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6 Top Tips Resolving Challenges of Droplet Generation

Updated: Mar 3, 2022

Double-Encapsulation:  Single-cell droplet in oil shell
Double-Encapsulation: Single-cell droplet in oil shell

Droplet generation studies have been growing considerably in the past few years mainly due to the significant advantages they 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.   But as with the evolution of any new field, there are also challenges so we've put this blog together to highlight some of the most common questions we get asked by our customers when setting up their own droplet generation experiments.

Cellix's Top Tips:

1. Protocol:  The order in which you fill the microfluidic channel with different solutions is critical! Flow sensors (which are connected to the pumps) are calibrated for different liquid types; e.g. for oil or aqueous-based solutions. It is a common mistake to connect both pumps (oil and aqueous phase) to the chip at the start of the experiment! If you then try to prime the chip by pre-filling with the oil phase while the chip is connected to the pump containing the aqueous phase; oil is likely to back-flow into the sensor connected to the pump for the aqueous phase.  The flow sensor connected to the aqueous 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 aqueous phase once you start your experiment. 

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 DropGen PreCoat for use with our DropChips.  

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

  • Connect your microfluidic pump for the water phase to the flow sensor and pre-fill with the aqueous 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. Achieving 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: