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Artificial intelligence applied to intravital video microscopy, 3rd edition
13 - 15 November 2023

This workshop will cover theoretical and practical aspects regarding the application of A.I. -based methods for the analysis of cell motility from microscopy data, with a focus on immune cells observed in vivo. Moreover, we will discuss the importance of Open Source and Open Data resources in image-driven immunological research.
The methods presented in the workshop can be applied also to other cell types and imaging modalities. A basic understanding of microscopy is recommended.
How: Best would be to attend in presence, with practical cases to be solved in small groups. We will provide data and tools. You have to bring your own laptop. You can also bring your own data to be analyzed with the help of the peers.
For those who cannot come to Lugano in person we will leave open the possibility to connect via Google meet, attending lectures and watching the demonstrations. However, interactions may be restricted.

Link to attend online https://meet.google.com/qiq-yuug-vif 

Where: USI Campus EST, Via La Santa 1, 6962 Lugano – Switzerland

Organizers:
Prof. Santiago Fernandez Gonzalez
Prof. Rolf Krause
Dr. Diego Ulisse Pizzagalli

Confirmed speakers:
Pau Carrillo Barbera (University of Edinburgh)
Raffaella Fiamma Cabini (University of Pavia & USI Lugano)
Diego Morone (IRB Bellinzona)
Santiago Fernandez Gonzalez (IRB Bellinzona)
Mariaclaudia Nicolai (USI Lugano)
Elisa Palladino (IRB Bellinzona)
Diego Ulisse Pizzagalli (USI Lugano)
Alain Pulfer (IRB Bellinzona)
Inge Wortel (Radboud University)

Contact: For questions do not hesitate to contact us at This email address is being protected from spambots. You need JavaScript enabled to view it.

DAY 1: November 13th. Introduction, intravital imaging and Open Data

Learning objectives. Understanding the applications and advantages of intravital microscopy / the importance of Open Data research / Practice with immunemap platform and other tools

09:00 - 10:00 Introduction - DUP
10:00 - 11:15 Principles of fluorescence and intravital microscopy – DM
11:15 - 11:30 Coffee break
11:30 - 12:30 Studying the immune system in vivo, surgical models and applications - SFG
12:30 - 14:00 Lunch Break
14:00 - 15:00 Open Data research, FAIR principles and Immunemap  DUP + EP
15:00 - 17:00 Practical session 1 in small groups: Reuse of datasets and Open source tools

DAY 2: November 14th: Analysis with tracking. 

Learning objectives. Understanding what can be extracted from IVM data using the classical image analysis pipeline (tracking and track measures), challenges and new perspectives.

09:00 - 09:45 Basic measures of cell motility and cell-to-cell interaction. DUP+MN
10:15 - 10:30 Advanced metrics of cell motility + introduction to CelltrackR. IW
10:30 - 10:45 Coffee break
10:45 - 12:30 Practical session 2: Exercises to quantify motility
12:30 - 14:00 Lunch Break
14:00 - 15:00 Tracking with FIJI/Trackmate – PCB
15:00 - 16:00 Practical session 3: Practice with FIJI and Trackmate
16.00 - 17.00 Deep learning based cellular action recognition (mock dissertation - AP)

DAY 3: November 15th: Analysis without tracking.

Learning objectives. Understanding how image processing and computer vision techniques can be applied to analyze IVM data without necessarily using of cell tracking

09:00 - 10:15 Image processing and supervised machine learning techniques to
                      - facilitate cell detection and tracking DUP+RFC
                      - quantify motility when it is impossible to track individual cells DUP
10:15 - 10:30 Coffee break
10:30 - 12:00 Unsupervised machine learning technique for
- cell segmentation RFC
- cell swarming analysis DUP
- identification of main motility patterns DUP
12:00 - 13:00 Lunch Break
13:00 - 14:00 Discussion: recent trends in computer vision methods
14:00 - 16:00 Practical session 3: Analyzing cell migration without tracking
16:10 - 17:00 Conclusion

PAST EDITIONS

Artificial intelligence applied to intravital microscopy of the immune system, 2nd edition 16 - 18 May 2022

This workshop will cover theoretical and practical aspects regarding the application of A.I. -based methods for the analysis of intravital imaging data.
We will focus on the migration and interaction of immune cells observed in vivo, explaining how to get accurate tracking analysis, or how to extract insights from videos without using cell tracking.
Moreover, we will discuss the importance of Open Source and Open Data resources in image-driven immunological research.
The topics and tools presented in this workshop can be applied also for other imaging modalities and cell types but a basic knowledge in microscopy is recommended.

How: Blend modality. Best would be to attend in presence, with practical cases to be solved in small groups. We will provide data and tools and can offer a few workstations to be used but is encouraged to bring your own laptop. You can also bring your own data to be analyzed. For those who cannot come to Bellinzona we will leave open the possibility to connect via Zoom and watch the demonstrations.

When: May 16th – 18th, optional excursion on May 19th

Where: Institute for Research in Biomedicine, Via Francesco Chiesa 5, CH-6500 Bellinzona (Switzerland) / or online on Zoom (we will do the possible to let you following, but consider that technical issues may arise for the practical sessions)
Registration: up to May 8th 2022 23:59 CEST at https://forms.gle/x3XqkdqG7PxMBVH56 15 places available, served on a first come first served basis. You will receive a confirmation via email.

Cost: Free of charge

Contact: For questions do not hesitate to contact us at This email address is being protected from spambots. You need JavaScript enabled to view it. with the subject of the email ai-ivm2022

Program is in CEST timezone. Program can be subjected to changes, here you will always find the most updated schedule.

DAY 1: May 16th. Introduction to intravital microscopy (IVM).

Learning objectives. Understanding the applications and advantages of intravital microscopy / Understanding the importance of Open Data research / Practice with the immunemap platform

08:45-09:15: Walk-in and welcome of participants

9:15-10:15: Introduction of the course and Ice-breaker activity (Pizzagalli DU)

10:15-11:15: Technical Introduction to IVM (Morone D)

11:15-11:30 Coffee break

11:30-12:30: Surgical models and biological applications of IVM (Virgilio T)

12:30-14:00: Lunch Break

14.00-14:25: Summary of the concepts presented during the morning, Q&A session (Pizzagalli DU)

14:30-15:15: Introduction to cell tracking and open data research (Pizzagalli DU)

15:15-15:45: Coffee break

15:45-16:00: Presentation of immunemap (Palladino E / Ceni K)

16:00-17:00: Practical session 1: Use of immunemap and open data resources (Palladino E / Ceni K / Thelen B)

 

DAY 2: May 17th: Analysis with tracking. 

Learning objectives. Understanding which type of information can be extracted from IVM data using the classical image analysis pipeline, challenges and new perspectives

9:00-10:30: Classical image analysis pipeline, artifacts, quality requirements (Pulfer A)

10:30-10:45 Coffee break

10:45-12:00: The relevance of cell tracking in life sciences, motility parameters (Pizzagalli DU)

12:30-14:00: Lunch Break

14.00-14:25: Summary of the concepts presented during the morning, Q&A session (Pizzagalli DU)

14:25-15:00: Practical session 2a, Facilitating automatic tracking using computer-assisted colocalization (Pizzagalli DU)

15:00-16:00: Practical session 2b, Analysis of motility measures and cell-to-cell interaction (Pizzagalli DU)

16:00-16:15 Coffee break

16:15-17:30: Practical session 2c, Manual tracking and crowd-based tracking using immunemap. Manual tracking in FIJI/Trackmate and Imaris on request. (Ceni K, Carrillo-Barbera P, Palladino E).

 

DAY 3: May 18th: Analysis without tracking.

Learning objectives. Understanding how image processing and computer vision techniques can be applied to analyze IVM data without the usage of cell tracking

9:00-10:30: From pixels to cell actions (Pizzagalli DU)

10:30-10:45 Coffee break

9:00-10:30: Recent trends in computer vision methods (Pulfer A)

12:30-14:00: Lunch Break

14.00-14:25: Summary of the concepts presented during the morning, Q&A session (Pizzagalli DU)

14:25-15:45: Practical session 3: Advanced analysis of cell migration (Motility heatmaps, Optical flow, Tracklet-based action recognition, Colocalization, Cell death detection, Swarm detection) (Pizzagalli DU, Pulfer A)

15:45-16:00: Coffee break

16:15-16:45: Practical session 4: Unsupervised machine learning to identify migration modalities (Pizzagalli DU)

16:45-17:00: Conclusion

 

DAY 4: May 19th: Social activity

09:00–12:00: Visit to the castles of Bellinzona

Organizers:
Dr. Santiago Fernandez Gonzalez - Institute for Research in Biomedicine (CH)

Dr. Diego Ulisse Pizzagalli - Institute for Research in Biomedicine and Euler Institute, USI (CH)

Tutors:
Dr. Pau Carrillo-Barbera – The University of Edinburgh (UK)

Kevin Ceni – Institute for Research in Biomedicine (CH)
Diego Morone – Institute for Research in Biomedicine (CH)
Elisa Palladino – Institute for Research in Biomedicine (CH)
Alain Pulfer – Institute for Research in Biomedicine and ETH Zurich (CH)
Benedikt Thelen – Euler institute (CH)

Dr. Tommaso Virgilio - Institute for Research in Biomedicine (CH)

 

 

---- MATERIAL FROM PREVIOUS EDITION (2021) ----

Description

This summer school covered theoretical and practical aspects regarding the analysis of immune cell migration and interaction, using intravital microscopy and new A.I.-based methods. We had 18 participants (4 from USI, 14 external), from 4 european countries including BSc, MSc, PhD students, postdocs and imaging specialists.

Organizers. Dr. Santiago Fernandez Gonzalez, Dr. Diego Ulisse Pizzagalli, dr. Pau Carrillo Barbera
Tutors. Diego Morone, Alain Pulfer, Kevin Ceni, Benedikt Thelen
Supported by. IRB PhD program    Cell Migration PhD program    USI - Biomedical PhD program    IMMUNEMAP consortia    Euler Institute

Program

DAY 1: June 14th. Introduction to intravital microscopy (IVM).

Learning objectives. Understanding the applications and advantages of intravital microscopy / Understanding the importance of Open Data research / Practice with the immunemap platform

Introduction and networking activity (Pizzagalli)

1

9:00 - 10:00

Physics of 2-photon intravital microscopy (Morone)

1

10:30 - 11:30

Intravital imaging of the immune system (Gonzalez)

1

11:45 - 12:45

Image formation / histograms / Properties of digital images (Barbera / Morone)

1

14:00 - 15:00

The role of Open Data and Open Source in biomedical research , FAIR principles and IMMUNEMAP project (Pizzagalli / Barbera)

1

15:10 - 15:45

FIJI introduction (Morone)

1

15:45 - 17:00

DAY 2: June 15th: Analysis with tracking. 

Learning objectives. Understanding which type of information can be extracted from IVM data using the classical image analysis pipeline, challenges and new perspectives

Exercise 1

  • Which histogram is saturated
  • Thresholding
  • Use pixel inspector

2

9:00 - 9:30

Image analysis pipeline, quantification of cell motility, dynamism, and interaction (Pizzagalli)

2

9:30-10:30

The possibilities of Machine Learning in IVM (Pizzagalli)

2

10:45-11:30

Excercise 2 (Morone / Pulfer / Barbera Pizzagalli)

  • cell detection (classical / Otsu etc)
  • Thresholding with TWS
  • stardist
  • trackmate
  • automatic tracking with trackmate
  • computation of measures: collective (i.e. preferential direction) and individual cell behavior (i.e. change of speed)

2

14:00 - 17:00

DAY 3: June 16th: Analysis without tracking.

Learning objectives. Understanding how image processing techniques working at pixel-level can be applied to analyze IVM data without the usage of cell tracking

Exercise: track-based measures on neutrophil chemotaxis

Exercise: migration differences between WT and KO cells in spleen

 

9:00 – 10:30

Discussion Exercise 1 (Pizzagalli)

Contact analysis in Exercise 1 (Pizzagalli)

Advanced analysis without tracking (of/recruitment) (Pizzagalli)

3

10:45 - 11:00

Image processing techniques (Pulfer)

3

11:15 - 12:15

Exercise 3

  • Quantify the recruitment in vessels
  • Analyze whole LN images with OF
  • Apply the bayes coloc plugin by Alain (cell-cell interaction)
  • Imaris workshop (tracking, coloc, heatmaps, and cell-cell interaction)

3

14:00 - 17:00

DAY 4: June 17th:  Beyond tracking.

Learning objectives. Understanding how computer vision methods for action recognition can be applied to analyze IVM data without the usage of cell tracking

Inside AI: neural networks and clustering (Pizzagalli)

4

9:00 – 10:00

Trends in computer vision: classic, deep learning, generative (Pulfer)

4

10:05 - 11:00

Action recognition applied to immune cells (Pizzagalli)

  • Review of motility patterns displayed by immune cells in vivo and their biological meaning
  • Application of action recognition to quantify neutrophil dynamics and migration morpho-phenotypes

4

11:15 - 12:15

Exercise 4 (Pulfer / Pizzagalli)

  • Usage of the apoptosis detection program (incl. challenge)
  • Discussion on the limits and potential usage of Deep Learning in IVM
  • Neutrophil swarm detection

4

14:00 - 16:00

Discussion on good practices for IVM analysis (Barbera + Pizzagalli)

4

16:15 - 17:00

DAY 5: June 18th: Workshop and conclusion

Workshop: analyze data by participants (Pizzagalli):

- Quantification of leukocyte migration within and outside blood vessels via track-based measures and pixel classification.

- Analysis of DCs – Macrophage interaction via supervised spectral unmixing, contact analysis, and bayesian colocalization.
- Quantification of neutrophil swarm dynamics via unsupervised machine learning.

5

9:00 - 12:00

Exam

5

14:00 - 15:00

Concluding remarks

5

15:30 - 16:30

 Project directors

  • Dr. Santiago F. Gonzalez is a group leader at the Institute for Research in Biomedicine (IRB, Bellinzona, Switzerland). Dr. Gonzalez has extensive experience in the field of imaging. During his postdoctoral research at Harvard University, he received complete training for the use of state-of-the-art imaging techniques, including MP-IVM. After establishing his own group in Switzerland, he has completed, with support from the Swiss National Science Foundation, the installation of a state-of-the-art MP-IVM facility at the IRB.
  • Prof. Dr. Rolf Krause is the director of the Euler institute and a professor with a chair in Advanced Scientific Computing at USI, Lugano - Switzerland.
    His research focuses on numerical simulation and mathematical modelling in scientific computing and computational sciences, leading the research group "High Performance Methods for Numerical Simulation in Science, Medicine and Engineering". He is also co-director of the recently installed Center for Computational Medicine in Cardiology - CCMC.
  • Dr. Diego Ulisse Pizzagalli is an independent researcher in Computational Medicine at the Università della Svizzera italiana (USI, Lugano, Switzerland) and visiting scientist at IRB. In his doctoral studies, supported by systemsx.ch, Diego focused on network theory to study the migration and interaction of immune cells in vivo, contributing to bridging the gap between computer vision and immunology. Moreover, he recently translated these techniques from cells to patients and is currently co-PI in a project aiming at monitoring longitudinally a cohort of patients with chronic liver disease. Diego serves as a teaching assistant and lecturer at USI in subjects related to machine learning in medicine, computing theory, and signal processing.

Project partners

Immunemap is a collaborative project supported by a network of international laboratories willing at making 2-photon imaging data of the immune system FAIR (Findable, Accessible, Interoperable, Reusable).

 

Britta Engelhardt (Theodor Kocher Institute, University of Bern, Bern, Switzerland). The group of Prof. Engelhardt developed an extensive in vivo imaging biobank that includes 2P-IVM imaging of T cell interactions in the cervical spinal cord microvessels of mice suffering from experimental autoimmune encephalomyelitis. Prof. Engelhard has a solid career as a relevant world expert on imaging autoimmune diseases. Mykhailo Vladymyrov (University of Bern), Neda Haghayegh Jahromi (System Biology Division, Philip Morris International, Neuchâtel, Switzerland).

Hans Uwe Simon (University of Bern, Bern, Switzerland). MD Uwe’s laboratory explores apoptosis and autophagy in inflammatory diseases and cancer, focusing on atopic dermatitis, hypereosinophilic syndromes, eosinophilic esophagitis, bullous pemphigoid, and malignant melanoma. They aim to identify novel drug targets for therapeutic interventions. Additionally, they have developed in vitro and in vivo systems to assess drug effects on the immune system. Engaged in clinical drug studies, their research bridges pathogenesis understanding with translational medicine for improved disease management. Nina Germic (Respiratory Infections Division, Roche, Rotkreuz, Switzerland).

Jens Volker Stein (University of Fribourg, Switzerland). Dr. Stein wants to clarify the molecular and cellular processes that govern adaptive immune responses mediated by cytotoxic CD8+ T cells. His research line examines lymphoid tissue using two-photon microscopy (2PIVM) to define the key regulators of T cell activation by using genetically modified CD8+ T cells. Petra Pfenninger, Jun Abe.

Cornelia Halin (ETH, Zurich, Switzerland). The group of Prof. Halin specializes in the study of intravital microscopy observations of migratory T cells and dendritic cells in and around dermal lymphatic capillaries in murine skin. Since the establishment of her lab in ETH, Prof. Halin has published her innovative research in the most important journal in the field of immunology. Morgan Hunter.

Wolfgang Kastenmuller (University of Bonn, Bonn, Germany). Prof. Kastenmuller directs a dynamic group that focuses on the characterization of the interactions between DC and CD8 and CD4 T cell subsets in the LN in response to different conditions. The Kastenmuller lab has significantly contributed to the field of immuno-imaging and has published innovative imaging articles in top journals like Immunity or Nature Immunology. Sarah Eickhoff (University Hospital Bonn, Bonn, Germany).

Andres Hidalgo (Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain). The group of Dr. Hidalgo is specialized in the study of the migration mechanisms that lead to the recruitment of inflammatory cells from the blood vessels following different insults. He has published his recent papers on neutrophil dynamics in journals like Cell or Science. Miguel Palomino-Segura, Georgiana Crainiciuc, José M. Androver.

Matteo Iannacone (San Raffaele Research Institute, Milan, Italy). MD PhD Iannacone directs a group specialized in studying the activation of adaptive immune cells and immunopathology during viral infection and cancer. He has developed specific models to study the dynamics of T cells in the liver in response to viral infection. Iannaccone’s group has recently published novel research that regards liver imaging in the journal Cell. Xenia Fitch, Federica Moalli, Alexandre Bénéchet (CHUV, Lausanne University Hospital)

Philippe Bousso (Pasteur Institute, Paris, France). Dr. Bousso leads the group Dynamics of Immune Response at the Pasteur Institute. By further developing functional in vivo imaging, they aim at identifying critical aspects of T cell and NK cell activation and function during tumor growth, during infection by an intracellular parasite, and transplantation. Hélène Moreau.

Ana-Maria Lennon-Duménil (Curie Institute, Paris, France). Dr. Ana-Maria Lennon-Duménil's scientific career has been built on the use of multidisciplinary approaches to understand, at different scales, the functioning of cells in the immune system. In 2008, she set up a consortium including cell biologist M. Piel (I. Curie) and theoretical physicist R. Voituriez (UPMC, Paris). Their goal was to apply quantitative imaging and microfluidics to study molecular mechanisms and physical principles governing the ability of cells of the immune system to move. Their work has opened up a still unexplored line of research into the mechanisms that allow dendritic cells, to coordinate their functions with their migration in time and space. They have given rise to more than 25 publications, including some in very prestigious journals such as Science, Cell, Nature Cell Biology, Nature Physics, and Developmental Cell. Dorian Obino.

Milka Sarris (University of Cambridge, Cambridge, UK). Dr. Sarris' laboratory investigates how immune cell movement is orchestrated during inflammation using zebrafish larvae as a model. Leveraging advanced microscopy and genetic manipulation, they decipher the molecular and physical cues guiding leukocyte recruitment to damaged tissues. Their goal is to obtain a better understanding of how leukocytes interpret complex environmental cues to generate effective immune responses. Antonios Georgantzoglou (University of Copenhagen, Copenhagen, Denmark).

Johannes Textor (Radboud University, Nijmegen, Netherlands). Dr. Johannes Textor leads the Computational Immunology group at Radboud University, employing simulation models, machine learning, and causal inference to study the adaptive immune system. Their research aims to understand immune responses to pathogens and tumors, informing immunological treatments. They develop Artificial Immune Systems to compare immune and nervous system information processing, investigating their distinct architectures and complementary roles. Collaborating with colleagues at Radboudumc's Tumor Immunology department, they provide computational support, advancing immunological research through diverse methodologies. Inge M. N. Wortel.

 

Thomas Murooka (University of Manitoba, Manitoba, Canada). The laboratory of Dr. Murooka specialized in the study of T cells and DC as well as their migration during HIV infection in lymphoid tissues. Additionally, they studied the effector and regulatory T cell behaviors during Leishmania major infection in mouse skin. During his career, Dr. Murooka has published high-impact factor papers in journals like Nature.

Michael Carroll (Harvard Medical School, Boston, USA). The group of Prof. Carroll has made important contributions to the field of antigen trafficking and B cell biology. He has extensive experience characterizing the behavior of DC and B cells in different models including infectious diseases and autoimmune diseases. The group publishes their research in a regular base in the highest impact factor journals in the field including Nature, Science or Cell.

Francesco Marangoni (University of California - Irvine, USA) . Dr. Marangoni studies immune-evasion mechanisms, especially the ones favouring the development of tumours. His primary technological approach combines multiphoton intravital microscopy with functional reporters of biological processes (functional intravital microscopy or F-IVM). His long-term goal is to devise new forms of immunotherapy through the manipulation of immune-evasion mechanisms. His Functional Immuno-Imaging Lab investigates whether checkpoint blockade immunotherapy enhances the function of tumour-associated Treg, which in turn may limit therapeutic efficacy. Moreover, their studies hinge on new mouse models allowing to map the function of TAM (Tumor-associated macrophages) to identify the factors leading to TAM reprogramming towards pro-inflammatory functions in vivo.

Michael D. Cahalan (University of California - Irvine, USA). Dr. Chalan's research incorporates single-cell approaches to investigate the immune response, using the patch-clamp method to characterize ion channels and a variety of imaging techniques to monitor motility, cellular interactions, Ca2+ signalling, and gene expression.  His pioneering work identified the pivotal role of ion channels in the immune response. He elucidated physiological functions and the molecular basis for calcium signalling that activates T lymphocytes. By imaging lymphoid organs, his work revealed an elegant cellular choreography that underlies immune responses in vivo. Shivashankar Othy.

Thorsten Mempel (Massachusetts General Hospital, USA). Dr. Mempel's Laboratory is seeking to understand how the function of T cells is regulated through their interaction with other cells, structural tissue components, and soluble mediators that they encounter in tissues. Their main approach to this end is direct dynamic in vivo visualization of immune processes at cellular and subcellular resolution in living mice, using multiphoton intravital microscopy.

Mauro Di Pilato (MD Anderson Cancer Center, Houston, Texas, USA). The laboratory of Dr. Di Pilato investigates and characterizes which immune signals are involved in the intratumoral infiltration of unstable Treg. Specifically, they want to focus on pathways that are involved in Treg intratumoral destabilization, proliferation and survival and define new strategies of Treg accumulation. Their goal is to develop new approaches that will increase the effectiveness of cancer immunotherapy and the number of cancer patients who will respond to immune checkpoint blockade inhibitors, and finally to prevent the progression of primary melanomas.

Chris Xu (Cornell University, Ithaca, NY, USA). Dr. Xu is currently a Professor of Applied and Engineering Physics, at Cornell University, the Mong Family Foundation Director of Cornell Neurotech– Engineering, and the Director of the Cornell NeuroNex Hub, an NSF-funded center for developing neurotechnology. Before Cornell, he was a member of the technical staff at Bell Laboratories, where he developed new long-haul fiber optic communication systems. His current research areas are biomedical imaging and fiber optics. He is a fellow of the Optical Society of America and a fellow of the National Academy of Inventors. Kibaek Choe.

 Michael Hickey (Monash University, Melbourne, Australia). Professor Michael Hickey is the Director of the Monash Centre for Inflammatory Diseases at Monash Medical Centre. The main focus of his current research is white blood cell-mediated inflammatory injury of the kidney, a major cause of kidney failure. He investigates the mechanisms whereby white blood cells are attracted to the glomerulus and understands their behaviour once they get there. His laboratory examines inflammatory disease of the skin and how regulatory T cells dampen down skin inflammation. The ultimate aim of this work is to develop a detailed molecular understanding of the mechanisms whereby leukocytes cause or control injury in inflamed tissues. Ursula Norman.

 

 

 Data curation and communication responsible

  • Pau Carrillo Barberà is a Bioimage Analyst at IRB. Pau is responsible for handling the curation and integration of IVM data from an international network of laboratories contributing to the IMMUNEMAP database and the establishment of a data annotation workflow. In his doctoral studies at BIOTECMED (University of Valencia) he worked on the deployment of microscopy-based screening protocols and bioimage analysis workflows for the study of neural stem cells.
  • Elisa Palladino is an Imaging Specialist at IRB. Elisa is responsible for gathering data and populating the IMMUNEMAP platform. She also takes care to establish collaborations with new partners, around the world. 

Imaging specialists and developers

  • Benedikt Thelen is a Biomedical Engineer at USI. Benedikt is responsible for the development of the IT infrastructure and software backend of the database. After completing his master at the university of Bern, he has participated in multiple scientific projects as researcher and programmer.
  • Kevin Ceni is a Biomedical Engineer at IRB. Kevin is responsible for the development of the client-side software of the project (frontend) and the implementation of web-based tools for the analysis of IVM data. In his master thesis at the University of Bern he worked on the development of a convolutional neural network able to classify and locate different diseases affecting the retina.
  • Alain Pulfer is a PhD student in Bioimage Analysis at IRB. Alain works on the design of diverse bioimage data analysis tools for IVM data, e.g., for quality assessment or action recognition.
  • Joy Bordini is a Bioinformatician and Bioimage Analyst at IRB. Joy works on the design of diverse bioimage data analysis tools for IVM data, e.g., segmentation or colocalization methods.
  • Enrico Moscatello is a Engineering of Computing Systems student at POLIMI. Enrico is responsible for the development of the software and debugging.

 

Project partners

Immunemap is a collaborative project supported by a network of international laboratories willing at making 2-photon imaging data of the immune system FAIR (Findable, Accessible, Interoperable, Reusable).

 

  • Britta Engelhardt (Theodor Kocher Institute, University of Bern, Bern, Switzerland). The group of Prof. Engelhardt developed an extensive in vivo imaging biobank that includes 2P-IVM imaging of T cell interactions in the cervical spinal cord microvessels of mice suffering from experimental autoimmune encephalomyelitis. Prof. Engelhard has a solid career as a relevant world expert on imaging autoimmune diseases.
  • Cornelia Halin (ETH, Zurich, Switzerland). The group of Prof. Halin is specialized in the study of intravital microscopy observations of migratory T cells and dendritic cells in and around dermal lymphatic capillaries in murine skin. Since the establishment of her lab in ETH, Prof. Halin has published her innovative research in the most important journal in the field of immunology.
  • Wolfgang Kastenmuller (University of Bonn, Bonn, Germany). Prof. Kastenmuller directs a dynamic group that focuses on the characterization of the interactions between DC and CD8 and CD4 T cell subsets in the LN in response to different conditions. The Kastenmuller lab has significantly contributed to the field of immuno-imaging and has published his innovative imaging articles in top journals like Immunity or Nature Immunology
  • Andres Hidalgo (Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain). The group of Dr. Hidalgo is specialized in the study the migration mechanisms that lead the recruitment of inflammatory cells from the blood vessels following different insults. He has published his recent papers on neutrophil dynamics in journals like Cell or Science.
  • Matteo Iannacone (San Raffaele Research Institute, Milan, Italy). MD PhD Iannacone directs a group specialized in studying the activation of adaptive immune cells and immunopathology during viral infection and cancer. He has developed specific models to study the dynamics of T cells in the liver in response to viral infection. Iannaccone’s group has recently published novel research that regards liver imaging in the journal Cell.
  • Johannes Textor (Radboud University, Nijmegen, Netherlands). Dr. Johannes Textor leads the Computational Immunology group at Radboud University, employing simulation models, machine learning, and causal inference to study the adaptive immune system. Their research aims to understand immune responses to pathogens and tumors, informing immunological treatments. They develop Artificial Immune Systems to compare immune and nervous system information processing, investigating their distinct architectures and complementary roles. Collaborating with colleagues at Radboudumc's Tumor Immunology department, they provide computational support, advancing immunological research through diverse methodologies.

 

  • Thomas Murooka (University of Manitoba, Manitoba, Canada). The laboratory of Dr. Murooka specialized in the study of T cells and DC as well as their migration during HIV infection in lymphoid tissues. Additionally, they studied the effector and regulatory T cell behaviors during Leishmania major infection in mouse skin. During his career, Dr. Murooka have published high impact factor papers in journals like Nature.
  • Michael Carroll (Harvard Medical School, Boston, USA). The group of Prof. Carroll has made important contributions to the field of antigen trafficking and B cell biology. He has extensive experience characterizing the behavior of DC and B cells in different models including infectious diseases and autoimmune diseases. The group publishes their research in a regular base in the highest impact factor journals in the field including Nature, Science or Cell.

 

Artificial intelligence applied to intravital microscopy of the immune system, 2nd edition 16 - 18 May 2022

This workshop will cover theoretical and practical aspects regarding the application of A.I. -based methods for the analysis of intravital imaging data.
We will focus on the migration and interaction of immune cells observed in vivo, explaining how to get accurate tracking analysis, or how to extract insights from videos without using cell tracking.
Moreover, we will discuss the importance of Open Source and Open Data resources in image-driven immunological research.
The topics and tools presented in this workshop can be applied also for other imaging modalities and cell types but a basic knowledge in microscopy is recommended.

How: Blend modality. Best would be to attend in presence, with practical cases to be solved in small groups. We will provide data and tools and can offer a few workstations to be used but is encouraged to bring your own laptop. You can also bring your own data to be analyzed. For those who cannot come to Bellinzona we will leave open the possibility to connect via Zoom and watch the demonstrations.

When: May 16th – 18th, optional excursion on May 19th

Where: Institute for Research in Biomedicine, Via Francesco Chiesa 5, CH-6500 Bellinzona (Switzerland)
Registration: up to May 8th 2022 23:59 CEST at https://forms.gle/x3XqkdqG7PxMBVH56 15 places available, served on a first come first served basis. You will receive a confirmation via email.

Contact: For questions do not hesitate to contact us at This email address is being protected from spambots. You need JavaScript enabled to view it. with the subject of the email ai-ivm2022

 

DAY 1: May 16th. Introduction to intravital microscopy (IVM).

Learning objectives. Understanding the applications and advantages of intravital microscopy / Understanding the importance of Open Data research / Practice with the immunemap platform

9:00-9:30: Introduction of the tutors and the course (Pizzagalli DU)

9:30-10:15: Ice-breaker activity (Pizzagalli DU)

10:30-11:30: Technical Introduction to IVM (Morone D)

11:30-12:30: Biological applications of IVM (Gonzalez SF)

12:30-14:00: Lunch Break

14:00-15:15: Open Data research and presentation of immunemap (Pizzagalli DU)

15:15-15:45: Coffee break

15:45-17:00: Practical session 1: Use of immunemap (Palladino E / Ceni K / Thelen B)

 

DAY 2: May 17th: Analysis with tracking. 

Learning objectives. Understanding which type of information can be extracted from IVM data using the classical image analysis pipeline, challenges and new perspectives

9:00-10:30: Classical image analysis pipeline, artifacts, quality requirements (Pulfer A)

10:30-10:45 Coffee break

10:45-12:00: The relevance of cell tracking in life sciences, motility parameters (Pizzagalli DU)

12:30-14:00: Lunch Break

14:00-15:15: Practical session 2a: Manual tracking using FIJI/Trackmate (Carrillo-Barbera P), Imaris (Palladino E), immunemap (Ceni K)

15:30-16:00: Practical session 2b: Facilitating automatic tracking using computer-assisted colocalization (Pizzagalli DU)

16:15-17:00: Practical session 2c: Analysis of motility measures (Pizzagalli DU)

 

DAY 3: May 18th: Analysis without tracking.

Learning objectives. Understanding how image processing and computer vision techniques can be applied to analyze IVM data without the usage of cell tracking

9:00-10:30: From pixels to cell actions (Pizzagalli DU)

10:30-10:45 Coffee break

10:45-12:30: Recent trends in computer vision methods (Pulfer A)

12:30-14:00: Lunch Break

14:00-16:15: Practical session 3: Analyzing cell migration without tracking (Motility heatmaps, Action recognition, Optical flow, Colocalization)

16:30-17:00: Conclusion

DAY 4: May 19th: Social activity

09:00 – 12:00: Visit to the castles of Bellinzona

Organizers:
Dr. Santiago Fernandez Gonzalez - Institute for Research in Biomedicine (CH)
Dr. Diego Ulisse Pizzagalli - Institute for Research in Biomedicine and Euler Institute, USI (CH)

Tutors:
Dr. Pau Carrillo-Barbera – The University of Edinburgh (UK)
Kevin Ceni – Institute for Research in Biomedicine (CH)
Elisa Palladino – Institute for Research in Biomedicine (CH)
Alain Pulfer – Institute for Research in Biomedicine and ETH Zurich (CH)
Benedikt Thelen – Euler institute (CH)

 

 

---- PREVIOUS EDITION 2021 ----

Description

This summer school covered theoretical and practical aspects regarding the analysis of immune cell migration and interaction, using intravital microscopy and new A.I.-based methods. We had 18 participants (4 from USI, 14 external), from 4 european countries including BSc, MSc, PhD students, postdocs and imaging specialists.

Organizers. Dr. Santiago Fernandez Gonzalez, Dr. Diego Ulisse Pizzagalli, dr. Pau Carrillo Barbera
Tutors. Diego Morone, Alain Pulfer, Kevin Ceni, Benedikt Thelen
Supported by. IRB PhD program    Cell Migration PhD program    USI - Biomedical PhD program    IMMUNEMAP consortia    Euler Institute

Program

DAY 1: June 14th. Introduction to intravital microscopy (IVM).

Learning objectives. Understanding the applications and advantages of intravital microscopy / Understanding the importance of Open Data research / Practice with the immunemap platform

Introduction and networking activity (Pizzagalli)

1

9:00 - 10:00

Physics of 2-photon intravital microscopy (Morone)

1

10:30 - 11:30

Intravital imaging of the immune system (Gonzalez)

1

11:45 - 12:45

Image formation / histograms / Properties of digital images (Barbera / Morone)

1

14:00 - 15:00

The role of Open Data and Open Source in biomedical research , FAIR principles and IMMUNEMAP project (Pizzagalli / Barbera)

1

15:10 - 15:45

FIJI introduction (Morone)

1

15:45 - 17:00

DAY 2: June 15th: Analysis with tracking. 

Learning objectives. Understanding which type of information can be extracted from IVM data using the classical image analysis pipeline, challenges and new perspectives

Exercise 1

  • Which histogram is saturated
  • Thresholding
  • Use pixel inspector

2

9:00 - 9:30

Image analysis pipeline, quantification of cell motility, dynamism, and interaction (Pizzagalli)

2

9:30-10:30

The possibilities of Machine Learning in IVM (Pizzagalli)

2

10:45-11:30

Excercise 2 (Morone / Pulfer / Barbera Pizzagalli)

  • cell detection (classical / Otsu etc)
  • Thresholding with TWS
  • stardist
  • trackmate
  • automatic tracking with trackmate
  • computation of measures: collective (i.e. preferential direction) and individual cell behavior (i.e. change of speed)

2

14:00 - 17:00

DAY 3: June 16th: Analysis without tracking.

Learning objectives. Understanding how image processing techniques working at pixel-level can be applied to analyze IVM data without the usage of cell tracking

Exercise: track-based measures on neutrophil chemotaxis

Exercise: migration differences between WT and KO cells in spleen

 

9:00 – 10:30

Discussion Exercise 1 (Pizzagalli)

Contact analysis in Exercise 1 (Pizzagalli)

Advanced analysis without tracking (of/recruitment) (Pizzagalli)

3

10:45 - 11:00

Image processing techniques (Pulfer)

3

11:15 - 12:15

Exercise 3

  • Quantify the recruitment in vessels
  • Analyze whole LN images with OF
  • Apply the bayes coloc plugin by Alain (cell-cell interaction)
  • Imaris workshop (tracking, coloc, heatmaps, and cell-cell interaction)

3

14:00 - 17:00

DAY 4: June 17th:  Beyond tracking.

Learning objectives. Understanding how computer vision methods for action recognition can be applied to analyze IVM data without the usage of cell tracking

Inside AI: neural networks and clustering (Pizzagalli)

4

9:00 – 10:00

Trends in computer vision: classic, deep learning, generative (Pulfer)

4

10:05 - 11:00

Action recognition applied to immune cells (Pizzagalli)

  • Review of motility patterns displayed by immune cells in vivo and their biological meaning
  • Application of action recognition to quantify neutrophil dynamics and migration morpho-phenotypes

4

11:15 - 12:15

Exercise 4 (Pulfer / Pizzagalli)

  • Usage of the apoptosis detection program (incl. challenge)
  • Discussion on the limits and potential usage of Deep Learning in IVM
  • Neutrophil swarm detection

4

14:00 - 16:00

Discussion on good practices for IVM analysis (Barbera + Pizzagalli)

4

16:15 - 17:00

DAY 5: June 18th: Workshop and conclusion

Workshop: analyze data by participants (Pizzagalli):

- Quantification of leukocyte migration within and outside blood vessels via track-based measures and pixel classification.

- Analysis of DCs – Macrophage interaction via supervised spectral unmixing, contact analysis, and bayesian colocalization.
- Quantification of neutrophil swarm dynamics via unsupervised machine learning.

5

9:00 - 12:00

Exam

5

14:00 - 15:00

Concluding remarks

5

15:30 - 16:30

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