Researching cancer in tiny droplets of water
We spoke to Dr. Nachiket Shembekar, a post-doctoral researcher in Dr. Christoph Merten’s group at the European Molecular Biology Laboratory who told us about his method of using tiny droplets of water to conduct his research.
“Our team uses Microfluidics technology to answer biological questions. Microfluidics broadly means miniaturizing scientific experiments. For example, using chemicals in nano to pico litre (10-9 or 10-12 litres) volumes in very tiny chambers.
I work on something called a 'droplet based microfluidic platform.' This involves generating millions of small water (aqueous) in oil droplets. Each droplet serves as an individual reaction tube. We can put human cells, chemicals or proteins inside these tiny droplets.
The advantages of this technology are that it allows us to carry out several million reactions in synchrony. This means conducting the research in less time and using less chemicals making it much cheaper and very accurate.
My project, funded by Worldwide Cancer Research, focuses on developing improved antibody therapies against cancer. Antibodies are something similar to bullets, shot by the cells of the immune system. These antibodies can specifically attach to the cancer cells and help in their destruction. Antibodies are available as treatments against several cancers. However, sometimes they are not as effective as they could be and the cancer cells develop strategies to overcome the treatment.
The goal of my project is to use droplet based microfluidics to screen (test on a large scale) and identify useful antibodies that can help in killing the cancer cells. We hope that this can lead to a novel therapy for cancer patients which would be natural and therefore have minimal side effects. To give you an idea of what this work entails, here is an insight into a typical day in my lab.
08:30 – I’m on the bus which takes me along a winding road, up a beautiful hill in Heidelberg to reach my lab situated in the European Molecular Biology Laboratory (EMBL). I usually begin my day by checking and responding to important emails. There are also various email notifications about the day’s events in the institute such as talks by distinguished scientists or by postdocs, PhD students or workshops and conferences. I make a point to attend as many talks as I can. It is really exciting to hear what other scientists are working on. It inspires me as well sometimes as it shows me the way to tackle my own questions. If the topic is close to my research field then it keeps me updated about the latest developments.
Next, I check my calendar for today’s reservations for various work platforms or instruments. If we want to perform any experiments on a particular instrument, we need to reserve a time slot in advance, which helps in planning the time nicely. Today’s experiment is to correctly identify and separate droplets, which should be containing an antibody against a cancer protein, from thousands of other droplets. To do this I have booked instrument #2 from 10:00 to 12:00 and instrument #3 from 13:00 till 18:00.
09:00 – The cancer cells that I using are kept in the “Cell culture room”. Here, the cells are routinely “grown” inside small plastic bottles, providing them with “nutritious” media, humidity and at a suitable temperature, something similar to farming crops. I observe the cells under the microscope and check if they are healthy and then collect them. I seed a few of these cells, like sowing plant seeds. This involves moving roughly a hundred cells into some fresh media in a new bottle, to keep growing them for future experiments. I am going to use 2 types of cells that look identical to each other under the microscope - just like tiny white shiny balls. In order to differentiate between them, I colour one of the cell types with a ‘dye’, which makes them look red or blue or green under the microscope. Cells don’t like being out of their nice warm environment for long so in order to preserve them for a short while, I quickly put them on ice, similar to putting food in a fridge. I take a note that I will soon be running out of the “nutritious” media and I need to buy more.
10:00 – I bring the harvested cells into the main lab on the instrument #2. Then using a “microfluidic chip”, containing small chambers or channels (size wise it is in the range of 10-6 metres) through which the liquids flow, I generate lots of water-in-oil droplets (Image A).
The droplets are generated in such a manner that the cells are trapped inside the droplets. Since it is a random process, few droplets will contain the cells whereas many will remain empty. In order to keep these droplets separate from one another, we also put a surfactant in the oil which reduces the surface tension thus avoiding the fusion of droplets. Then I store the droplets at 37⁰ C (body temperature) to allow cells to release the antibodies inside the droplets.
11:30 – I go “shopping” to buy the nutritious media in the stores department of EMBL. All the chemicals that are commonly and routinely used by the labs are available in the stores. It is just like a supermarket experience, I take a trolley, and collect the chemicals that I need from various chambers. The only difference is I don’t have to pay anything immediately, but I have to make an invoice mentioning my budget number to which I will be charged later. A 500 ml bottle of a nutritious medium, which lasts for a month or two, costs anywhere between 10 to 40 Euros.
12:00 – Its lunch time! All the members of the Merten group usually have a delicious meal together in the EMBL canteen. While having lunch, a few of us discuss international issues or interesting travel plans, places or food, others talk about sports or movies and a few always end up discussing science!
12: 45 – Back from lunch and I start preparing for the next part of my experiment on instrument #3. This is situated in a “dark room” (as the name suggests, it is simply a room without light) and contains microscopes, lasers and photomultiplier tubes known as PMT's. PMTs are devices that can amplify light signals used for detecting fluorescence signals. The dark room is required since the fluorescent signal coming from a droplet is rather weak compared to ambient (natural) light.
The microscope is fitted with high speed cameras, so that we can actually see the droplets moving in a “chip” [Image B].
I am going to scan all the droplets through a laser to check which contain an antibody that binds to my cancer target. If the antibody binds to a cancer target inside the droplet, a sharp fluorescence peak will be displayed on the recording software just like an electrocardiogram (ECG). Based on this fluorescence peak, the desired droplet will then be pulled apart from the rest by an electric pulse. I can see all this happening live on the computer using high speed cameras. The whole process takes about 3-4 hours. I have to monitor all the parameters continuously. Sometimes the droplets do not flow at a consistent speed or a channel of the chip is blocked, so on and so forth.
17:00 - In the last step, I try to capture the individual droplets that were pulled aside and take a nice photograph under the microscope to make sure I have pulled out the correct ones [Image C].
17:30 – 18:00 - Finally, I note down my days’ experimental observations. I record any errors or problems that I faced while performing the experiment and a brief summary of the results in a lab book. This record is really helpful for future experiments to avoid repeating any mistakes. Before leaving the lab, to catch the 18:11 bus, I once again check my e-mails and make any future experiment bookings.
Making progress to reach our ultimate goal is always satisfactory. We are constantly improving the droplet microfluidics platform to help us identify useful antibodies.