The Conduct Science Podcast: Endangered Species

00:00 – Intro

02:05 – Announcement

06:30 – News

17:45 – Not so fun facts

20:35 – What are endangered species

25:00 – IUCN Red List

44:16 – How animals become endangered?

59:16 – Conservation strategies

1:09:00 – What YOU can do

1:11:45 – Ending and outro

You can listen to The Conduct Science Podcast by using the player above, searching for “The Conduct Science Podcast” on any place you listen to your podcasts, using any of the links below, or you can download it HERE!

Links from the show:

• Revenge Bedtime Procrastination
• IUCN Red List
• IUCN Red List Criteria
• Best ways to conserve
• Scientists debate conservation triage
• Instagram
• Twitter

This week on The Conduct Science Podcast, Mitch and Tom take a look at endangered species! What and why are species considered endangered and who gets to make that decision? The IUCN red list denotes those species that we should pay close attention to, but how do species make it on and off the list, and how do governments use that list? Mitch takes a look at the main reasons that cause population decline and Tom takes a look at the different strategies that can be used to help conserve populations. Do you think that its ethical to abandon some species to extinction in an effort to save others? Join the debate with #ConductScience. Music: WhiteBatAudio.com

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Figure 1:  Example of an electropherogram from the sequencing of an unknown DNA fragment by automated capillary electrophoresis.

High Surface-To-Volume in CE Increases the Efficiency in Heat Dissipation

Capillary electrophoresis uses the same principles as other types of electrophoresis and is subjected to Joule heating, the conversion of electrical energy into thermal energy or heat.^[1,2] In electrophoresis, it occurs when the current meets the resistance of the components in the system.

The generated electrical energy is subsequently converted into thermal energy, which heats the fluid in the electrophoresis system.^[2,3] The heated fluid diffuses from its source and is replaced by the cooler fluid from the other part of the system. The replacing fluid is heated and subsequently displaced by cooler fluid, creating a cycle of convection currents that continue until heat is evenly distributed throughout the system.^[3]

Excessive heat can damage heat-sensitive samples, disrupt the separation and destabilize the system. As a result, the application of high power to the system, which can accelerate the ion mobility and decrease the separation time is generally restrained in electrophoresis.^[1]

However, in CE, electrophoresis is performed in a capillary tube, this affords the system with a high surface-to-volume environment. Thus, the distribution and subsequent dissipation of thermal energy are more efficient. It enables CE to be performed in a high electrical power field, and it reduces the separation time without having to compensate for the resolution.^[3]

Electroendosmosis Flow Adds to The Speed and Versatility Of CE

Apart from the high surface-to-volume ratio, the capillary also affords CE with a moving force that drives the ion’s migration, in addition to the electrical force. On the interior surface of the capillary, the ionized solution causes the surface to be ionized, and the solvated ions are adsorbed onto the interior wall of the capillary. The ions on the stern layer, the interior wall of the capillary, attract the ions of the opposite charge in the solution to build up nearby, forming a diffuse layer. Thus, an electrical double layer is formed at the interface of the surface and the solution.

When voltage is applied, the ions at the diffuse layer are pulled towards the electrode of the opposite charge, creating an electroendosmosis flow, also termed electroosmotic flow (EOF) that tows other solvated ions to the same destination, whether their net charge is the same or opposite. EOF is considered to be a hallmark of CE, which further reduces the separation time and allows all ion species, anions, cations and neutrals, to be separated in a single electrophoresis run.^[3,4]

Instrumentation

Figure 2: Diagram showing the components of an electrophoresis unit in capillary electrophoresis.

CE requires a high-voltage power supply, an electrophoresis unit, and a detector where the result of the electrophoresis is recorded. The basic components in the electrophoresis unit include a cathode, an anode, and a long capillary tube that is in contact with the electrophoresis solution buffer. Nowadays, CE is automated and requires very little monitoring, and the basic components in CE have been modified to accommodate it. For example, buffer reservoirs have been added, each sample is placed in a vial in an autosampler that automatically feeds each sample to the analysis.^[4]

From an operational perspective, CE instrumentation can be viewed as follows:^[3]

   1.        Sample injection

Since the capillary is narrow and small, only a small amount of sample is required for CE. Excessive amounts of samples are not only wasteful but may also interfere with the resulting electropherogram, which may be misleading. To ensure good sample injection quality, two methods are used:

• Hydrodynamic injection: This is the application of pressure at the injection end of the capillary and the use of vacuum or siphon at the other end. Typically, the capillary is first filled with the separation buffer before the sample is injected into the capillary. The amount of the sample injected is controlled via the applied pressure and the duration of the injection.
• Electrokinetic injection or electromigration is the application of the voltage across both ends of the capillary. However, the injection end of the capillary is in contact with the sample before the voltage is applied to the system. Here, ions enter the capillary-based on their electrical mobility and subsequent EOF.^[3]

   2.        Separation in the capillary

In CE, the capillary is considered the most important element of the system since this is where the separation takes place. Fused silica is often used in CE because it’s chemically and electrically inert, transparent and flexible, making it the ideal capillary.^[3,4]

A part of the capillary where the separation takes place, including the detection window, is typically packed in a cartridge, to ensure that the temperature and the condition of the system are maintained throughout the separation, and that external changes will not destabilize the system or interfere with the electrophoresis. Most capillaries are coated with a layer of protective polymers to ease the handling. Capillaries should be regularly conditioned to ensure that any residual ions on the surface wall are removed and will not interfere with the electrophoresis.^[3]

   3.        Detection

The method of detection in CE is determinant in the overall sensitivity of the analysis.

Popular detection methods for CE are:^[3]

• UV-absorption is the most widely-used detection method in CE due to its universal nature. UV-absorption is based on the absorbance properties of the electrophoresed component and rarely requires sample modification. It is estimated that the detection limit of UV-absorption is about 10^-13-10^-15 moles in mass or 10^-5-10^-8 molars in concentration.^[3]
• Fluorescence and laser-induced fluorescence (LIF) are popular for works that require high sensitivity. Both methods are based on the emission of light following an excitation. It is estimated that LIF has a detection limit range of 10^-18-10^-20 moles or 10^-14-10^-16 molars, while fluorescence is 10^-15-10^-11 moles or 10^-7-10^-9 molars. Both methods require some form of sample modification or derivatization, which is the main disadvantage because not all samples can be modified or derivatized.^[3]

Apart from the main instrumentation, other devices can be added to the CE. For example, a fraction collector can be added to collect fractions of samples for reanalysis, or a mass spectrometer can be connected for structural identification and characterization.

Modes of Operation in Capillary Electrophoresis

CE is regarded as the most versatile electrophoresis-based technique because it can be easily adapted without a need to change the instrumentation. Instead, it only requires small modifications in the electrophoresis system and alteration in certain operating parameters. For example, the interior capillary wall can be coated or uncoated to accommodate the nature of molecules in the sample, or certain operating parameters can be adjusted to increase the resolution or decrease the separation time.

The following are popular modes of operation in CE:

1.   Capillary zone electrophoresis (CZE)

Capillary zone electrophoresis (CZE) is a form of zone electrophoresis and is regarded as the basic mode of operation in CE. In CZE, the capillary is filled with a solution buffer and molecules are separated in the free solution in the capillary. Ionized molecules migrate into distinct zones based on their electrical mobility.

CZE has been used to analyze amino acids, peptides, proteins and other ionizable molecules, including the detection of drugs and drug metabolites in a forensic application. It’s also used in the separation and characterization of polar, non-volatile compounds from environmental samples. ^[3,4]

2.   Capillary gel electrophoresis (CGE)

Capillary gel electrophoresis (CGE) is the embodiment of gel electrophoresis in CE. In CGE, fluid crosslinking or linear polymers are filled into the capillary, samples are injected using electrokinesis and separated by size and charge. While a support matrix is not necessary for CE, the gel in CGE provides a molecular sieve which adds to the resolving power of the technique.^[3,5]

CGE has been applied to the analysis of nucleic acids, DNA fragments and protein. Its high resolution, sensitivity, reproducibility and automated nature has made it the technique of choice for Sanger sequencing and for forensic DNA profiling.^[3]

3.   Micellar electrokinetic capillary chromatography (MEKC)

Micellar electrokinetic capillary chromatography (MEKC) is the use of surfactant in the running buffer above the critical micelle concentration. The added surfactant induces the formation of micelles in the electrophoresis system. The interaction between ionized molecules in the sample and the micelles affects the separation time, enabling positively, negatively, and neutrally-charged ions to be eluted at different retention time.^[3,4]

Since MEKC can be used to separate and obtain all ion species, it has been used for a wide range of molecules, including forensic analysis of illicit compounds.^[3]

4.   Capillary isoelectric focusing (CIF or CIEF)

Capillary isoelectric focusing (CIF) is the separation of amphoteric molecules such as peptides and proteins based on their isoelectric point (pI) using CE. Here, the acidic buffer is placed at the anode and the basic buffer at the cathode. The capillary is filled with a mixture of the running buffer and amphoteric molecules before an electrical field is applied to establish a pH gradient in the capillary. Amphoteric molecules migrate through the gradient until they reach the point where the pH is equal to their pI. Detection is then performed when all molecules have successfully migrated.^[3]

In Conclusion

All in all, capillary electrophoresis is a versatile electrophoresis technique that uses a capillary to single out molecules in the sample of interest. Despite the need for sophisticated instrumentation, the sensitivity, speed and automated nature of CE enable it to accommodate high-throughput analysis, without sacrificing the resolution, making it an all-around tool that applies to both research and routine works.

References

1. Westermeier, R., Gronau, S., Becket, P., Buelles, J., Schickle, H., & Theßeling, G. (2005). Electrophoresis in Practice: A Guide to Methods and Applications of DNA and Protein Separations (4th, revised ed.). Wiley-VCH Verlag
2. Jorgenson, J. W. (1986). Electrophoresis. Analytical Chemistry, 58(7), 743A-760A. https://doi.org/10.1021/ac00298a001
3. Heiger, D. (2000). High Performance Capillary Electrophoresis: An introduction. Agilent Technologies.
4. Capillary Electrophoresis. (2020, August 16). Retrieved March 10, 2021, from https://chem.libretexts.org/@go/page/294
5. Xu, A. (2008). Development in electrophoresis: instrumentation for two-dimensional gel electrophoresis of protein separation and application of capillary electrophoresis in micro-bioanalysis (Iowa State University). Retrieved from https://lib.dr.iastate.edu/rtd/15688

Hello! Who are you and what are you working on?

I am a social worker in the San Francisco Bay Area working at the intersection of climate, health and social issues. My team and I recently finished up at qualitative research study looking at the health and social impacts of the recent wildfires that we have had in California. In addition to looking at how these fires are impacting health and well-being in the surrounding communities, we also looked at the perceived effectiveness of the response and recovery resources that survivors utilize after the wildfire.

                       Fig 1. 2017 North Bay Fires Burn Area (Tubbs, Atlas & Nuns).

                                   Fig 2. 2017 Thomas Fire Burn Area.

What’s your backstory and how did you come up with the idea?

I graduated with my Masters in Social Welfare from UC Berkeley in 2019. I had always been interested in how our social environment impacts health outcomes, which is what led me to go back to school. While I was at Berkeley, Northern California saw two of its most destructive wildfire seasons on record. I kept thinking about what short and long-term impacts these fires would have on various social issues that I was learning about in school–mental health, housing, substance use, etc. As social work students, we also think a lot about the effectiveness and sustainability of health and social programs, so I was also interested in the role of response and recovery resources in the aftermath of wildfires. I teamed up with a professor of mine, Dr. Rohini Haar at Berkeley’s School of Public health, who is an emergency room physician, and together we decided to conduct a research study on the health and social impacts of climate change with a focus on the role of response and recovery resources.

                                     Fig 3. 2018 Camp Fire Burn Area.

                                     Fig 4. 2018 Woolsey Fire Burn Area.

Please describe the process of learning, iterating, and creating the project

We read a lot of research articles about the health and social impacts of wildfires and natural disasters prior to beginning our own study. We found many articles discussing some of the immediate, physical health impacts of wildfires, such as issues related to smoke inhalation. But we found less articles that talked about long-term health and social impacts, so we knew we wanted to make sure we asked participants about those outcomes. We also read a lot of articles that talked about housing and employment issues quantitatively, such as how many homes burned, or how the wildfire impacted local employment rates. But we were really interested in how wildfires impact those same social issues qualitatively. What kind of impacts do wildfires have on housing and employment that cannot be captured in numbers? What we found through our qualitative research was a series of what we called  “ripple impacts” in the communities surrounding the wildfire.

Please describe the process of launching the project

First we submitted a proposal to our University’s IRB. Once that was approved, we began recruiting participants to be interviewed for our study. We sent out some recruitment emails on health and social service professional listservs asking. We also attended community meetings in disaster impacted areas in Northern California to recruit participants and to learn more about the long-term impact that the fires were having on the community. We conducted interviews for four months in late 2019 and then began the process of analyzing our data and writing up our study.

Since launch, what has worked to make your project grow?

It’s helpful to send your work around to your personal and professional networks to increase exposure and, hopefully, impact. We hope to be able to continue to do more research on the health and social impacts of climate issues going forward!

How is everything going nowadays, and what are your plans for the future?

Our research study was published a few weeks ago. It is always nice to finish a project you have worked hard on and be able to send it around to family, friends and colleagues. Next, we hope to continue to conduct more studies on the health and social impacts of the California wildfires. There are so many ways that these disasters disrupt the surrounding community and it is critical that we have an understanding of the extent of these issues and how social programs can better meet the needs of survivors. But this is not just a California issue or a wildfire issue– climate change is threatening the safety and wellbeing of communities all over the world. It is critical, at this moment, that we gather more information both about how we can prevent the climate crisis from worsening, and about how we can support the individuals and communities that are most vulnerable to the impacts of climate change.

Through your science, have you learned anything particularly helpful or advantageous?

Our research has taught us a lot about what we call the “ripple impacts” of wildfires and other disasters. We already know many of the immediate health and social issues that result from wildfires. For example, smoke inhalation issues or people being displaced because their homes burn down. But there are so many health and social issues that don’t begin to surface until weeks, months, or even years after the wildfire is over.

For example, wildfires create many complex, ripple impacts on housing accessibility. There is often an influx of people moving from wildfire-impacted areas into surrounding communities, which can cause the local rental market to essentially evaporate. Local rent prices can then skyrocket because the demand on housing is so high. These new rent prices can eventually displace people not initially impacted by the wildlife because they can no longer afford the price of housing in their area.

That is just one example. Our research found that for almost every immediate health or social issue that a wildfire creates, there are ripple impacts of that issue on the broader community.

We’d like to know more about your lifestyle and habits. Please let us know what is your morning routine like.

I usually get up around 7 a.m. or 7:30 a.m. and try to go outside for a walk first thing. I am lucky to live in Northern California where the weather is pretty nice year-round. I grew up in New England, so try to be extra appreciative of the good weather out here! Walking first thing also really helps to get the blood and creativity flowing. I find myself much more focused and present at work when I find time to walk in the mornings. After that I will make breakfast and coffee and get started on work, usually around 8:30 a.m. I usually listen to music while I work if I am not in meetings. It really helps me stay focused and energized.

And how does a typical day look for you?

My COVID remote working routine is obviously different than it was when I was going into my office. I get up in the morning, go for a walk, usually while listening to a podcast or music. Then I will come home, make coffee and breakfast and get started on work. Before I get started on tasks, I will try to plan out my day in a way that feels productive and realistic. I will make a list of everything I need to get done that day, and space out those tasks throughout the day so it doesn’t feel overwhelming. Then I will fill in the rest of the agenda with other, less urgents tasks, and breaks. It is important for me to build breaks into my agenda, otherwise, I forget to take them. Breaks are so crucial for productivity. I try to get outside a few times throughout the day.

What does your workstation look like?

During COVID, my workstation looks a lot like my kitchen table! I try to keep my table as clear and clean as possible to minimize distractions. It’s typically just my computer, a charger and some plants! I also try to keep my phone in another room so I am not distracted by that. I sit in my kitchen because it has the best light. I also have a big window looking out onto the Oakland hills, so I am very lucky!

What platform/tools do you use for your professional life?

We used the platform, Dedoose, to conduct our coding and qualitative analysis for our research paper, which we found to be extremely helpful and easy to use. Other than that, the tool we used the most for our research was Microsoft Word, where we drafted and redrafted our research study many, many times.

How do you stay up to date on News and resources?

I check a few new sources in the morning, and I might listen to a news podcast while I am on my working walk. I will spend another 15-20 minutes reading the news in the evening and that is usually it. I do not do Google alerts, push notifications, banners or even Twitter because I find that I can get easily overwhelmed with news that way. I think it’s good to stay informed, but to limit daily news intake so that I can actually take in what I am reading.

What have been the most influential podcasts, or other resources?

Amy Goodman and Democracy Now always does a really good about covering climate issues in a way that many other news sources do not. I especially appreciate the way that program covers climate disasters such as the California wildfires. Instead of only covering the damage and destruction that the disasters create, Democracy Now never fails to remind their listeners that nothing about these disasters are actually “natural.” They are created or exacerbated by climate change, which is caused by human activity such as the burning of greenhouse gases. It is important to remind the public during climate disasters, while their attention is piqued, that these are human-caused issues that require human-generated solutions, like clean energy usage.

What tools do you use in your personal life? Cook? Self-Care? Hobbies?

An important question! I try to get plenty of outdoor time and exercise. Even if it is cold or rainy I try to get outside. It really helps me clear my head and re-center. I also try and play tennis a few times a week with friends. That’s a great way to socialize and be active. It has also been a great, safe activity for COVID times.

I also like reading books that have nothing to do with my work. It’s really great to get lost in a story that totally takes you away from the stressors of everyday life.  Also, I am a very big advocate of lying on the couch and binge watching TV after a hard day. Sometimes that’s honestly the best thing for mental health!

Advice for other scientists who want to get started or are just starting out?

You don’t have to be in the hard sciences to research and write about climate issues. I’ve heard a lot of comments from friends and colleagues in the social sciences, for example, who believe that climate change is an issue to be researched and worked on by those with hard science degrees/ backgrounds. This couldn’t be less true! Climate change issues intersect with all kinds of different professions, so the approach to researching and solving climate issues needs to be multidisciplinary. Further, because climate issues amplify many pre-existing health and social conditions, I believe it is especially important to have social workers and healthcare workers involved in climate solutions.

Thank you very much for your time, Annie. Where can we go to learn more?

Our paper on the health and social impacts of the 2017/18 California wildfires was published a few weeks ago in PLOS one. Berkeley News also published a QandA about it.

The Conduct Science Podcast: Microfibers with Dr. Barbara Martinez and Krystle Moody Wood (Interview)

00:00 – Introduction

01:30 – Background

09:20 – What drew you into conservation?

15:40 – Microfibre pollution

27:45 – Microfibre innovation challenge

39:30 – How has conservation changed over time?

46:15 – What can we do?

52:35 – COVID pollution

54:15 – Cool new research

1:00:00 – Ending and outro

You can listen to The Conduct Science Podcast by using the player above, searching for “The Conduct Science Podcast” on any place you listen to your podcasts, using any of the links below, or you can download it HERE!

Links from the show:

• Microfiber Innovation Challenge website (due June 25)
• Microfiber Innovation Challenge contact: microfibers@conservationxlabs.org
• Conservation X Labs website
• The Trident Prize (due April 30)
• The Microfibre Consortium
• Conservation XLabs Socials:
  • Twitter
  • Facebook
  • Instagram
  • LinkedIn
• Materevolve Website
• Materevolve Contact: krystle@materevolve.com
• Article on UCSB/Nature Conservancy Study
• Fibershed Website: microfibers blog
• Experiential Learning Program: Building Regional Textile Systems for Healthy Soils and Healthy Seas
• NOAA + Materevolve California MIcrofiber Report: A Textile Perspective Report
• Materevolve Socials:
  • Instagram
  • LinkedIn

This time on The Conduct Science Podcast, Tom and Mitch are joined by two amazing guests: Dr. Barbara Martinez from Conservation XLabs and Krystle Moody Wood from Materevolve! Join them as they discuss the very important issue of microfibre and microplastic pollution! How is the fashion industry responsible for polluting the environment through the clothes we wear? what are they doing about it? What can we at home do to reduce the amount our clothes affect the environment around us? Think you have an innovative solution to this worldwide problem? Then check out https://conservationxlabs.com/microfibers for your chance to win part of a $650,000 prize! What do you have to lose? Nothing! So check it out. Music: WhiteBatAudio.com

Instagram     Twitter     Facebook

Thanks for Listening!

Feel free to share your thoughts on our Twitter here:  @ConductScience

Use #ConductScience on twitter to:

• Suggest a guest
• Suggest a topic
• Ask a question we can answer on the show
• Or to just get in touch!

Learn about our products:

• Visit our Lab
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The Conduct Science Podcast: Astrobiology

00:00 – Intro

05:15 – News

12:00 – What is astrobiology

19:45 – what astrobiologists do

33:45 – NASA’s astrobiology roadmap

38:45 – The search for life

45:30 – Is there life out there? The drake equation and Fermi paradox

1:03:00 – Ending and outro

You can listen to The Conduct Science Podcast by using the player above, searching for “The Conduct Science Podcast” on any place you listen to your podcasts, using any of the links below, or you can download it HERE!

Links from the show:

• Natural blue food coloring
• Muon G-2 experiment
• History of astrobiology
• Biosignatures
• The Fermi Paradox (video):
  • Part 1 – Where are all the aliens?
  • Part 2 – Solutions and ideas
• Instagram
• Twitter

This week on The Conduct Science Podcast, join Mitch and Tom as they join the search for life in the universe and explore astrobiology! This is an incredibly interesting scientific field gaining more and more traction in recent decades but what is it? What do astrobiologists do if we haven’t yet found life? What scientific fields are involved? How is the search for life carried out both within and outside of our solar system? NASA has an astrobiology roadmap but what if life is predicted to be so abundant, why haven’t we seen any? And what happens when we actually meet life, and what might that life look like? Music: WhiteBatAudio.com

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Thanks for Listening!

Feel free to share your thoughts on our Twitter here:  @ConductScience

Use #ConductScience on twitter to:

• Suggest a guest
• Suggest a topic
• Ask a question we can answer on the show
• Or to just get in touch!

Learn about our products:

• Visit our Lab
• Find Resources for your lab including protocols, methodologies, and more.
• Are you a scientist with a new device that you would like tech transferred? Join our Creators Program

(Image courtesy: Leica Biosystems.)

Components of the Cryostat-Microtome, their Uses, and Care Instructions

1.    Waste container

This holds the water generated by the continuous frost-defrost cycles inside the cryostat. This water can be a source of sample contamination; thus, it is important to dispose of it in the waste container.

2.    Freezing chamber

As the name suggests, it is the chamber that houses the microtome and keeps it at a low temperature while sectioning.

3.    Specimen disc

This holds the tissue specimen in place while it is being sectioned with the knife.

4.    Microtome blade and blade holder

In order to observe tissue under microscopes, it needs to be sliced to an appropriate width (in order of a few microns in width). This requires precision cutting with very sharp blades. The blades are held in position by the blade holder.

5.    Glass slide door

This is basically a cover for the cryostat.

6.    Anti-roll plate

The anti-roll plate prevents curling of the tissue sections after they have been sliced by the blade.

7.    U.V source

The ultraviolet light is an efficient sterilizer of surfaces and is, thus, used to sterilize the cryostat chamber. Usually, a 35-minute U.V irradiation is sufficient for sterilization.

8.    Source of freezing temperature

Low temperatures are maintained inside a cryostat with various modes of refrigeration. Liquid nitrogen or low temperatures by condensers. Helium is also used in cryostats, especially for cooling cryogenic detectors. The liquid cryogens are held in Dewar’s, which are built to match safety standards. It is essential to carry out regular checks to ensure that there is no physical damage to the Dewar.

9.    Peltier stage

This is a thermo-electric device which gets rid of the heat from the cryostat chamber by increasing diffusion towards a cold stage.

10.   Control panel

The control panel, as the name suggests is the hardware interface for the user to operate the Cryostat. It displays the following options:

• Lamp On/Off switch: to illuminate the Cryostat.
• V On/Off switch: to put on the U.V when sterilization is needed.
• Manual defrosting button: used to control manual defrosting. The cryostat defrosts once the experiment ends.
• Control panel lock: prevents accidental modifications in the Cryostat’s protocol set.
• Timer: the cryostat can be programmed to run for a specified length of time. It also manages the defrost timing.
• Temperature control: as the name suggests, it controls the temperature.
• Controller: for movement of sample towards the knife.

(Control panel of Leica CM1950. Image courtesy: Leica biosystems)

11.    Specimen chuck and chuck holder

Chuck is the adapter on which the tissue sample is mounted. It has a crossing grid pattern which provides better gripping. These chucks are made of stainless steel, which makes them more durable. Chucks are available in different sizes depending on the size of the sample to be processed. When in use, the chucks are kept at low temperatures in the cryostat.

The Science Behind Cooling the Cryostat

The selling point of any cryostat is its ability to maintain a low temperature. An underlying theory of a cryostat is the application of the concepts of heat exchange and gas laws. At its core, a cryostat has a couple of tubes. The first tube (tube 1) has liquid nitrogen while the second tube (tube 2) contains nitrogen gas, which is maintained slightly above the atmospheric pressure. Tube 1 is placed inside tube 2. The liquid nitrogen is vented out via a small opening in a stopper known as Flange.^[3] Due to this leak, the heat from tube 2 is conducted to tube 1 and is absorbed by the liquid nitrogen. The vaporization of liquid nitrogen removes heat from the system, thus, cooling the cryostat. The rate of heat transfer depends on the material of the tubes. Usually, silver or copper tubes are best for tube 1, but Aluminum is used in most cases to cut cost. Tube 2 is generally made of stainless steel.

Figure adapted from reference [3]

How the Cryostat-Microtome Works

Note: How various parts of the Cryostat-microtome works are briefly described in the section below. 

1.   Preparing and mounting the samples in the cryostat-microtome:

The tissue samples are flash-frozen in liquid nitrogen at a temperature of -196-degree C.  This method of freezing ensures that there is no build-up of ice crystals in the sample. These frozen samples are mounted on metal blocks with an initial chemical fixation step. The fixative is usually FAE (Formalin, Acetic acid, Ethanol, and water). The tissue sample is embedded with an embedding medium (also called OCT, which stands for Optimal Cutting Temperature agent).  Essentially, this OCT compound acts as a glue, which sticks the sample to the tissue sample holder (chuck). The chuck is cooled to the temperature of the cryostat (this is also referred to as CT or Cryostat temperature). The chuck is then mounted on the chuck holder and the position of the sample is kept perpendicular to the blade. The blade is clamped in the blade holder. The low temperature of the cryostat leads to the preservation or fixation of the ultrastructure of the specimen. This mounted specimen is then cryo-sectioned at -20 to -30-degree C, the temperature of the microtome.

2.   Adjusting the sample and optimizing the parameters for sectioning:

The mounted block of tissue samples is mechanically moved forward towards the blade for sectioning. The thickness of the slices is chosen according to the desired micron for the particular application. There is a wheel mechanism in the cryostat, which mechanically controls the movement of the chuck apparatus. This wheel is kept locked when not in use and has to be unlocked and rotated for getting the right distance from the blades and eventually cutting the ultra-fine slices to get the thickness of the desired micron. The wheel is designed in such a way that with every single rotation of the wheel, it can be programmed to advance the specimen holder by a specified micron. The section thickness is adjusted to usually 5-10 microns once the knife starts peeling the tissue.^[4,5]

3.   Adjusting the anti-roll plate or brush; a critical step:

Another important feature of the cryostat microtome is an anti-roll plate. This plate is brought down and adjusted on the blade in such a way that the slices fall underneath it and over the blade cover. In case there is no anti-roll plate provided with the instrument, one can use a brush to hold the edge of the sectioned tissue sample and place it flat on the blade cover.

4.   The golden rule:

According to Abraham Lincoln, “Give me six hours to chop down a tree and I will spend the first four sharpening the axe”. This golden rule must be followed. The blade of the knife needs to be very sharp to get thin sections. It is advisable that the blade is constantly monitored for bluntness and replaced when needed. Since the blade is not fixed, it can be moved horizontally to replace the blunt portions with sharp ends. The blade also needs to be cleaned with ethanol before sectioning to remove any dirt or adherent tissue debris. The blade is very sharp and great care needs to be taken while inserting and removing it from the blade holder. Often, the end of a brush is used to nudge it out of the holder rather than attempting to do the same with fingers. The speed of slicing by the blade must be optimized by first keeping a steady speed and then cutting the tissue with a faster speed. This ensures that the tissue sectioning generates a clean cut every time.

5.   Final piece of advice: Don’t lose your sample!

Many tissue sections are cut so that at least one of these sections provides any worthwhile information when viewed under the microscope. The sample is never left in the cryostat overnight since the machine is usually programmed to defrost and this is going to damage the sample. Usually, if the sample is required to be reused, it is kept back in a low-temperature freezer.^[6,7]

Conclusion

The contribution of Cryostat to the healthcare industry is quite significant. Analysis of a small tissue section from the patient is often used to determine the next course of treatment. In the case of diseases such as cancer, the treatment of a patient cannot resume unless the results of tissue biopsy are back from the immunocytochemistry lab. Thus, it becomes important that diagnosis is concluded early using techniques that involve the Cryostat-microtome.

Furthermore, the cryostat is very important for tissue section preparation for Fourier Transform Infra-Red (FTIR) spectroscopy, where even a small temperature difference between the blade and the sample holder can have huge quality issues in the sectioned samples.^[8]

References

1. Parma, V. (2013) Cryostat Design. CERN Technology Department, Magnets, Superconductors and Cryostats Group
2. II, J. G. W. (2016) Principles of Cryostat Design, Springer
3. Joiner, B. B. (1968)Design and construction of a liquid nitrogen cryostat for use in a swimming pool type reactor, Missouri University of Science and Technology
4. Hyam, P. (2010) Understanding and Maintaining the Cryostat, Springer
5. Gschwendtner, A., and Mairinger, T. (1995) How thick is your section? The influence of section thickness on DNA-cytometry on histological sections. Anal Cell Pathol9, 29-37
6. Dey, P. (2018) Frozen Section: Principle and Procedure, Springer
7. Son, A. I., Sokolowski, K., and Zhou, R. (2013) Cryosectioning. Methods Mol Biol 1018, 301-311
8. Liyanage, S., Dassanayake, R. S., Bouyanfif, A., Rajakaruna, E., Ramalingam, L., Moustaid-Moussa, N., and Abidi, N. (2017) Optimization and validation of cryostat temperature conditions for trans-reflectance mode FTIR microspectroscopic imaging of biological tissues. MethodsX4, 118-127

Hello! Who are you and what are you working on?

I am a product designer and ergonomist. I am currently pursuing a Ph.D. in Health Technologies at the Universidad Politécnica de Valencia. I have a grant from the Ministry of Sciences of Colombia and the Universidad del Norte teacher training program.

I started this Ph.D. program in 2019 and aim to complete it by the end of 2022.  My research focuses on neuropathic hand injury rehabilitation, therapeutic adherence and assistive product development. We have recently conducted a systematic review on the efficacy of electrophysical modalities for the treatment of radial, ulnar and median peripheral neuropathies.

What’s your backstory and how did you come up with the idea?

My interest in upper limb rehabilitation began in 2007 after I was involved in an automobile accident. I had multiple fractures and paralysis due to radial nerve injury. This experience allowed me to analyze orthopedic products’ characteristics from the designer’s and user’s point of view. A decisive event was designing an immobilizer to consolidate my ribs and clavicle fractures because none of the prescribed ones worked adequately.

My goal is to improve people’s quality of life with motor dysfunction through the design of assistance and rehabilitation products.

Please describe the process of learning, iterating, and creating the project

I believe it is essential never to lose curiosity and the desire to learn. Identify what knowledge and skills could nurture your life project. In my case, I have opted for postgraduate studies that did not have thematic lines associated with product design, but that has allowed me to have a comprehensive vision as if it were a macrosystem.

It is crucial to keep in mind that learning, creating, or researching are cumulative processes and their advances arise from trial and error.

Please describe the process of launching the project

To disseminate our results, we used academic or professional networks such as LinkedIn or ResearchGate and other social networks such as Twitter.

How is everything going nowadays, and what are your plans for the future?

My mid-term plan is to validate the design I am currently developing, finish my thesis and graduate from my Ph.D. In the long term, I want to continue working in biomechanics and hand rehabilitation, including pathologies such as stroke or Parkinson’s disease.

Through your science, have you learned anything particularly helpful or advantageous?

Each experience brings with it lessons learned. Facing a Ph.D. that is not entirely linked to my training has allowed me to grow academically and personally. For example, I have known that having reservations and fears about our expertise in the face of a new challenge prevent us from taking the first step. It is convenient to take the plunge and look for the necessary mechanisms to solve the problems as they arise (they are always lighter than the imaginary ones :D).

We’d like to know more about your lifestyle. Please let us know what is your morning routine.

The first thing I do is walk my dog, then after breakfast, I take a shower and start my day. I usually list the tasks I expect to do during the day and cross off the ones I complete. I set ambient sounds according to the task. For example, if I am going to read papers, I listen to binaural sounds; if I am in the design phase, I listen to music that inspires me.

And how does a typical day look for you?

Due to the pandemic, I have been working from home since March 2020. I am not much of a night owl, so I prefer to make the most of the day. I tend to start with the activities that demand the most attention and leave for the end those that are simple or not urgent.

At the end of my day, I go out to train (3 times a week), play with my dog, watch a movie, read a book, and do a short meditation session before going to sleep.

What does your workstation look like?

My workstation at home is very simple. I have a desktop computer, a notebook where I keep all my notes, and an acrylic whiteboard. When I’m designing, I have sketch pads and materials for mock-ups.

What platform/tools do you use for your professional life?

I use tools like Excel, Word, PowerPoint, R Studio, and Illustrator, platforms such as Zoom or Google meet, and professional networks such as LinkedIn and ResearchGate.

What secondary software and apps do you use daily?

Gmail, Google Calendar, Binaural Beats, and Meditopia app.

How do you stay up to date on News and resources?

I keep up to date with the news through newscasts and online media, although I have tried to filter a bit for mental health in the wake of the health crisis.  In terms of academia, I use Google Scholar alerts and receive newsletters from journals that publish interest information.

What have been the most influential podcasts, or other resources?

I’m not very into podcasts. The book that got me interested in my field was “Fundamentals of Biomechanics“. In the challenge of publishing, it helped me to have the Cochrane and Grade manuals.

What tools do you use in your personal life? Cook? Self-Care? Hobbies?

I do functional exercises and sparring three times a week. On weekends, I take long walks with my dog and spend time with my boyfriend, family, and friends.

I enjoy cooking, hiking, singing, and when I feel inspired, writing poetry.

Advice for other scientists who want to get started or are just starting out?

I think it is essential to find a topic that genuinely interests you, have clear goals but not lose sight of the day-to-day. Learn from mentors, be receptive to their suggestions and rejoice with each new learning. Be flexible and open-minded to all possibilities and objective enough to doubt them all. Keep abreast of developments in your field of study. In moments of mental exhaustion, do not continue working on the same activity but alternate it with another task that requires a different skill. Try to manage stress by whatever method works best for you (meditation and coaching have helped me).

Thank you very much for your time, Ena. Where can we go to learn more?

Social media:
https://www.linkedin.com/in/enabula/
https://twitter.com/enabula87

Research:
https://www.researchgate.net/profile/Ena-Bula
https://orcid.org/0000-0001-9158-5830
https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0248484