 2 x Flow sensors:  required for active feedback of the flow control of both the oil and water phases.  Fluidic capacitance in microfluidic devices can often introduce instabilities when trying to vary flow rates.  Therefore, it is extremely important to have accurate microfluidic pumps with flow sensors to limit such instabilities enabling you to product uniform, stable droplets of a consistent size, volume and frequency.  

  • Microfluidic Chip with appropriate geometry to create droplets: the geometry of the junction (where the oil and water phase meet) is the most influential factor affecting droplet size.  Once this is fixed, the droplet size is affected (to a lesser extent) by surfactant concentration and the ratio of flow rates of the oil and water phases.  

  • Stable Channel Surface Chemistry to ensure droplet stability:  stabilises interface between oil and water phase giving stability to the droplets.

    • Water-in-oil droplets: you will need a hydrophobic channel surface and this can be achieved by pre-filling the microfluidic chip with DropCoat.

    • Oil-in-water droplets:  in this case, you need a hydrophilic channel surface.  Cellix's DropChip channels are sold pre-treated so that the surface of the channels are hydrophilic.  

  • Surfactant:  stabilises the interface between oil and water phase giving stability to the droplets.  

  • Oil for continuous phase:  surfactant is often added to the oil to improve droplet stability.  

  • Tubing to connect from your pumps to the microfluidic chip


Flow-focusing set-up:  water-in-oil droplets

  • 4U Pressure pump (using 2x channels) or 2x ExiGo syringe pumps

  • 1x Sample Reservoirs (with 4U pump) or 2x syringes with ExiGo pumps

  • 2 x Flow Sensors

  • Microfluidic chip with hydrophobic channel surface treatment:

    • Flow splitter to split the flow from the continuous (oil) phase​

    • "Droplet generator" junction as shown in the image

  • Surfactant, oil, tubing connections.

Flow-focusing set-up:  water-in-oil droplets with 2 aqueous phases

  • 4U Pressure pump (using 3x channels) or 3x ExiGo syringe pumps

  • 2x Sample Reservoirs (with 4U pump) or 3x syringes with ExiGo pumps

  • 3 x Flow Sensors

  • Microfluidic chip with hydrophobic channel surface treatment:

    • Flow splitter to split the flow from the continuous (oil) phase​

    • "Droplet generator" junction as shown in the image

  • Surfactant, oil, tubing connections.

What do I need to get started?
Example experimental set-ups


  • Cell / Protein encapsulation and sorting on-chip

  • DNA sequencing for target selection / enrichment.

  • Sample / reagent mixing and control of reactions on-chip

  • DNA amplification

  • Biomolecule synthesis

  • Diagnostic chips

  • Drug discovery


CELLIX TOP TIPS - Resolve common challenges with Droplet Generation

1. Protocol:  The order in which you fill the microfluidic channel with different solutions/liquids (e.g. solution to ensure channel hydrophobicity, oil and water phase) is critical as flow sensors are calibrated for different liquid types (e.g. for oil or aqueous-based solutions)!  Before you start, ensure the chip is not connected to the pump which contains the water phase.  It is a common mistake to connect both pumps (oil and water phase) to the chip at the start of the experiment!  If you try to coat the chip with a solution to ensure channel hydrophobicity or pre-fill the chip with the oil phase while the chip is connected to the pump containing the water phase; oil is likely to back-flow into the sensor connected to the pump for the water phase.  The flow sensor connected to the water phase is only calibrated for aqueous-based solutions.  If a non-aqueous solution, such as oil, flows into this flow sensor, it will result in incorrect flow rates being delivered for the water phase once you start your experiment. 

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Flow sensors are calibrated by liquid type

Cellix's top tips:

  • Disconnect the chip from the pump which contains the water phase

  • Pre-coat the microfluidic channels ​of your chip with solution to ensure it is hydrophobic.  Cellix recommends DropCoat for use with our DropChips.  

  • Pre-fill the microfluidic channels of your chip with the oil phase - remember to keep the pump for the water phase disconnected!

  • Connect your microfluidic pump for the water phase to the flow sensor and pre-fill with the water phase until you see a droplet coming out at the end of the tubing.  You may now connect this tubing to your microfluidic chip.

2. Uniform, stable droplets:  Two of the top factors affecting droplet stability are channel surface chemistry (hydrophobic for water-in-oil droplets and hydrophilic for oil-in-water droplets) and addition of surfactant in the oil phase which stabilises the interface between the oil and water phases.  After this, it comes down to tweaking the ratio of the flow rates of the oil and water phases.  

Cellix's top tips:

  • Water-in-oil droplets need a hydrophobic channel:  Ensure the surface of the microfluidic channel is hydrophobic:  The surface of the microfluidic channel must be hydrophobic as this prevents adhesion of your continuous (oil) or dispersed (water-based) liquids on the surface which will disrupt the formation of  your droplets.  PDMS is naturally hydrophobic but is often sealed with a glass coverslip which is hydrophilic.  To ensure you have a completely hydrophobic surface, the first thing you should do is fill the microfluidic channel with a solutions such as Cellix's DropCoat.  Leave this in the channel for at least 10 minutes, then flush it out with air or immediately prime with the oil phase before starting your experiment.  If you have a PDMS microfluidic chip, you can try drying it out in an oven (~85C) for a few minutes but we don't recommend this for plastic chips.  

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Coating a biochip with DropCoat ensuring hydrophobic channels

  • Prime the channels of your microfluidic chip: Also make sure that you fill your channel with your continous phase liquid first; i.e. oil for water-in-oil droplets or water phase for oil-in-water droplets.  

  • You can also try increasing the concentration of your surfactant in oil.

3. Droplet generation stops mid-way through experiment and both oil and water-based phases become laminar flow:  The flow is unstable.  There can be a number of reasons for this.  

Cellix's top tips:

  • Water-in-oil droplets need a hydrophobic channel:  Did you coat the microfluidic channels first with DropCoat or other similar reagent to ensure hydrophobicity?  If not, start again...

  • Check for leaks or blockages:  If you did coat the microfluidic channels with a reagent to ensure hydrophobicity (for water-in-oil droplets only); then you should check to ensure that there are no leaks or blockages (e.g. air bubbles) in your system - this can often result in a change in flow rate​ which disrupts the formation of droplets.  

  • Optimise the flow rates:  If there are no leaks or blockages, try keeping the flow rate of the oil phase constant while slowly ramping up the flow rate of the aqueous phase but make sure you do not allow the oil phase to back-flow into the sensor controlling the water phase.  See our tip above about protocol.

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ExiGo syringe pumps &

UniGo pressure pumps

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4U Pressure Pump

4. Controlling Droplet Size:  The most important factor affecting droplet size is the geometry of the microfluidic chip, in particular, the junction where the oil and water phase meet.

Cellix's top tips:

  • Geometry:  Select the channel geometry (particularly the junction where the oil and water phases meet) of the microfluidic chip such that it can easily create the droplet sizes that you want.  Check out our DropChip page for a table with details of the droplet diameter and volumes that can be achieved with Cellix's DropChip. 

  • Surfactant:  To a lesser extent than geometry, the surfactant concentration (normally added to the oil phase) will affect the size of the droplets.  By altering the surfactant concentration, you will slightly alter the ratio of flow of the different phases enabling you to achieve very slightly different sizes; i.e. you can achieve more water in your droplet, thereby making it a slightly bigger size.  Cellix's recommends DropSurf surfactant for droplet generation studies.

  • Ratio of flow rates of oil and water phases:  If the geometry of the microfulidic chip is fixed, altering the ratio of flow rates of the 2 phases has a much lesser affect on droplet sizes.  However, it can still produce a minor change but there is typically only a very small range of flow rates for a specific fixed geometry that will work so this will only give you limited flexibility.

Droplet Generator 1 and 3_Flow Focusing.

Geometry of the junction is one of the biggest factors affecting droplet size

6. Measuring droplet stability (size, distribution) and monodispersity:  There are lots of software packages available which work well, including Image-Pro Premier offered by Cellix.  However, it should be noted that all of these software programs work in 2D and so precision is often compromised.  Even a very small change in diameter of the droplet can have a significant effect on the volume of the droplet. 

Cellix's top tip:  When developing our DropChip and characterising the droplets which were generated, we used scattering (3D) - this enables us to accurately measure the size, volume and frequency of droplet generation.  There is a significant amount of work associated with characterisation of droplets but if you want to be absolutely confident, we recommend using a 3D method such as scattering.

Droplet Generation Frequency Graph.jpg

Droplet generation frequency dependent on ratio of flow rates

Cellix Top Tips

5. Biocompatible oil and surfactants for droplet generation:  This is a common question for those interested in applications related to cell-based work.  

Cellix's top tip:  Mineral oil works very well for the continuous phase and is widely available but biocompatible surfactants are not as widely available and tend to be quite expensive.  Cellix recommends DropOil for droplet generation studies.


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4U Pressure Pump

Droplet Generation - Microfluidic Syring

ExiGo Syringe Pump

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Flow Sensors



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