Hemdeep (00:12)
Welcome to Big Ideas in Microscale, the podcast where we explore groundbreaking research happening at the microscale where micro innovations makes a big impact. We're excited to showcase the incredible work being done by our users from around the world who are pushing the boundaries of microfluidics, lab on a chip, organ on a chip and beyond. Through these conversations, we hope to learn from their experiences, uncover their insight and bring their big ideas to wider audience. So whether in a lab,

Hemdeep (00:41)
On the go, or just curious about the future of microtechnology, join us as we dive into big ideas at Microscale.

Robin (00:53)
Welcome back to Big Ideas at Microscale.

Hemdeep (01:12)
I'm your host Hemdeep, co-founder of Creative Cat Works, Cat Works 3D, and Resin Works 3D.

Robin (01:17)
And I'm Robin, co-host as well as the technical writer within the marketing team.

Hemdeep (01:21)
In the last two episodes, we've had the pleasure of hearing from Jack Koch, an assistant professor at the Aquatic Germplasm and Genetic Resource Center at Louisiana State University Agricultural Center. Jack shared his fascinating journey from childhood fascination with marine life to his current research in cryopreservation and germplasm repository for aquatic species.

Robin (01:45)
We also discussed the vital role mentorship played in shaping his career and his passion for wildlife photography. Then last week, we explored the challenges of preserving genetic material of aquatic species like sperm, eggs, and embryos. Jack highlighted the gap in preservation efforts between terrestrial and aquatic species and discussed emerging cryopreservation techniques that are advancing the field. If you don't know what we're talking about, go back and listen to our previous two episodes.

Hemdeep (02:14)
This week we're going to explore the exciting development Jack and his team have been working on to standardize chiropraservation across species. They've developed open hardware kits designed to make the process more affordable and accessible.

Robin (02:27)
version of these kits even encourages users to 3D print components themselves, allowing for further customizations which could revolutionize genetic preservation in aquatic species. Jack will also share how 3D printing is playing a crucial role in creating cost-effective devices like sperm counting chambers and tools to streamline cryopreservation techniques. So let's jump right in to big ideas at Microscale.

Hemdeep (02:52)
these kits and these open hardware has a series of different type of modules or devices. Can you describe some of them? There's one that I've read up. It's called the strand cryopreservation shuttle system or the single sperm counting chamber. How did you develop those devices in terms of the brainstorming or the ideas that were used to develop them? And then how did you get it to a commercial object at that point?

Jack (03:20)
Sure, so let's start with the single piece sperm counting chamber. And that evolved from a need to give people access to a way of counting sperm, finding out the concentration. So just like we talked about, knowing the concentration of the sperm, if it's modal, what the motility information is, those are also pieces of important quality information. There's some commercial.

Options out there, there's something called a hema cytometer, which was developed for counting blood cells. There's also something called a Mackler chamber, which was developed for counting sperm cells. Those costs like $300 and $700 each. And so if we can make something that's 3D printed and costs, you know, 10 cents, 20 cents, you know, a dollar that someone can print anywhere in the world that will allow them to count sperm repeatably and standardized across many laboratories.

Like that's, that's wonderful. And so that's where the single piece sperm counting chamber came from was this need to have some sort of device that was low cost accessible to people anywhere that could be packaged in a kit like this. The Strand cryopreservation shuttle system also came from a need that we recognized in trying to develop cryopreservation processes for aplasia. Before I talk about the shuttle system though,

When we work with outside groups for this project, for the Applesia project, we work really closely with the National Applesia Resource, which is based out of the University of Miami. And people, including ourselves, have created a device that we think is going to be useful for a community. You give it to them and they're like, we don't need this. Like that's not a problem. Or that doesn't fit into our workflow. Like that's going to cost us another person or.

And so we work really closely with these national centers in something called user centered design, where we form relationships with them. We talk to them, we ask them questions, we go see their facilities, we make observations. And we find out what types of hardware are needed to help them in their facilities, especially as they move to incorporate cryopreservation. So the Strand cryopreservation shuttle system is

perfect example of that. So we have many meetings with the folks at the National Applesia Resource. We've visited their facilities, we've made observations, and then we bring it back to the AGGRC. And while we're developing cryopreservation processes for Applesia, we keep in mind what their needs are, what the environment is. And that's what this came out of. So to give you guys a mental idea of how difficult

it is to work with plesia egg masses. Imagine you have that coffee straw and you have a little piece of wet noodle and you need to package that piece of wet noodle into the coffee straw repeatably over and over again.

Robin (06:29)
How many eggs do you need in the straw? Or noodles do you need in the straw?

Jack (06:33)
Let's say you want to put one noodle per straw, one one centimeter piece of noodle per straw, and you have 100 straws, and you have 10 minutes to do it. Like, it's just, it's not possible, right? And then if one one centimeter piece of noodle in a 20 centimeter straw, that seems like a huge waste of space, right? So wouldn't it be better if you could package multiple noodles in the straw? But now if I asked you to package three noodles in 100 straws in 10 minutes, you're like, Jack, 100 wasn't.

possible in the first place. Like you want me to do 300? Like no. So we created this device that put your egg strand pieces into a set cassette and you take this device and you just push down and you load 10 egg strands at a time into these cassettes, into these shuttles. And now, now I can do something with them. I can take these 10 and I can put them back in the seawater. I could load a hundred egg strands ready to go.

And now I'm not, I'm not on the clock, right? I haven't started my 10 minute timer. I can take those hundred strands that are packaged ready to go. But now I take my straws and I put them in something that has them in the same spacing as the shuttles. When my 10 minute timer is up, I take and I just snap 10 off at a time. And now I can get three noodles in each straw and my packaging efficiency goes through the roof. That's the basis of this sort of system.

is just efficiently sample handling and packaging things. We developed a way to seal those straws. Traditionally, you would use an ultrasonic sealer, which uses like high frequency vibrations to like melt the straw closed, but without heat. An ultrasonic straw sealer costs $10,000. And so in most parts of the world, including the United States, if people don't have access to that, they might take like a pair of

of tweezers and heat them up over a flame and use heat to seal their straws. But heat's not good for sperm. So you probably are affecting the sperm in some way sometimes. So we made something that we can 3D print. We have 3D printed ceiling balls. They interact with this system. So you have 10 straws that are spaced appropriately. You have 10 ceiling balls. So now you can seal 10 at a time. And again, rapidly increasing the time or decreasing the time that it takes to

fill, seal, and prepare them for crop reservation. And you're increasing access because these are all printed on a $300 consumer scale printer with basic resin. it's just, opens the door to so many possibilities, not only for Aplasia, but for other animals all around the world.

Robin (09:18)
So for a lot of these devices, did you immediately jump into using 3D printing as a means to fabricate these? Or were there experimentations with other fabrication methods?

Jack (09:31)
While I've been there, we've only used 3D.

Robin (09:34)
3D printing.

Hemdeep (09:36)
And what was the previous, before 3D printing, those cassettes that you mentioned or the straw that you mentioned, was there a tool set that was used for this or is it something like, is this all just a brainchild of necessity, I guess?

Jack (09:50)
Yeah, it came out of my brain one night.

Hemdeep (09:56)
Technobiologist.

Jack (09:58)
It goes back to the limits of 3D printing are your imagination because you can print pretty much whatever you want. It's incredible.

Robin (10:05)
But even then, I guess 3D printing is not at its peak yet in terms of technology. Do you find that there's still kind of limitations that 3D printing has with your research? Yeah. And what would be the main ones?

Jack (10:22)
For crop preservation, reducing the amount of thermal mass around your sample is very important. So if a 3D printer can only print something that is, let's say, one millimeter wide, let's say it can't print anything smaller than one millimeter, I have to have a one millimeter thing next to my sample. That one millimeter thing holds heat at some amount, right? And so if there was some way for me to cut it in half, so half a millimeter,

and still have the same structural integrity, like that'd be great. That's what we see a lot is just being able to reduce how much is there while still having the same structural integrity. This is when we bring in engineers, right? I am not an engineer, but this is when we bring in engineers and they say, yeah, you know, if you add some cross-links in the back, you get that rigidity back and you can reduce how much material you have. The technology is growing very rapidly for like metal 3D printing and polymer 3D printing.

especially on these smaller scales. And metal is great because it's a heat sink. So you can rapidly pull heat away from your sample much better than you can with a traditional resin. Incorporating things like graphene, different types of mixing resin with these different materials to increase its ability to cool samples quickly or its structural integrity. And then toxicity of resin.

is something that people think about a lot, or I guess toxicity of any sort of material, even the plastics that we use in the fused filament fabrication printing. You just have to do a test, right? And a lot of the times your sample may not be in contact with that device for more than a couple of seconds, or you're using a sub sample of your sample and you don't care if it ends up non-viable in the end.

You have your sample that you're freezing over here and maybe you're running a sample through a microfluidic device and it's just going to run through. You're going to do some sort of analysis in two seconds and then you don't care about that piece of sample anymore.

Robin (12:32)
So you said that the AGGRC has what, over 300 3D printers, right? Like that? And like, how does that?

Jack (12:40)
No, I think we might have just broken 100.

Robin (12:44)
sorry, I totally misheard that. I guess 100. But anyway, that's a lot of printers. how does that look like? Is this all in like one gigantic room or warehouse? how does that work?

Jack (12:59)
Well, you guys will have to come down to Baton Rouge at some point to come visit our center. But we have a printer farm, which is one big room with maybe 50 3D printers in it. And they're just on shelves, right? If you go look up on a printer farm online, that's what we have is a bunch of 3D printers on shelves. But we have printers in many other rooms. So I have a couple of 3D printers in my office. We have a training and prototyping space, which has about a dozen 3D printers in it.

And so we have this training and prototyping room where people learn to break printers. They learn how to fix them. They learn how to do their first print. They learn CAD, they learn slicing. And then we also have our industrial space, which is the other end of the building. And that has a room that has a six or eight 3d printers in them. And then another room that has a couple of resin printers and a polymer printer in it. So they're, they're spread out in different places. And it's usually based on like.

what level of 3D printer it is and then what its purpose is.

Robin (14:02)
So talk about the AGGRC a little bit more. Clearly 3D printing is very important for a lot of the research that's going on there. But maybe you could talk a bit more about like what the AGGRC kind of, you know, its mission essentially.

Jack (14:19)
Yeah. So I mentioned it earlier, but we help other people and communities around the world form germplasm repositories. And that includes thinking about the process of crop reservation, technology development. How do you bring other users in? Where you're going to store the samples, how you're going to track the samples, how many people you need, what type of equipment you need. So that's what we help people do for aquatic species specifically.

Robin (14:47)
You mentioned something about four pillars of the center. What is that, example?

Jack (14:51)
Yeah. So we support our mission through four major programs. First one is basic biological research. So we study animals to learn about their biology, reproductive biology, or evaluate how different chemicals interact with their germplasm to prepare them for crop preservation. We have a technology development program and that's sort of split into open hardware and industrial processing. We have outreach.

because it's super important to talk to people in other industries, other universities, other levels of education to not only improve how we think, but also let people know that these technologies and resources are out there. And then the last one is industrial scale processing. We could run a company out of our building. We have the equipment necessary to double or triple.

the total global aquatic species, germplasm samples, just in a couple of weeks. We could double or triple what's out there. Obviously we'd need enough sperm to do the freezing of, but we have some high level equipment that we do use for certain projects and we train people on that sort of equipment as well. Because at some point people want to go from 10 to a hundred samples, a hundred to a million samples and...

There's different levels of scale that need to be recognized and thought about because 10 samples, you know, you might be able to hire one person, a hundred samples. Maybe you need two people, but when you get to a million samples, like you can't just hire more people. You need some technology to help you accomplish some of those tasks.

Hemdeep (16:34)
The center has grown significantly, it sounds like, and it's quite important for this repository. I'm wondering, how did the growth of the center sort of follow the trend line of the adaptation of 3D printing? Did it grow hand in hand? Was it a necessity? Or if you were to remove 3D printing from the equation, was the center had its own inertia that it was going to meet these mandates that it had set out for?

Jack (17:01)
The center definitely would have continued without the 3D printing intervention, but the growth has been much more rapid and has allowed us to address more communities, more diverse communities in different places than if we didn't have 3D printing. There are many commercial technologies out there for crowd preservation that we probably could have used. When 3D printers became accessible to more people, I don't...

I'm going to put a number out here, but I don't remember when the first 3d printer was purchased at the center. Let's say it cost a thousand dollars. And that was like a big turning point because they used to be a lot more than that. They 3d printed something ran down the hall and dunked it in liquid nitrogen and to see if it would still stretch and it stretched. Right. You know, you usually you put something in liquid nitrogen and you try to like end it and just shut it. And that was, that was a huge realization.

Hemdeep (17:56)
better.

Jack (18:00)
to the people at the center at that time was that, wow, we can make devices that are gonna be near and in contact with liquid nitrogen and we can bring it to more people and make standardization and access just has driven or helped direct where we go along our mission.

Robin (18:20)
community that you're trying to bring this to, do you find that they already have experience with 3D printing or is it something that you kind of teach them about?

Jack (18:32)
It's a mix. are some people who know about 3D printing, but have never done it. There are people who have like printed one or two things. There's people who have never heard of it or seen a 3D printer before, but more and more local libraries have 3D printers. A lot of universities have 3D printing facilities that their students are learning on. Any university with an engineering program.

I'd say typically has some sort of 3D printers. There's a lot of groups of students that are forming like maker clubs that have 3D printers. So more and more people know about it. We don't necessarily like teaching people about 3D printing. We like helping them and making the environment such that they can succeed in 3D printing because we find a lot of power in having people learn on their own.

while providing support for them. Like, you know, if you have questions, like we're here for you, but we're not going to tell you what buttons to press. We can help guide you, but we're not going to like give you some recipe or some formula because that's not going to work every single time. Right. 3D printing, you need different parameters for different things. so having an innate understanding or a learned understanding of what those parameters mean. We can't help every single person either. Right.

We need it to be a network approach where we help one person, they help two people, and eventually everyone has the same background.

Robin (20:06)
And guess that's why it's so important for your team to like bring this more to the commercial scale as opposed to industrial 3D printers. And I think that's what your latest paper was about.

Jack (20:19)
Mm-hmm. Yeah.

Robin (20:20)
What sort of devices did you kind of test between the two types of 3D printers essentially? And like, what were the results essentially?

Jack (20:32)
Yeah. So that goes back to the single piece sperm counting chamber. type of device. One of the reasons why devices like hemocytometers and Mackler chambers are so expensive is because they're highly precise devices. The reason you can calculate concentration of a sample on a device like that is because the volume underneath the cover slide is very precise. It's exact.

And that has to do with the way it's manufactured. The weight of the coverslip on a hemostatometer is very precise. You can't just use any run of the mill coverslip on a hemostatometer. They have special coverslips that are weighted properly so that the volume of the sample that you're counting is known, which allows you to do an accurate concentration. So when you're trying to 3D print something, there's a level of tolerance. There's some tolerance that you have to achieve.

to guarantee or ensure that the volume underneath your cover slide is within some sort of known region so that when you count the concentration, it's real. And so printing something like that on a consumer printer, it may be difficult, right? And so, you know, that's, we have a CADworks printer, right? And we bought it through a seed grant.

that Louisiana State University Agricultural Center provided to us. The purpose of that paper that you were referencing was to see what the capabilities of consumer level and in this case, industrial or commercial level printers are. We didn't want to compare them because it would be an unfair comparison. If this is our scale of sizes that we're interested in, we wanted to know maybe consumer scale covers this much.

And industrial or commercial scale covers that much. And there's some overlap in the middle. And that was what was super important to us. Not all of the sperm that we work with is very, very tiny. And so the consumer scale printers are completely capable of printing some devices for these larger sperm. I'll send you guys a picture of a variety of sperm, but ambistumous sperm are absolutely massive. We sort of call them like aircraft carriers. are.

huge compared to human sperm, even frog sperm. They're just, massive. so printing on a consumer level printer is relatively easy when you're trying to achieve sizes for that scale. For zebrafish, which have sperm that are like maybe two microns big, you're going to be hard pressed or you may be hard pressed to use a consumer scale printer to create a device to count concentration for you. And that is where

having something like the industrial scale printer might, or commercial scale printer, might actually be needed because the size of those sperm are just so small that you have to create a gap that's small enough that you don't have a bunch of sperm stacked up on each other. So that was the purpose of the paper, was just to find where they overlap and so that we can make informed decisions on when we create new devices in the future, knowing, ⁓ okay, we can create this on a consumer scale.

It's going to be accessible to a lot of people around the world, or we're creating this device and it's just, it's small, but it's not soft photolithography small. And we can use a commercial scale printer and either print devices for people or going back to that molding capability, print molds for people and let them pour a device themselves.

Hemdeep (24:15)
I think you probably have a significant amount of experience based on the wide range of printers that you've been using and the type of work you've been using. What do you sort of see in the future for yourself, your team? Are there any new sort of features that you guys are hoping to develop or developing this repository?

Jack (24:35)
think having standardized devices are one place that we're really trying to push and making sure that the communities that are using them, that they're serving the needs that the communities want. In terms of the printers themselves, access to different resins, all of the printers that we have in our building are open material printers.

which means that we can use any resin on the market. We've just may need to dial in the settings a little bit, which again goes back to our philosophy of open hardware, right? We don't want to be stuck in one ecosystem. Just being able to control every aspect of the printer is great. And I think having more companies that allow you to tweak your printer to your liking is something that would be good.

at an even higher level scale than our center, right? That's like an industry level thing. But I understand that, you know, companies need to, they need to make a buck. And so a lot of those printers that are closed material ecosystems, those companies put a lot of money into like dialing those printers in and making them work exactly as you expect. But there's also plenty of open material printers, both in fuse filament fabrication and resin that.

They just work out of the box because they're just workhorses, right?

Hemdeep (26:03)
What is next for the germplasm repository? What do you envision in terms of your work? And I guess in any institution or team, there's always a changing mission statement, you know, that when they sort of start attaining some of the goals or the milestones that they had set out, they start building brand new ones. And what would be the next ones look like?

Jack (26:26)
Our mission won't really change that much because the number of communities and number of species is just so large that we could spend decades working on it. But where we may change is in how we enable other communities and the speed that we enable other communities to create their own processes and repositories.

to address their own species, right? So that's like a distributed workflow where rather than us being the only group cryopreserving species, helping to accelerate the number of groups that are cryopreserving different species because we have expertise in a lot of different things, but there's not enough time in the day or in our lives that we can do that. So we need more people. So getting the word out, doing more outreach.

think we sort of envision every middle elementary high school, every college student knows how to 3D print. know, they're familiar with CAD, whether they're a accounting major, a psychology major, a nursing major, a biology major, an engineering major, because it's really a life skill at this point.

Robin (27:43)
you're trying to build this wider community. Where is kind of like your home base? Is there is on your website? So say I was a aquatic researcher for a specific species and I want to learn more about your cryokits or anything like that. Do I need to come across you based off your website or are you on a forum or some other platform where AGGRC can be visible?

Jack (28:08)
Yeah. So aggrc.com is probably the best place to go. We do have a community forum that we can give you an invite link to that if people are interested in communicating with us and with other people in our community. Every time we publish a new open hardware device, it gets posted on there. And then we also have a variety of different social media. So think we're on LinkedIn, Facebook, X, Blue Sky, Instagram.

Right. And so I think there's messaging on those so you could get in touch with someone here.

Hemdeep (28:43)
All right. That was a fantastic conversation, Jack. I am so glad that we had this opportunity. We're definitely going to be posting all the links that and the articles and the images that we discussed during this podcast on the show notes. Jack, why don't you take a moment to give shout outs to all of those that have made your work and research possible.

Jack (29:08)
Cool. Thanks, Hemdeep. Yeah. Thank you so much to the Louisiana State University Agricultural Center. Provided some seed funding for some work that was very relevant to this conversation. A big shout out to the National Institutes of Health Office of Research Infrastructure Programs. They help support us in our biomedical work, helping the national centers across the United States. And those include the National Pleasure Resource, the Xenopus,

National Resource Center, the Ambistema Genetic Stock Center, the Zephyrus Genetic Stock Center, and the Zebrafish International Resource Center. Also thank you to all of our collaborators at Louisiana State University, Dr. Monroe's group, collaborators around the world, and other funding sources from USDA, National Science Foundation. Really appreciative of everything that

those groups do for us and help us with. We love helping people all around the world to protect the genetic resources of aquatic species.

Hemdeep (30:17)
Amazing. So what we'll do is that we'll add the links to all of those teams as well. I'm so happy that we had this chance to speak and I think you guys are doing some amazing work and I look forward to seeing some of the more interesting work as a techno biologist. I certainly am interested in seeing what we do next. Thank you very much.

Jack (30:42)
Thanks, Hemi. Thanks, Robin.

Robin (30:44)
And with that, that's a wrap on our series of Jack Koch here on Big Ideas at Microscale. Over the past few weeks, we've explored the world of genetic preservation and cryobiology research. Jack has taken us on a journey through his career, from his early curiosity about marine life to the groundbreaking work he's doing at the AGGRC, the Aquatic Germplasm and Genetic Resource Center.

Hemdeep (31:10)
We've discussed the growing need for germplasm repository to support the breeding and conservation efforts of aquatic species and the challenges that they face due to complex reproductive biologizes. We then discussed emerging cryopreservation techniques and how they are helping to overcome these hurdles.

Robin (31:29)
We dove into the use of 3D printing open hardware and how it is helping to make cryopreservation tools more accessible, affordable, and customizable for researchers around the world. Jack and the AGGRC's work is an exciting glimpse into how 3D printing technology is driving forward solutions to some of the most pressing challenges in conservation and genetic diversity.

Hemdeep (31:52)
We hope you've enjoyed this deep dive and the vital work being done to protect aquatic species. Our next guests to be featured on the podcast are assistant professor Veronica Mendez and master student Noah Franco from the University of Waterloo.

Robin (32:07)
This episode is particularly exciting for us as they have spent the past several months limit testing and validating CADWORK3D's new 3D printing material, the Cyto-clear photopolymer resin. Here's a little snippet of how that conversation went.

Hemdeep (32:38)
So there was a material that my brother was sort of manufacturing and he goes, look, you know what? This thing has got amazing components and you know how we have to use IPA to clean our models. we were on this large workbench and right beside a wall and the workbench itself had IPA sort of, you know, there's a very thin layer of IPA on the workbench. We didn't notice it, but my brother wanted to sort of take a model and light it.

just to sort of showcase and he didn't realize it but a drop of it fell on the bench and we didn't notice anything until this blue flame just went right up the wall and we're like ⁓

Robin (33:22)
Was this at your mother's house or was this going into office?

Hemdeep (33:25)
No, no,

no, this was at the office. My mother's house, we burnt her kitchen.

Robin (33:33)
Thanks for tuning in to Big Ideas on Microscale. If you enjoyed the episode, make sure to follow us and stay up to date. You can listen on Apple Podcasts and Spotify, or watch the full video on YouTube. You can also follow us for more updates and behind the scenes content on LinkedIn, Instagram, Blue Sky, and X. We're Cadworx3D across the board. Let's spell C-A-D-W-O-R-K-S-3D.

for show notes, paper references, and bonus resources on today's topic, visit our website, catworks3d.com. That's spelled C-A-D-W-O-R-K-S 3D.com.

Hemdeep (34:17)
Thank you for tuning in and as always stay curious, keep exploring and never stop asking the big questions that are shaping our world. Whether you're in a lab or on the go, we're just curious about the...