Secondment: Droplet Sorting in Cambridge

Today’s post was prepared by ES-Cat ESR Berndjan Eenink

A few weeks ago, I went to the Hollfelder lab for a week as part of my ES-Cat secondment. I plan to visit for a longer time (around a month) later in the Summer.

For my project, I use microfluidic droplets to screen large amounts of library variants. In our lab we can do most of the upstream and downstream work, but for microfluidics know-how and equipment I rely on our collaborators in the Cambridge group in the network.

The plan was to go for a short visit and to do some sorting on one of the libraries that I made to test if the whole process works, and if not, what I need to fix and/or improve before coming back for a longer visit in which I will sort through all my libraries.

In addition, we wanted to test a different method to express our cells for droplet sorting. Up until now we had encapsulated a single cell in each droplet. This works but has some issues. One of them is that cells seem to clump together after expression starts. This makes encapsulation tricky, as the cells have the tendency to stick together and form a biofilm, rather than moving neatly across the microfluidic channels as they ought to. Another problem is that after sorting the cells, the recovery is below 100% (not every cell that is sorted grows back.)

Growth and expression in droplets could potentially fix both of these issues. If no enzyme is expressed during the droplet formation, the sticking together is not an issue, and once expression starts the cells are free to stick together in the droplet however much they want. The recovery could also be helped, if each droplet contains (50-100) cells, 10-20% recovery still means getting back multiple copies of each variant. The reason to use microdroplets in the first place is to link the phenotype to the genotype (there is only one E. coli cell expressing a variant per droplet). Having many cells in each droplet may then sound counter-intuitive, but since a single cell is encapsulated, every cell that will grow inside the droplet is identical to the other cells in the droplet, thus phenotype and genotype are still linked. The advantage is that now we can oversample at a step where it is not that much extra time, instead of oversampling by sorting 5-10 times longer or with 5-10 times less coverage, now we can oversample by simply plating out more of the sorted cells (which is only a little bit of extra work in the evening).

Droplet sorting setup ready for action (as soon as the light goes out)

The visit as part of the secondment would started on a Tuesday. I arrived in Cambridge Monday afternoon, got to my airB&B and prepared a bit for starting the next day. Since we only had 6 days to make all droplets, do incubations and sort, the schedule was quite packed. I arrived on Tuesday morning to the lab and started preparing buffers, substrates, and whatever else was needed. I also got the samples that I had sent on dry-ice shipment a week ahead of me.

Immediately I (re)discovered the biggest problem of working someplace new. Where do these guys keep their Eppendorf tubes? Where is the medium? In which cupboard is this component of my buffer? Where are my tips? What is shared and what do I need to prepare myself? While I like to believe that as I get further along in my education and career I’m getting better at coming up with solutions to scientific or practical challenges, the answer to ‘where is this particular piece of material stored in this lab?’ remains as insurmountable as when I stepped into a lab for the first time.

The other thing that does not change when coming to a new lab (though this one pleasant, if at times a bit overwhelming) is meetings lots of new people and hearing about the stuff they are working on. I knew the people from the network, and some from previous visits during the network meetings and my previous sorting visit, but there were some new faces, as well as lots of faces I had to remember. ††

Droplets under the microscope

Since there was a limited amount of time, we set up both the envisioned new method and a repeat of the previous method as a comparison. This would also let us fall back to a method proven to be mediocre if the method thought to be great turned out to be dismal. We could look at some of the droplets under the microscope and confirm that something was actually growing. Over half were empty (they should be, as a side effect of preventing a lot of your droplets containing multiple cells), but the rest were all ‘teeming’ with cells. Some clumping (aforementioned problem) was observed, but since the cells are already in droplet this should not affect the encapsulation anymore.

A droplet being sorted

We did the screening at multiple timepoints, and using two different protocols for droplet growth and recovered the sorted cells, which I plated on LB-agar. The sorting seemed to work, and in all cases at least around 10 cells per droplet (based on calculations and estimations) were recovered. Due to the packed time-schedule I did not have time to do follow-up experiments to verify the recovery while in Cambridge, but luckily all the sorting work results in just a few Eppendorf tubes worth of material per sorted library, so these would be sent after me back to Munster, where I could do the follow-up experiments to verify that we actually sorted what we thought.

On Sunday, I managed to wrap everything up and get ready for the first train Monday morning to make it to the plane back to Germany. That Tuesday, I was scheduled for a regular research update for my group (either ideally or horribly timed, depending on which way you look at it). Since then, I did follow-up checks on the cells were recovered and found out that one of the protocols for the new method worked quite well, although there are still some issues that could be fixed to improve efficiency.

All in all, it was a short and busy but both productive and enjoyable stay.

Accurate science in video games: a new tool for sharing science to make it more accessible?

Today’s post was prepared by ES-Cat ESR Nikolas Capra

Video games made their breakthrough in the 80s, but have been part of our daily life and pop culture for almost 10 years. Like comic book characters and superheroes, they have become popular, and have an estimated 2.5 billion players worldwide1 (Fig. 1). Of course, video games cover a wide range of genres and topics, and luckily science found a place among all the shooters and well narrated, breath-taking, plots.

Figure 1: Number of video game players around the world in 2016

Before 1998, scientists in video games were almost always evil and villainous (like Dr. Eggman from the Sonic series) but this situation was saved by Dr. Gordon Freeman, a fictional theoretical physicist featured in the Half-Life series. Unfortunately, he had to fight aliens with a crowbar, not with science.

Again, in 1998, a masterpiece appeared on the console market by the name of Metal Gear Solid. Although the game is tactical-action and has absolutely no glimpses of scientists, the plot starts to introduce the topic of genetic engineering and nanotechnology (Fig. 2). The main character Solid Snake, who has been injected with nanorobots, must infiltrate a facility and fight villains that have been enhanced genetically, making his way among the genomic soldiers. This time there are no rogue scientists but a misuse of science. It might be that I overthink things, but the message that struck me, other than feeding my interest in science, is how science can be used for the wrong purposes, even if the technologies were made with the purpose of improving life quality. Besides the message, it has been a powerful vehicle for science by inspiring young players to know more about it, including me.

Figure 2: Sorry Snake.

This mixture of science, action and sci-fi opened the doors to science in the video game world.

Since the release of Metal Gear Solid, the topic of science in games has reached a crescendo. Many video games over the next years used science to enrich their plots. Some accurately, some…originally.

Covering a wide range of topics, from biology to engineering and physics, games are becoming more accurate and some of them are totally focused on the science, like in Kerbal Space Program where you need to build a space rocket, take off and land, adjusting the flight according to gravity and your orbit. To be honest, it was too difficult for me to play so I couldn’t try all the features and explore the physics behind the game.

But since we are way fonder of biology than physics and there are a huge number of games that involve science, it’s easier to make a list of the most (in)famous biology moments in video games.
Therefore, here is my personal list of the “Best Biology in Games”.

+ Resident Evil (1998): This famous game (and spin-off movie), paints a portrait of a world where a modified virus capable of bringing dead cells back to life, makes its way to the outer world. Despite the fictional idea of bringing back dead cells, the so-called T-virus, being modified from Ebola, can be spread only though saliva or blood and modifies the host on a cellular level. In the years after, the infectious virus is replaced by a parasite that modifies the host’s behaviour. Considering that parasites that changes host behaviour exist in nature (like the Cordyceps, also known as “zombie fungus”), I would consider the biology quite accurate.

+ + Spider-Man (2018): There is no need for introduction. Spider-Man has been part of many people’s childhoods. I’m not here to discuss whether being bitten by a radioactive spider gives you superpowers or a horrible and painful death. We all like to dream that if it ever happened, we would wake up with the amazing features of the animal that bit us. Except if it is a skunk. That’s the worst superpower. EVER. Terrible for social interactions too. Anyway, in this video game, Norman Osborne finds a cure that can correct genetic abnormalities. I was like “Hey, that’s cool but tell us how!” and apparently this cure uses CRISPR genome editing combined with an AI controlled gRNA to identify and edit mutations in the DNA. I would say that is quite an impressive mention in a video game. But the even cooler thing is that machine learning coupled with genome editing is not sci-fi but actual science and you can find it in here. This is shockingly accurate for being a video game on a non-biology topic.

+ + + Plague Inc. (2012): This is absolutely the most accurate non-educational game featuring biology. The game is very simple: select a host, infect people, change your features, annihilate the whole world (Fig. 3). The starting choices are “the usual suspects”: bacteria, virus or parasite. Each one of them has different traits such as fast replication rate and sturdiness for the bacteria, high mutation rate for the virus, and low detection for the parasite. Through the game you have to manage virulence, infectiousness and lethality. The goal is to infect and kill every human on Earth but as the plague spreads, the humans will respond by closing airports, harbours and they’ll start cooperating to find a cure to the disease and stop it from destroying humanity as we know it. The game was so realistic that has even attracted the attention of the CDC!

Figure 3: Host selection screen in the game Plague Inc.

Of course, there are more games concerning science and game designed by scientists to allow people to help with research, like the protein-folding game Foldit. There are also a variety of educational games that unfortunately are not well-know or promoted.

I will leave you then with a question: can we scientists use video games as a tool to inspire young players to become scientists? Can we use games to explain complex concepts in an easy and appealing way to share science with non-scientists?

Science and knowledge belong to the world and as a scientist, I think that our duty includes gifting our knowledge to everyone with every means we can use.


MTR – Cambridge, 2019

The ES-Cat Midterm Review (Interim) Meeting was held at the Department of Biochemistry, University of Cambridge, on the 14th and 15th of January, 2019.

This meeting was an opportunity for the ESRs to meet up, present their research progress to each other, and meet with EU representatives. As we are at the midway point of the ES-Cat Consortium, a major focus of this meeting was career opportunities for ESRs. They were encouraged to be proactive in planning for life after their PhDs, and to make use of LinkedIn to highlight their broad skill sets and seek out opportunities.

So… ES-Cat is now on LinkedIn!

During the MTR, ESRs also attended a facility tour of Johnson Matthey. Solid state chemistry falls a bit outside of many of the ESRs’ specialties, so it was a great opportunity to see a very different type of lab (including some stellar small molecule crystallography equipment) and learn more about the pharmacy industry.

We will be posting more frequently here, so if you’re interested in the work being done as part of the ES-Cat network, add us to your RSS feed, or check back often!