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Dionis Minev on Using Crisscross Assembly for Diagnostics

The Humans of the Wyss (HOW) series features members of the Wyss community discussing their work, the influences that shape them as scientists, and their collaborations at the Wyss Institute and beyond. 

Dionis Minev truly values the creative design process, whether he’s using it to produce a beautiful painting or to build a new DNA structure. He is the team lead on the Crisscross Nanosciences Validation Project and is working with his team to create a rapid, low-cost, point-of-care diagnostic device. Learn more about Dionis and his work in this month’s Humans of the Wyss.

What are you working on?

Dionis Minev on Using Crisscross Assembly for Diagnostics
Dionis Minev, Postdoctoral Fellow. Credit: Wyss Institute at Harvard University.

I’m working in William Shih’s lab on DNA nanotechnology to create tools and devices engineered to overcome specific bottlenecks in the development of new therapies and diagnostics. Specifically, I’m working on a system called crisscross cooperative self-assembly, also known as crisscross polymerization, with Anastasia Ershova, Chris Wintersinger, and Jim MacDonald, who is part of the Advanced Technology Team, along with some others in the Shih lab.

Crisscross is a programmable DNA self-assembly method that allows researchers to initiate the assembly of large DNA structures, starting from a tiny seed structure. I spend the majority of my time working on crisscross with single-stranded DNA. The strands­—which we call slats—weave over and under each other forming large, micron-scale DNA ribbons. We believe that crisscross assembly is the third step in the evolution of DNA nanotechnology tools after DNA origami and DNA bricks. This method can be used to precisely assemble structures from single strands of DNA all the way to multicomponent DNA origami assemblies. Crisscross solves the difficult problem of precisely controlling the formation—otherwise known as nucleation—of higher order nucleic acid assemblies. The formation is entirely seed dependent under a wide range of reaction conditions making it a robust self-assembly tool.

Though this method has a wide variety of applications, I’m specifically using crisscross assembly to work on the detection of biomarkers, whether they are DNA, RNA, or proteins. The idea is to build a system that amplifies a signal once you’ve captured a single biomarker molecule of interest. The signal is amplified by this assembly method as a large DNA structure that’s easily detectable on a gel, plate reader, or low-cost microscope. What’s novel about our system is that it is extremely sensitive, by eliminating unwanted background noise: it makes single biomarker molecules easily detectable and works without any enzymes which could make it a really powerful diagnostic tool. Most DNA amplification strategies, such as the PCR-based diagnostic assays that are used to diagnose COVID-19 and many other diseases, rely on enzymes that can be expensive, and are often stable in very specific storage conditions, so being able to function without them could make a diagnostic assay much more robust, cheaper, and thus accessible.

What real-world problem can this solve?

The COVID-19 pandemic has intimately demonstrated the importance of point-of-care diagnostics to the public. My team is trying to build a diagnostics platform technology using crisscross assembly that can be used at the point-of-care to detect various pathogens as part of a Wyss Validation Project. Our tool would allow people to test themselves at home or in another regular setting for some diseases. Not only would this be helpful in the United States and Europe, but it would be extremely beneficial for low-resource communities where there is less access to clinical diagnostics.

How did you end up in this field and in William Shih’s lab at the Wyss?

Dionis Minev on Using Crisscross Assembly for Diagnostics
An aerial view of the 2014 Wyss Retreat, where Dionis was inspired to pursue his Ph.D. at Harvard University and the Wyss. Credit: Wyss Institute at Harvard University

I decided to study mechanical engineering during my undergraduate studies because I was interested in engineering in general but in particular, I was curious to learn how to design and construct machinery that we use or is around us every day. I did my master’s in medical engineering, which mainly focused on medical devices, such as prosthetics and imaging equipment. During that time, I had the opportunity to get involved in wet lab work and be a Visiting Graduate Student in Neel Joshi’s group at the Wyss. When I got here, I saw this amazing variety of people from different backgrounds working on a multitude of problems. I was especially fascinated by synthetic biology. I’d heard of synthetic biology and read papers about what it could do before, but I didn’t understand the breadth of the field until I got to the Wyss. As a mechanical engineer, I was used to designing and building macro-scale machines. I was fascinated by the idea that with synthetic biology, you could use parts of biology to rationally build machines on the nano and micron-scale. Early on during my time at the Wyss I was invited to attend the 2014 Wyss Retreat. At this event, I looked around and was in awe of the diversity of people and projects. There was such a high concentration of impactful work on display in the space. Attending the retreat solidified my desire to try and return to Harvard and the Wyss for my Ph.D.

I got into Harvard and soon took a course with William Shih and Peng Yin on synthetic biology that was focused on DNA self-assembly. The idea of designing nanostructures with DNA was fascinating. I had the opportunity to talk to William about my ideas, which was really exciting and gave me good insight into how he interacts with people in his lab. He was so motivational. At the end of the course, I asked to do a rotation in his lab, meaning I would join for three to four months to see if it was a good fit. Quickly, I knew I wanted to formally join the Shih lab and it’s been a great experience.

What continues to motivate you?

I think what motivates me the most is the progress we’ve made. I initially started graduate school not even knowing if we could get crisscross assembly to work. I got my Ph.D. last year and stayed on as a postdoc with the intention to finish our work started during my graduate studies and further work on applying crisscross as a biosensor. Now, not only do we know it works, but we’ve also written a paper and published it. Moving forward, we get to take it even further and explore the applications of this technology, like diagnostics and building larger structures. Taking something that was a proof-of-principle and turning it into an application that makes a real-world impact motivates me.

What excites you the most about your work?

I’m really excited by the amount of creativity that goes into designing new DNA structures. This all goes back to what initially fascinated me when I was in college – designing and building. Those are huge elements in this work. The fact that I can be in a research setting where I’m exploring interesting and important questions while still having fun with it is exhilarating. There’s also a lot of freedom to think about your own ideas, just like in the visual arts, like painting and drawing, or in engineering. DNA nanotechnology is a relatively young field, so there’s a lot of room to innovate and build new things.

Most of the time we don’t really know the answer, but it’s fun to give it a shot and try to come up with a solution. For every failure in the lab, I get to wake up the next day and try again! Eventually, something does work.

Dionis Minev
Dionis Minev on Using Crisscross Assembly for Diagnostics
Dionis and his colleague Anastasia Ershova working through the design process, which Dionis finds incredibly exciting. Credit: Wyss Institute at Harvard University

Using the design process to try to find solutions to various challenges is also very exciting. You can wake up one morning, have an idea about how to solve a problem, try it, and if it doesn’t work, you try again. Most of the time we don’t really know the answer, but it’s fun to give it a shot and try to come up with a solution. That process of brainstorming with others or sitting at home and drawing something on my iPad is what I find so exciting. For every failure in the lab, I get to wake up the next day and try again! Eventually, something does work. You go from designing something to ordering it, testing it, and then seeing the results. In my field, the whole cycle is pretty short, because we design a lot of the structures on a computer using CAD-like software. When everything comes together and you see the results of that creative design process on the transmission electron microscope, it’s amazing.

What are some of the challenges that you face?

The challenge is that there are still a lot of things we don’t understand about the crisscross assembly process. Now that we’ve gotten this first piece of the puzzle, making the 1D linear filaments and tubes, we’re trying to make more complicated 2D and 3D structures and introduce other mechanisms that further diversify the features crisscross can incorporate. It’s a challenge to find clever ways to make the system work or understand how it’s functioning so we can alter it and make it do what we want.

How have the unique aspects of the Wyss impacted your work?

The Wyss is unlike anything I’ve experienced anywhere else. As I said, my first Wyss Retreat in 2014 was the defining moment that made me decide to apply to Harvard for my Ph.D., in the hopes I could come back to the Wyss in particular. That in and of itself has had a huge impact on me. At the time, I thought the whole premise of Biologically Inspired Engineering was fascinating. I loved that there was such a diversity of projects and groups working on different things.

I’ve always been fascinated with scientific translation but wouldn’t have even known where to start before I got to the Wyss. Valuable resources are more tightly integrated into the Wyss’ structure than they are at any other place I’ve seen, which means the process works as rapidly and as efficiently as possible.

Dionis Minev
Dionis Minev on Using Crisscross Assembly for Diagnostics
One of Dionis’ paintings, acrylic and oil on canvas, 36” × 48” Credit: Dionis Minev

Since I’ve been fortunate enough to come back and join William’s lab, I’ve benefited from the extra talks, the community events, and the open collaborative atmosphere. There is such a high concentration of labs doing really high-impact work close together, and if you’d like to, you can chat with anyone. That’s really unique and has led to some scientific collaborations that wouldn’t have happened otherwise. Now, I’m also starting to benefit from the Wyss’ commitment to teaching entrepreneurship. I’ve always been fascinated with scientific translation but wouldn’t have even known where to start before I got to the Wyss. When you have something interesting and have proven that it could make an impact in the world, like we have done with our Validation Project, the Wyss has a process in place for allocating resources to help advance projects towards commercialization, including additional funding, a dedicated Business Development Lead, and assistance with filing your IP. These resources are more tightly integrated into the Wyss’ structure than they are at any other place I’ve seen, which means the process works as rapidly and as efficiently as possible. It’s just incredible.

What do you like to do outside of work when you’re not social distancing?

I like to go outside and bike or run. I enjoy traveling. In terms of hobbies, I enjoy painting, drawing, and digital art. I think that the creative design process involved in my scientific work is directly related to the creative process used to make art. That’s probably why I love both of them so much.

What’s something unique about you that someone wouldn’t know from your resume?

I speak North Macedonian, and I can read and write it. It’s a Slavic language that’s closely related to Bulgarian, Croatian, and Serbian with a similar alphabet to Russian Cyrillic script. My dad is from North Macedonia, and growing up in Germany, he was adamant that I learn this language. I didn’t appreciate it then, but now I’m really glad that I know it.

What does it feel like to be working on cutting-edge technology that has the potential to have a real and significant impact on people’s lives and society?

Even making a small contribution to something that could impact a lot of people feels amazing, especially if you’re having fun while you’re doing it. I can’t imagine a better way to spend my time.

Dionis Minev

I’m humbled. I feel like I’m in a privileged position. When I applied and got into school at Harvard, I was amazed. Back when I was starting college in Germany, I never thought I’d be at a place like the Wyss. Achieving that was amazing. I’m so happy that I can contribute to some of these cutting-edge technologies, in whatever way that may be. Even making a small contribution to something that could impact a lot of people feels amazing, especially if you’re having fun while you’re doing it. I can’t imagine a better way to spend my time.

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