Meet the people behind Volume EM community (part 2)

In this second post, we continue talking with some of the scientists involved in the Volume EM initiative. If you missed the first part of these series of interview, you can find it here.

Raffa Carzaniga and Errin Johnson, Training Working Group

What are the aims of your WG?

The main aim of the vEM Training WG is to provide resources and training on both the technical skills and managerial skills relating to all aspects of vEM.

Which types of people are in your WG? 

Our WG has over 20 members from across the UK, Europe and the US. We have a wonderful mix of electron microscopists from EM Facilities, industry and research. As such, there are a range of different perspectives on vEM training within the group, which has led to some very interesting discussions and lots of great ideas for facilitating vEM training for both novice and experienced users.

How did you come to be involved with the vEM Community Initiative?

EJ – I attended the first Town Hall meeting in 2019 and thought it was a brilliant initiative that I wanted to contribute to. Helping people understand not just how to use electron microscopes, but how they work, and how different electron microscopy techniques can be applied to specific research questions, is something I really enjoy.  When the opportunity came up to co-chair the Training WG with Raffa, I jumped at it!

RC – From the early conceptual stages and discussions I felt that I wanted to contribute and to be part of this exciting initiative. Training and knowledge transfer is one of my passions – it’s great fun to co-chair the Training WG with Errin and enjoy collaborating with a lot of imaging colleagues. I also think that Volume EM has a lot of untapped potential and raising awareness through training is a great way to expand the community.

What are you doing to accomplish your WG’s aims?

We decide to pursue 4 main areas to achieve our aim. Firstly, to establish a database for people to deposit training resources, such as courses, conferences and instructional videos. Secondly to create an vEM introductory pack to help people get started in the field. The pack will include recommended tools and consumables, as well as key papers covering each area of vEM. We’re also exploring setting up regular technical seminars and generating new training related content for the vEM website, such as tips/tricks and short technique highlights. Finally, we are in the process of creating a series of primer videos on vEM for novice users, as well as recorded seminars for advanced users, which will cover: the history of biological EM, sample preparation for vEM, FIB-SEM, array tomography, serial block face SEM, 3D correlative microscopy and data analysis. We’ve made good progress in these areas this year, much of which will come to fruition next year!

Challenges/Opportunities?

Everyone in the WG is really keen to contribute to our aims, but we’re all volunteering our time for this initiative on top of heavy workloads. So the main challenges are logistical: finding times for meetings that work across multiple time zones, and that suit the schedule of as many people in the group as possible, as well as making the time to action our initiatives. For the videos in particular funding will be required to help make these possible, so that is something we will need to focus on in 2022.

Which vEM discovery has struck you the most?

EJ – I’d like to answer it more from a technical point of view about applications of vEM that have really inspired me to learn more about the techniques and implement them in my own facility. In this case, there are two pieces of work that showcase the power of vEM for me. The first is by Eija Jokitalo’s group at the University of Helsinki, where they were an early adopter of serial block face imaging and published an excellent paper (Puhka et al. 2012, Molecular Biology of the Cell) that showed the variation in 3D ER ultrastructure across different cells lines. The second is by Yannick Schwab’s group at EMBL in Heidelberg, where they correlated two-photon intravital microscopy, x-ray micro-computed tomography and FIB-SEM to target single mouse brain tumour cells in vivo and image their invasion through brain capillaries at ultrastructural resolution in 3D. This paper is a stunning example of both CLEM and vEM (Karreman et al. 2016, Journal of Cell Science)!

Tell us something about yourself that not many people know? Or What’s your favourite ice-cream flavour?

EJ – I’m training to be a fitness instructor. Don’t worry, I won’t give up my day job!

RC – Black sesame with white miso caramel

Michelle Darrow and Carles Bosch, Sample prep Working Group

What are the aims of your WG?

The remit of the Sample Prep working group is the curation of sample preparation resources for the vEM community, including a protocol library that encompasses preparation steps through to imaging, with a focus on discussion and dissemination of knowledge around why each step is done, health and safety information, and annotations to indicate options and when they might be useful.

Which types of people are in your WG?

We have around 30 members in our working group, spread primarily across Europe, the US and Australia and ranging from academia to industry and from post-doctoral research associates to heads of facilities.

How did you come to be involved with the vEM Community Initiative?

MD: My path to the vEM community is a bit serpentine – my current research focuses on the development of an instrument for single particle cryo electron microscopy (cryoEM) sample preparation and software tools for 3D visualisation, annotation, and segmentation. However, in my past research I dabbled in many of the techniques used in the vEM community and I see many overlapping benefits across imaging communities, so I decided to get involved!

CB: I have been using different kinds of volume electron microscopy since my PhD to study neuronal circuits, with a growing interest on correlating different imaging modalities together in a single experiment (be it light microscopy, X-rays, or others). Correlating imaging modalities brings added value to the experiment – but also forces you to create sample preparation protocols that are compatible with the different modalities you want to employ. The vEM community seemed like a great initiative where I could bring a little and learn a lot from, so the decision was clear the moment I heard about it – sure I’m in.

What are you doing to accomplish your WG’s aims?

We’ve gone in two different directions to start with. The first thing we’ve done is to collaborate with EMBL-EBI and the vEM Data working group to develop and test a Sample Prep Protocol widget. This widget allows capturing and storing the specimen preparation details (including metadata) that are associated with a given dataset – which will be stored in EMPIAR. This tool will allow individuals and groups to track, publish and share their protocols more easily. It will also make accessing and re-using that pooled knowledge in the future a simpler task.

The second thing we’ve been focusing on is a meta-analysis of various established sample preparation protocols. We started with seven common protocols for serial-block face scanning electron microscopy and have compared them to identify steps with high and low agreement. We’ve been able to combine these protocols to create a single merged protocol that highlights “mutation hotspots” where variations in the protocol are most common. We will soon expand this analysis to include other protocol families, next ones being focused-ion beam SEM and array tomography.

In addition to the current goals discussed above, we’ve made plans to start a new program to create a sample library. This will provide the community with standard samples, prepared and imaged in standard, well-documented ways, and an associated image library, all available on request. We envision this community resource will enable training new users, troubleshooting sample prep and imaging issues, prototyping of new functionalities, testing new microscopes and the acquisition of benchmark data for software development, among many other things!

Challenges/Opportunities?

We’re excited about the progress we’ve already made, about the future plans, and about the extraordinary group of individuals from all over the world we come to share ideas with through this initiative. However, as a group of volunteers, we sometimes struggle to find the time and resources to get things done.

Which vEM discovery has struck you the most?

CB: I can mention two papers that, using volume EM, were influential for the neuroscience field in two topics I have worked on. The first one is Harris et al. 1992. Using serial section TEM they reconstructed in 3D dendritic spines of a specific cell type and zone across two developmental ages. Spines are the receiving structure of the vast majority of excitatory synapses in the brain, and their structure is highly related to their function. Some of their dimensions escape the resolution of conventional confocal microscopes, making EM a good choice to resolve them fully. This study ensembled a collection of morphological parameters to describe the structure of spines, and paved the way for many future studies exploring those tiny structures. The second one is Briggman, Helmstaedter and Denk 2011.This work was the first time a neuronal circuit was studied correlating physiological function and synaptic-level dense ultrastructure. The result was impressive – they described how two cell types in the mouse retina (amacrine and ganglion cells) established connectivity patterns according to their physiology, and how those connectivity patterns helped refine hypotheses explaining the circuit’s computational logic. But more importantly, it proved the point that circuit-level systems neuroscience could use volume EM, and that in vivo readouts could be combined with ultrastructural insight to decipher how neuronal circuits operate in the brain. Certainly an inspiring challenge.

What’s your favourite ice-cream flavour?

MD: My favorite dessert flavor is the chocolate-peanut butter combination! The ratio is important and I’m happy to discuss the obvious flaws in the ratio associated with the mini-cup design 🙂

CB: That is a difficult choice. My vote would probably go for “Marc de Cava Sorbet” – an amazing ice-cream cocktail made with lemon sorbet, distilled cava… but well, from here on we should probably compare recipes to give an accurate description. Maybe the widget will help! 

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