Workshop 4

Phenotypic screening of complex cellular or whole organism model systems is increasingly employed in biomedical and pharmaceutical research. To fully exploit their potential, novel imaging workflows and tailored tools compatible with the automated imaging and spatiotemporal scoring of cell- and tissue-specific phenotypes are needed.
The ACQUIFER Imaging Machine (IM) is a versatile and flexible automated wide-field fluorescence microscope for high-content imaging of 2D/3D cell-cultures, fixed tissues sections and small model-organisms. The system features a unique moving optical unit in combination with a static sample holder, rendering it ideal for non-adherent and motion-sensitive specimen. For optimal time-lapse in-vivo imaging, the sample chamber is equipped with a Peltier device for highly precise temperature control.
In this workshop, we will demonstrate the unique hardware design and exemplify automated imaging applications. We will illustrate our workflow concept for automated imaging, feedback microscopy and High-Content-Screening using the ACQUIFER IM and associated software tools. Furthermore, we will discuss the optional laser-module that allows scaling-up of challenging photomanipulation experiments, such as photodamaging or photoconversion.
The IM control software ensures simple and intuitive configuration of standard imaging protocols. Complex acquisition procedures can be customized thanks to built-in scripting support (C#-based) or via dedicated extensions for external applications (Fiji, Python etc). The powerful imaging capabilities of the IM are complemented by the ACQUIFER PlateViewer, an easy-to-use software for browsing and visualization of large high-content screening datasets, and an interface for supervised feedback microscopy. The PlateViewer includes a plugin mechanism, allowing interfacing with external software for custom image-analysis. The open interface enables remote-control of the system by external software or integration in a robotic setup. It also allows feedback microscopy workflows for pre-scan/re-scan applications, such as automated centring applications, using Fiji and the PlateViewer.

Refer to our website at www.acquifer.de

One of the core facilities’ duties is to provide end-users, usually researchers in life sciences, a fleet of microscopes at a level of performance compatible with their experiments. This is not an easy task because the performance of such devices tends to fluctuate or deteriorate over time for many reasons: misusing, aging, environment fluctuations, etc. This is especially true for high end imaging systems such as confocal or super resolution microscopes. Having access to a unique, reliable, and easy-to-use device to ensure microscopes’ performance would certainly make easier this tedious task.

To get quantitative and reproducible data, assessing the performances of fluorescence microscopes is a prerequisite before any imaging campaign. For example, system co-registration accuracy should be evaluated before any co-localization study; System field uniformity and intensity response before any study where intensity in the image matters; Spatial resolution before any study aiming at counting objects close to each other, etc.

Three years after our last presence at ELMI, we are happy to come back and present during this workshop the novelties we have developed in the meantime: improved hardware products, new analyses from 3D patterns, new analyses related to non-Argolight products, improvements in user interface and user experience, etc. Above all, the presentation aims to show how the quality control of fluorescence microscopes can be standardized with Argolight software solutions, and how the generated quality control data can be managed and centralized for later reporting.

In this workshop we will demonstrate Bruker super-resolution and light-sheet microscopy solutions (Luxendo SPIM family).
The SPIM demonstration will cover different sample types and potential applications ranging from life imaging of delicate live specimen such as organoids and embryos (mouse, zebrafish etc. ) to whole in-toto imaging of large cleared samples such es entire adult mice. We will be showing our 3D Super-resolution system, the VXL with patented 3D technology suitable for STORM and PALM, along with an integrated
microfluidics system for highly multiplexed, DNA-PAINT research.

Workshop 1/4 : TruLive3D SPIM – light-sheet microscopy for live samples
Workshop 2/5 : VXL – 3D super-resolution microscopy
Workshop 3/6 : LCS SPIM – light-sheet microscopy for cleared samples

Image:

Top, left image: live zebrafish embryo H2A::GFP recorded on Luxendo MuVi SPIM LS.
Top, right image: Mitochondria stained for TOM20 with Alexa 647 and imaged with STORM. Bottom, left image: cleared adult mouse with GFP labeled nerves and recorded on Luxendo LCS SPIM.
Bottom, right: Cleared YFP expressing transgenic mouse brain recorded on Luxendo MuVi SPIM CS

High content screening (HCS) is increasingly adopted for experiments demanding the statistical power of many repeats and the ease of management of thousands of specimens, from acquisition through to automatic analysis, quantification and data management. Whilst drug discovery has been one of the key drivers in the widescale adoption of HCS, many other applications now also benefit from similar approaches for the generation of robust experimental datasets and results.

Robust analysis relies on high image quality of the raw data and powerful image analysis solutions that quickly and reproducibly extract the required data from each image. Celldiscoverer 7 is a powerful HCS boxed microscope solution that uses unique optics to generate unmatched image quality. Teaming these high-quality images with the comprehensive data management solutions provided by arivis VisionHub generates a powerful HCS engine with the flexibility needed for a wide range of applications, from 2D single cells to 3D organoids.

Join this workshop to see the entire HCS workflow in action and experience the benefit of teaming high image quality with powerful data management.

Fast and highly resolved volumetric microscopy is essential to examine various biological aspects in their complexity, ranging from whole animal imaging, to live phenomena, to thick samples to image at the different depth. During this workshop we demonstrate how X-light V3 spinning disk and DeepSIM add-on can be combined to work with deep samples of thickness comparable to those normally used in CF microscopy. This means that more meaningful data can be obtained from native heterogeneous complex (over 100µm thick) samples using routine preparation protocols.
#deepimaging #clearedtissues #spinningdisk #SIM

Characterising, comparing, and ultimately understanding, differences in dynamic live cell behaviour is critical in many fields, from cancer research to regenerative medicine.

In this workshop, we explore new ways to characterise cell behaviour, enabled by the Livecyte Kinetic Cytometer system. We look at how simple label-free assays, in 96-well plates, can be used to independently measure cell growth and cell proliferation, automatically characterise both random and sheet cell migration and identify other phenotypic changes. We also show Livecyte’s correlative fluorescence capabilities, and how these measurements unlock additional powerful new insights.

Presenter:
Bülent Peker, Evident Europe

Organoid imaging requires a lot of time, not only capturing images in 3D but also finding target objects especially if the vessel is microplate. With its ability to create excellent low magnifaction images the OLYMPUS confocal laser scanning microscope FLUOVIEW FV3000 has an advantage for organoid imaging, as this feature can be used for creating overview maps to find objects for high resolution imaging with higher magnification.

We automated the process by the integration of an object detection algorithm, and the connection of manual processes into a single step process. This means you can walk away from the system once you set up and start your experiment. No time required to find objects in each well anymore. The FV3000 automatically captures images at low magnification and then finds your objects of interest automatically and captures them at high magnification. This automated process dramatically reduces your time for microscope operation.

High-resolution microscopy of cells requires imaging chambers that are specifically tailored to your application.

Are you interested in learning about…
… easy to handle all-in-one solutions for live cell imaging?
… the advantages of using channel slides for immunofluorescence stainings?
… solutions for 3D cell culture and spheroid imaging?

Join our workshop and find the answers to these questions!

First, we will introduce a variety of ibidi chambers with different geometries and surfaces and help you to choose the products that are best suited for your assay. This will be followed by a hands-on-training where you get the chance to test a selection of our labware for 2D and 3D cell culture and imaging.

Confocal microscopy has become a staple for fluorescence imaging in the life sciences, owing to the optical sectioning, sensitivity, and the temporal and spatial resolution capabilities. With STELLARIS, we have introduced key innovations such as the Power HyD family of detectors (1), the next generation of white light lasers (WLL), and the TauSense technology (2).
The fully digital and photon counting proprietary approach, Power Counting, allows straightforward and robust quantification of fluorescence signals in STELLARIS. Together with the WLL excitation flexibility, STELLARIS delivers extended near infrared capabilities for expanding the palette of fluorophores in every experiment., In addition, . TauSense opens the door to functional imaging insights thanks to straightforward access to fluorescence lifetime-based information.

In this workshop we will learn how all the technological innovation in STELLARIS facilitate fluorescence quantifications . We will also unveil a new TauSense tool built to evaluate molecular interactions. Join us for a first-hand view of what STELLARIS can bring to functional imaging.

1. V. Schweikhard, L. A. J. Alvarez, I. Steinmetz, M. J. Roberti, Holger Birk, A. Giske. The Power HyD family of detectors. Nat. Methods. 2020. Doi: d42473-020-00398-0 (2020).
2. M. J. Roberti, L. Ortiz Lopez, G. Ossato, I. Steinmetz, P. Haas, F. Hecht, and L. A. J. Alvarez. TauSense: a fluorescence lifetime-based tool set for everyday imaging. Nat Methods. 2020. Doi: d42473-020-00364-w

Visualizing the three-dimensional architecture of complex organisms while analyzing biological processes in a whole-organ fashion are becoming the new standard in research. To provide a complete, smooth, and hassle-free 3D imaging workflow, Miltenyi Biotec covers this entire process offering solutions for sample staining, clearing and imaging. In this workshop, we will guide you through the most important steps of sample preparation, including a live demonstration, and will demonstrate how easily high quality 3D images of large samples can be acquired with our cutting-edge light sheet system, the UltraMicroscope BlazeTM.

In this workshop, we will illustrate how deep learning can be implemented in microscopy. Hands-on will be provided with a software environment, which does not require specific computing configuration or programming skills. Examples will include signal to noise and contrast enhancement, image prediction, and segmentation of objects in images. After this workshop, you will be able to train your own neural network with NIS-Elements for your own application. Possible implementation in feedback microscopy will be discussed. One-to-one or small group sessions can be arranged on request. Visit us at our booth and let us know if an additional session works better for your schedule!

Bring your own sample to the conference to image it with Prospective Instruments MPX series Two-Photon Microscope. Contact us if you would like to take this opportunity.
Of course, we will also bring a selection of samples to image.

Deconvolution, Restoration and Analysis of all your microscopy images Everywhere.

Tissue Cytometry permits to determine the in-situ phenotype of cells as well as histological entities, like glands, vessels or tumor foci. Applications include but are not limited to the exploration of the cellular/tumor microenvironment and/or the spatial organization of cellular subpopulations, assessment of different bone structures, quantification of blood vessels and neovascularization as well as analysis of samples in multiplexing or multispectral mode.
Earlier attempts to analyse single cells in tissue have mostly been subject to visual estimation, or – at best – to manual counting for decades. Hence, experts usually had the choice of the “least of evils” between guessing and endless (manual) counting. In (tumor) immunology, infiltrating inflammatory cells need to be phenotypically characterized on a quantitative basis. To better understand the function of inflammatory cells in tumor development, type and number of inflammatory cells and their proximity to glandular/tumor structures have to be analyzed in-situ and correlated with disease state. Using TissueFAXS™ Cytometry the time-consuming and error-prone human evaluation of stained histological sections can be approached with an observer-independent and reproducible technology platform, offering a high degree of automation, paired with user interaction at relevant points of the analytical workflow. TissueFAXS Cytometry is the “one-stop shop” for all kinds of microscopic analysis of tissue sections, organoids and adherent cell cultures.

Studying spatial protein organization or protein copy number, clustering and mobility at the molecular level is extremely important from both a diagnostic and treatment perspective. Looking at cellular structures beyond the diffraction limit with a resolution of 15-20 nm with robust quantification has been enabled with Single-molecule localization microscopy (SMLM) like STORM or PALM. ONI has created the world’s first desktop super-resolution microscope, the Nanoimager, which is designed to operate on a standard lab bench and has a footprint smaller than a piece of A4 paper, with the most intuitive and simple software interface, making it also accessible to researchers not specialized in high end microscopy techniques with one major goal: make it accessible and simple to use.

It is really important to enable users to extract quantitative information from their localization-based images. We have done this by designing a cloud-based analysis software called CODI (COllaborative DIscovery). CODI helps users analyze their single-molecule data by extracting quantitative information on protein clustering, cluster size and area, cluster density, and so much more. It enables all researchers to upload their data, share or collaborate with other scientists, whilst gaining meaningful insights about their data.

During this workshop we will perform a two-color STORM on mitochondrial samples (TOM20 and HSP60) and reveal the sub-mitochondrial compartmental organization with 20 nm resolution. The aim is to introduce SMLM imaging as a powerful tool for rapid and multiplexed characterization of cellular structures along with a visual and quantification analysis platform CODI developed by us. We will follow up the workshop with an open discussion on the wide-ranging applicability of the Nanoimager in different biological applications encompassing the field of immunology, neurobiology, cell biology and cancer.

1. AIMS
This workshop will give the listener an overview of new developments in Spinning Disk Microscopy and complementary micromanipulation and imaging methods.

2. SCOPE
Spinning Disk Confocal Microscopy allows researchers to image fast processes in living cells with minimum photobleaching [1]. This makes it an ideal imaging approach to combine with advanced sample manipulation methods like Fluorescence Recovery After Photobleaching (FRAP), Photoactivation (PA) and Ablation.
Over the past two decades, Visitron Systems GmbH has been pioneering new technical developments in this field. One recent result of this effort is an integrated FRAP/Ablation System that can be combined with TIRF and Widefield Imaging all on the same port. At the click of a button, researchers can now cut microscopic structures in 3D space, image membrane processes in TIRF mode and rapidly switch to fast confocal imaging to detect cellular events in confocal mode.
The new Yokogawa-SoRa optics for Super-Resolution based on via Optical Re-Assignment technique with realtime 2D deconvolution processing.