Welcome to FOCUS In Sound, the podcast series from the FOCUS newsletter published by the Burroughs Wellcome Fund. I’m your host, science writer Ernie Hood.
In this edition of FOCUS In Sound, we meet a Burroughs Wellcome Fund grantee who participated in the development of a remarkable new pathogen detection technology that may be vastly important in the detection and surveillance of COVID-19, among several other pathogens. He and his colleagues have just had a landmark paper published in Nature, describing the technology and its importance in the battle against the pandemic pathogen. With this story moving so rapidly, I should include that we recorded this interview on April 17, 2020.
Dr. Paul Blainey is a core member of the Broad Institute of MIT and Harvard, and a tenured associate professor in the Department of Biological Engineering at MIT. He is an expert in microanalysis systems for studies of individual molecules and cells. He is applying this technology to advance the understanding of DNA-protein interactions, evolutionary processes, functional differences between cells, disease processes, and drug target discovery.
Paul holds a BS in chemistry and a BA in mathematics from the University of Washington, and earned his PhD in physical chemistry from Harvard University. He completed his postdoctoral research at Stanford University in the laboratory of Stephen Quake. He was recognized by the Burroughs Wellcome Fund in 2011 with the Career Award at the Scientific Interface, when he was still at Stanford. The award description in 2011 was particularly prescient, as it talked about “from single cells to populations: using microfluidics, genomics, and culture to better understand infectious disease in the post-genomic era.”
The just-published Nature paper is titled “Massively multiplexed nucleic acid detection with Cas13.” The new platform it describes may be a huge game-changer in the quest for large-scale testing for COVID-19 infections.
Paul Blainey, welcome to Focus In Sound…
Thank you very much, I’m so glad to be here.
Paul, introduce us to this new testing platform that you and your colleagues have developed called CARMEN-Cas13…
This has been a wonderful effort by a really amazing team, and so let me start by acknowledging the rest of the team. This was a really tightly integrated collaboration between my research group, Pardis Sabeti’s research group at the Broad, and Deb Hung’s research group at the Broad. And in particular I want to acknowledge a lot of the junior team members, particularly postdocs Cheri Ackerman, Cameron Myhrvold, and two students, Gowthan Thakku and Catherine Freije, who made just essential contributions along with the broader team.
Now, the system that we put together is a really interesting and powerful fusion of new microfluidic technology and an amazing CRISPR-based nucleic acid detection assay called SHERLOCK. What we did was find a way to implement this SHERLOCK assay chemistry in teeny-tiny, nanoliter-scale microdroplets that are spread out in an array that enables us to read them out using microscopy. And so what this has enabled is the capability to run many, many different nucleic acid detection tests, and here we do have a focus on detecting viral sequences, and to do this quickly and at a low cost per test.
How has CRISPR been incorporated into the system, and what role does it play?
In our system, CRISPR enzyme Cas13 plays a role in the nucleic acid detection chemistry. So it binds a nucleic molecule, and helps generate a signal that we detect using optical microscopy.
Was the ability to detect the COVID-19 coronavirus something you were already working on, or was it a situation where you could expand your system to include it?
I’ll back up and give a little more history on the microfluidic platform. So we started work on this quite a number of years ago, for a totally different application. We invented this platform for small molecule drug screening, and it was still several years ago that we decided to repurpose the microfluidic platform from small molecule screening to infectious disease diagnostics. COVID-19 and the SARS-CoV-2 virus were not on our radar at all. Our interest with the platform was to develop and deploy what we refer to as a pan-viral assay panel, that includes assays for essentially all the important viruses in the world that infect humans. And so from the beginning, we did have a coronavirus panel, but at that time CoV-2 was not known to the world. And so while the paper was in review, we learned about the outbreak, of course, and moved to really quickly develop a new assay for the SARS-CoV-2 virus, and incorporate that into our large pan-viral assay panel.
Did you have to make adjustments on the fly to accommodate the pandemic virus?
One of the things that’s really wonderful about our assay system is that it’s actually quite easy to incorporate new content, and we don’t have a lot of issues with crosstalk among different assays. So it was actually relatively straightforward to incorporate a new test to cover the strains involved in the outbreak, and we were able to do that in a short period of time.
Paul, I do want to hear more about the CARMEN-Cas13 platform, but before we expand our discussion, tell me, what do you see as the headline here, particularly given the current pandemic?
I think the headline here is really one about hope. We’re incorporating here not only a new assay chemistry, it’s actually not CRISPR technology but PCR technology that’s on the front lines of nucleic acid diagnostics addressing the outbreak today. And so there are these powerful new CRISPR-based assay chemistries. And then also on the instrumentation side, we’re introducing this microfluidic platform for microscale testing, which has some advantages in terms of reducing the cost per test and enabling many tests to be done. And so I think in the context of the pandemic, the headline is really that there is a lot of room for creativity and innovation to develop new types of tests to address it. And given the current situation, I think it’s quite clear we’re going to need an all-of-the-above strategy to beat back the pandemic.
How will use of this system contribute to improvements in COVID-19 testing? I know that’s been a major issue worldwide…
Our system is really focused on test throughput, and it can very efficiently test each patient sample for a large number of target sequences, which could be different parts of one virus, different strain variations of one virus, or a whole diversity of viruses in each sample. And so it’s fairly unique in that regard. We have not constructed it in a way that’s suitable yet for rapid point-of-care testing. And so we see the role for the reported version of the system as being in surveillance, either for clinical screening tests or for public health screening and surveillance testing.
Is the turnaround rapid in this case?
The test in the microwell system currently takes several hours to return a result. So it’s not really suitable for rapid point-of-care testing. Now we’re already working on future versions that would be much faster and use really simple instrumentation that would be well-suited to point-of-care implementation, or testing at the point of care.
I understand from the paper that the platform is “sensitive, specific, and statistically robust.” Can you tell us a little bit more about that?
Absolutely. We were really glad to see that even in the microscale, nanoliter format, that the CARMEN testing platform maintains the nanomolar sensitivity that had previously been demonstrated for the SHERLOCK assay chemistry. And so it really has this exquisite sensitivity while maintaining really high specificity at the same time. Now, one of the unique features of our droplet array platform is that each test actually consists of many of these nanoliter droplet replicates. So that gives us a lot of statistical robustness in order to make sure that we make the right call for each test, because we can check the result across many nanoliter-scale replicate assay reactions. Another feature is that that average number of reactions per test is continuously adjustable, and so we can dial in the statistical power of the test case by case, so that it’s most suitable for the application that it’s being used for.
Paul, the experts keep claiming that the only way we’re really going to get ahead of this pandemic, and perhaps prevent a recurrence, is a massive expansion of testing, both point-of-care testing and surveillance testing as you’ve been describing. Is this the vehicle to accomplish at least part of that daunting task?
I think our system plays an important role alongside some of these other testing systems and modalities, and in its current form, yes, particularly for surveillance, as you described. And I do agree that this kind of widespread testing is going to be really important in helping re-open the world’s economy as we recover from the outbreak. Now on top of that, I think it’s also pretty important the we continue with surveillance, and with broad surveillance for many different types of viruses, such as our platform can enable, in order to make sure that we detect and get ahead of the next outbreak that heads our way.
So how will the platform be rolled out and scaled for widespread use?
Well thanks for asking. We’re already moving in that direction at the Broad Institute to test whether the system is going to be suitable for screening clinical samples from the outbreak. Now beyond that, we're looking at partnering, and are already in advanced discussions, with the government and with different companies that have appropriate instrumentation and manufacturing systems in order to in order to deploy our system.
So you are already thinking in those terms, then?
Not even just thinking, Ernie, we’re really taking strong action, pushing hard to make the most positive impact on the outbreak that we can.
Paul, obviously CARMEN-Cas13 is going to be tremendously useful for much more than just COVID-19. Would you elaborate on that aspect for us?
Yeah, that is one of the unique advantages of our system, is it’s so flexible in terms of adding new tests, and it’s so efficient when conducting many different tests per sample. We think it does have a great role to play there. And so in addition to the pan-viral panel that we report, it will also likely be useful to create region-specific or outbreak-specific assay panels that really target the most relevant pathogens that are circulating in a given location at a given time. So with respect to the current pandemic, we’ve created now a respiratory panel that includes the outbreak strain as well as other respiratory viruses against which clinicians will need to make differential diagnoses.
What are the advantages of the CARMEN-Cas13 platform compared to others in terms of costs and sample volume requirements? I saw some reference to those aspects in the paper and found it very interesting.
Yeah, so the CARMEN-Cas13 platform does have exceedingly low costs and low sample volume requirements per test. So this is enabled by the low volume of each test reaction that’s carried out. That’s 2 nanoliters in the current system. And it results from the microfluidic automation that the platform enables. And so that eliminates the requirement for complex liquid-handling equipment or a lot of manual labor, even when many tests are being conducted.
One of the challenges when one is interested to carry out a lot of different tests per sample is the sample consumption. And so our pan-viral test panel includes more than 150 assays, which could be a problem in terms of sample consumption using a conventional approach. But because our reactions are so small, it’s no problem at all to carry out hundreds of tests, even from a small clinical sample.
So this is a real victory for microfluidics, isn’t it?
Yeah, I think it might be, although only time will tell. I’m a big fan of microfluidic technology myself, and it’s frustrating sometimes to see how much of a challenge it is for the world to adopt the most advanced microfluidic technologies. And I think there are different theories about why that is the case. One has to do with the complexity of the instrumentation that’s required to run microfluidic devices. And so one of the focuses we had in developing the microdrop array platform was really keeping it as simple as possible. And so it will be really interesting to see how that potentially enables greater adoption than other microfluidic systems.
The other interesting aspect of the platform we’re using for CARMEN-Cas13 testing is that it’s actually a fusion of two different flavors of microfluidics. We use both microdroplets, which are small water droplets surrounded by a carrier oil, and microwell arrays, which are cleaner surfaced studded with small microwells into which we put the droplets.
Tell us also about the massive-capacity chip you designed, which allows more than 4,500 tests per chip…
Yeah, absolutely. So I want to give credit to Cheri Ackerman, the postdoc in my lab who actually designed and developed the procedure for using that chip along with support from our broader droplet array project team. So this is a chip that we refer to as the M chip. It’s about the size of a smart phone and has a capacity to handle hundreds of thousands of droplets, just on the one chip. And so that’s really the capability that enables us to process so many samples and so many assays for different viral targets all in one go.
That is very impressive, absolutely. So where is your and your team’s research going from here?
Well, in the immediate term, we are very focused on deploying CARMEN-Cas13 technology to make a difference in the context of the current outbreak. And so we do have a big focus on these conversations with government and corporate partners about how to deploy this technology for large-scale testing here in the U.S. and around the world.
Now stretching out to the intermediate time scale, we are very interested in developing the next generation of CARMEN testing technology, and our focus there is reduce costs even further, generate systems suitable for deployment at the point of care, and improve the turnaround time of the tests.
We will certainly look forward to those developments. Paul, it’s been an enlightening conversation, and believe me, we are all pulling for you and your group to keep the important innovations coming as we face this incredible global challenge. Thank you for speaking with us!
Well it’s been a pleasure.
We hope you’ve enjoyed this edition of the FOCUS In Sound podcast. Until next time, this is Ernie Hood. Thanks for listening!