COMPARE Workshop on Single-Molecule imaging and spectroscopy of membrane proteins
March 12th – 13th 2019
University of Birmingham
We are very pleased to announce our 2019 COMPARE Workshop, which this year will focus on Single-Molecule Imaging & Spectroscopy of Membrane Proteins.
Single-molecule microscopy and spectroscopy are very powerful techniques that allow investigating the organization and dynamics of membrane proteins in cells and tissues with unpreceded spatiotemporal resolution. This workshop will bring together world leaders in single-molecule imaging and spectroscopy to focus on key questions about the spatiotemporal organization of receptors and other signalling proteins in nanodomains on the plasma membrane.
Day 1 - Tuesday 12th March 2019
A first day of talks covering both recent exciting biological findings and new methodological developments will be followed on day two by unique practical sessions. These will include in-depth demonstrations of cutting-edge single-molecule microscopy and spectroscopy techniques in collaboration with leading microscopy vendors as well as of new open source software for the analysis of spectroscopy and single-molecule data.
|08:45 - 09:25
||Registration and Coffee
|09:25 - 09:30
||Welcome: Steve Thomas and Davide Calebiro
||Research session I: Single Molecule microscopy – Chaired by Steve Thomas
||HAWK analysis of localisation microscopy data, Susan Cox, KCL
|10:00 - 10:30
||Super-resolution imaging to study four-transmembrane proteins, Annemiek van Spriel, Radboud Institute for Molecular Life Sciences
||Quantitative analysis of single-molecule localization microscopy: A Voronoï tessellation story, Florian Levet, University of Bordeaux
||Research session II: Single Molecule microscopy – Chaired by Natalie Poulter
||Live-cell SMLM: tracking protein nano-cluster dynamics and interactions with sub-second resolution, Juliette Griffié, EFPL
||Optimising single molecule Localization experiments using NanoJ-SQUIRREL, Siân Culley, UCL
||Industry Session 1 - Chaired by Natalie Poulter
||Single Molecule Super-Resolution Imaging, Kate Lewis, NIKON
||Entering the nanoworld: imaging single molecules in 3D, Camille Clement, Abbelight
||The Nanoimager, Siobhan King, ONI
||Research session III: Single molecule dynamics – Chaired by Davide Calebiro
||Heterogeneous lateral diffusion of membrane receptors, Carlo Manzo, Universitat de Vic
||TRamWAy: probabilistic pipelines and unsupervised approaches to single biomolecule dynamics, Jean-Baptiste Masson, Institute Pasteur
||The dynamic nano-organization of synapses tunes synaptic transmission properties, Benjamin Compans, KCL
||Research session IV: FCS – Chaired by Dee Kavanagh
||Combining imaging and pharmacology: subcellular monitoring of receptors, ligands and their interactions, Steve Briddon, University of Nottingham
||Oligomerization of class C GPCRs in neurons, Emmanuel Margeat, Montpellier
||Industry session II – Chaired by Jeremy Pike
||The Elyra 7: Imaging at the speed of life, Nick Sergent, Zeiss
||3D SML in thick samples, Marc Koch, Bruker
||Spinning TIRF for Simultaneous Multi-Channel Single Molecule Imaging, Martyn Reynolds, Cairn
||Scientific Cameras for Single Molecule Imaging, Louis Keal, Photometrics
||Close of Day 1 – Davide Calebiro
Day 2 - Wednesday 13th March 2019
Venue: The Imaging Suite, Institute of Biomedical Research, Medical School
Attendees can choose up to 5 workshops for day 2 (maximum of 3 microscopy workshops).
Microscopy based workshops
Workshop 1: Using N-STORM for Single Molecule Location Microscopy.
Hardware: NIKON N-STORM.
Microscopy Specialist: Kate Lewis, NIKON.
Overview: This workshop will focus on the using the NIKON N-STORM system for Single Molecule Location Microscopy. 2D and 3D single molecule acquisition and analysis will be demonstrated.
Workshop 2: Spinning TIRF for Simultaneous Multi-Channel Single Molecule Imaging.
Hardware: Cairn Quad TIRF.
Microscopy Specialist: Martyn Reynolds, Cairn.
Overview: The Cairn TIRF system on demo is configured with a fast scanning TIRF/HILO head for excitation with detection from up to four cameras simultaneously. We will walk through the use of the Cairn spinning TIRF system and use the resulting high uniformity illumination for quantitative TIRF imaging, followed by a demonstration of single molecule localization approaches, including single particle tracking, that can benefit from the spinning TIRF approach.
Workshop 3: Evaluating Scientific Cameras for Single Molecule Imaging.
Hardware: Photometrics CMOS and EMCCDs, NIKON Ti2 Eclipse, Cairn TwinCam.
Microscopy Specialist: Louis Keal, Photometrics.
Overview: Learn more about how to evaluate camera technology for single-molecule imaging applications. We’ll also be performing a live comparison between back-illuminated CMOS cameras and EMCCD cameras to demonstrate their effectiveness for low light imaging.
Workshop 4: The Nanoimager: A demonstration of single-molecule localization microscopy carried out on a desktop compatible super-resolution microscope.
Hardware: The Nanoimager
Microscopy Specialist: Siobhán King, ONI.
Overview: A workshop on ONI’s Nanoimager offering various modes of operation including single-molecule localization microscopy (SMLM), TIRF, smFRET, smTracking, dSTORM and PALM. We will provide a brief overview of the Nanoimager and its applications in single-molecule localization microscopy and invite you to test out the system with your own samples.
Workshop 5: Application of FCS and FCCS to investigate receptor dynamics.
Hardware: Zeiss LSM780.
Microscopy Specialist: Nick Sergent, Zeiss.
Overview: This workshop will give an introduction into the background of FCS and FCCS, followed by a live demonstration of technique to probe molecular diffusion and interactions.
Software based workshops
Software workshop 1: Coordinate-based quantification of 1 and 2 color single-molecule localization microscopy data.
Developer: Florian Levet
Overview: In this workshop we will present various analytical methods designed to quantify single-molecule localization microscopy (SMLM) data directly from the localization coordinates. It will focus on various freely available methods (DBSCAN, CBC, Clus-Doc, Voronoï-based quantifications, K-Ripley’s function, etc.), presented in the framework of SR-Tesseler (http://www.iins.u-bordeaux.fr/team-sibarita-SR-Tesseler). We will discuss the pros and the cons of the presented methods by summarizing their capabilities. It aims at guiding users for choosing the most appropriate methods to their problem, whether it be for structural or colocalization analysis. As a support, we will use simulated and experimental data.
Software workshop 2: Image Quality Assessment with NanoJ-SQUIRREL: Data Preparation, Running the Software and Interpreting the Results
Developer: Sian Culley
Overview: Further details can be found at https://www.nature.com/articles/nmeth.4605.
Software workshop 3: HAWK analysis of localisation microcroscopy data
Developer: Susan Cox
Overview: HAWK analysis allows localisation data to be analysed with confidence that the results will not contain artifacts. This workshop will cover the details of what HAWK analysis does and practical demonstrations of the results it gives on different data of varying quality. Participants are welcome to bring their own data to analyse during the workshop.
Software workshop 4: The TRamWAy Python library: inferring maps of local diffusivity and force from single molecule trajectories
Developer: Francois Laurent
Overview: After an introduction on Bayesian inference of single molecule dynamics, we will install the TRamWAy open-source library from GitHub (available at https://github.com/DecBayComp/TRamWAy) and run standard use cases in Jupyter notebooks, including:
- loading simulated and experimental sets of trajectories.
- segmenting space and time.
- generating and visualizing maps of dynamical parameters such as diffusivity and force.
- determining the significance of a force using a Bayes factor technique.
We will face several limiting experimental conditions and give some guidance.
Software workshop 5: RSMLM: An R-package for pointillist based analysis of single molecule localization microscopy data
Developer: Jeremy Pike
Overview: This workshop will use Jupyter notebooks to introduce RSMLM. This R package allows users to easily and efficiently design customised analysis workflows for high throughput and efficient analysis of complex 2D and 3D single molecule datasets. We will cover commonly used cluster analysis techniques as well as more advanced methods for quantifying the shape or topology of biological nano-structure. For more details see our pre-print: https://www.biorxiv.org/content/10.1101/400275v2.
To register for this event, please visit the University of Birmingham Shop
Registrations can be made for both days or for day 1 only.
Please note, there will be a limit of 80 spaces for day 2
If you have a disability, specific access requirements or any special dietary requirements we need to be made aware of please contact the organisers using the details above.
We look forward to welcoming you to Birmingham in March 2019
Prof. Davide Calebiro
Dr Steve Thomas
Dr Dee Kavanagh
Dr Jeremy Pike
Dr Natalie Poulter
The organising committee would like to thank our industry partners for their generous support.