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Quality Assessment and Reproducibility for Instruments & Images in Light Microscopy

The Consortium for Quality Assessment and Reproducibility for Instruments and Images in Light Microscopy (QUAREP-LiMi), formed by the global community of practitioners, researchers, developers, service providers, funders, publishers, policy makers and industry related to the use of light microscopy, is committed to democratising access to quantitative and reproducible light microscopy and the data generated by it.

Our mission is to provide a comprehensive set of community-agreed guidelines, protocols, automation procedures and other resources to improve quality control, quality assurance, and instrument/method calibration. The stakeholders advocate for the use of the highest level of reproducibility for data generated by light microscopy methods in scientific research and biotechnology applications. To support the achievement of its mission, QUAREP-LiMi:

 

  1. sanctions Working Groups to form teams that focus on specific technical or outreach topics,
  2. maintains, disseminates, and teaches best practices, protocols, and software tools and
  3. collaborates with providers of reference materials or instrumentation or other tools that facilitate the implementation of best practices at the community level.

 

QUAREP-LiMi operates a NextCloud and an OMERO server, which serve as a central repository and exchange point for our collective work. These platforms facilitate collaboration and data sharing between our members. 

QUAREP-LiMi is open to new members, and we look forward to your contribution. If you are interested in joining our initiative, please fill out our membership form. If you would like to change your Working Group (WG) or join a new WG, please use our form. For any further questions, please contact us.

Breaking News

The Light Microscopy Metadata Model (LiMi-Model): Enabling the Generation and Sharing of Reproducible, Community-Harmonized FAIR Bioimage Data (QUAREP-LiMi WG7-Matadata) ELMI Workshop
🎤 Caterina Strambio De Castillia

Presenter(s): Caterina Strambio-De-Castillia *1

*- the workshop will presented on behalf of QUAREP-LiMi WG7-Metadata.

1. Program in Molecular Medicine, UMass Chan Medical School

A major output of QUAREP-LiMi is the Light-Microscopy Metadata Model (LiMi-Model) [1, 2], developed by the QUAREP-LiMi Metadata Working Group (WG7) in collaboration with BioImaging North America, the 4D Nucleome consortium and OME. The LiMi-Model was designed to harmonize the description of light microscopy hardware, acquisition settings, and quality-control metrics in order to enhance image quality and reproducibility, and to fulfill the FAIR (Findable, Accessible, Interoperable, and Reusable) data principles.

Specifically, the LiMi-Model aims to: (1) promote the harmonized generation and pre-publication management of image datasets that are FAIR by design; (2) facilitate the deposition of FAIR microscopy datasets to public image data repositories; (3) facilitate data reuse and the extraction of quantitative information from image data using advanced bioimage analysis techniques, including AI/ML; and (4) define the light microscopy implementation of the Image Acquisition module of the Recommended Metadata for Biological Images (REMBI) guidelines [3].

The workshop will open with a general introduction to the purpose and scope of the LiMi-Model, situating it within the broader landscape of community initiatives. These include the recently published minimal reporting recommendations for light microscopy [4], REMBI, the International Standards Organization, imaging modality ontologies (such as FBbi and EDAM-bioimaging), and an emerging vision for next-generation metadata schema definition languages. This will be followed by a presentation of the LiMi-Model website [1] and interactive LiMi-Model Viewer, a tool designed to help community members navigate and understand the content of the model. The workshop will conclude with an overview of the LiMi-Model’s governance structure and an explanation of how community members can engage with and contribute to its ongoing development. As a concrete example of this process, the workshop will introduce the recent Camera Module revision, conducted in partnership with camera manufacturers whose outcomes will also be presented in a related poster presented in collaboration with QUAREP-WG2.

The formal presentations will be followed by an open discussion in which attendees are encouraged to provide feedback and help shape the future direction of the LiMi-Model.

[1] https://quarep.org/working-groups/wg-7-metadata/limi-model

[2] https://doi.org/10.1038/s41592-021-01327-9

[3] https://doi.org/10.1038/s41592-021-01166-8

[4] https://doi.org/10.1083/jcb.202601032

WG4 Workshop at ELMI 2026: “Assessing the co-registration accuracy of a microscope”
🎤 Hans Fried1, Roland Nitschke2, Kees van der Oord3, Maria M. Azevedo4,5

1. CRFS – Light Microscope Facility, German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany

2. Life Imaging Center and Signalling Research Centre CIBSS, University of Freiburg, Germany

3. Nikon Europe BV, Amstelveen, Netherlands

4. Advanced Light Microscopy Scientific Platform, i3S – Instituto de Investigação e Inovação em Saúde | Porto, Portugal

5. University of Porto

Accurate multi‑colour imaging requires that signals from the same physical location co‑localise across channels. Chromatic aberration is a major source of spatial error in multi‑colour fluorescence microscopy. Therefore, rigorous co‑registration is essential for reliable interpretation of spatial relationships in biological specimens.

In this workshop, we will briefly explain the key steps of preparing a bead sample for co-registration assessment, show the main considerations for image acquisition (1), and show you how to analyze the obtained images using Fiji/ImageJ (2). We will also demonstrate how specific objective‑related factors can influence co‑registration performance. Finally, we will share practical hints and tips to support routine co‑registration analysis.

Watch Presentation

(1) Dauphin A., Azevedo, M., et al. Protocols.io, Ensuring accurate co-registration measurement for quality control of Single Point Confocal Laser Scanning Microscopes – V1, dx.doi.org/10.17504/protocols.io.q26g7yrj8gwz/v1

(2) Faklaris, O. et al. Quality assessment in light microscopy for routine use through simple tools and robust metrics. J Cell Biol 221, doi:10.1083/jcb.202107093 (2022).

WG5 Workshop at ELMI 2026: “Monitoring Microscope Performance Using PSF Measurements”

🎤 Glyn Nelson, Ioannis Alexopoulos, Yury Belyaev

The Point Spread Function (PSF), which describes the response of an optical system to a point source, is a key quality control of a microscope. Regular measurements, conducted with consistent tools, methods, and protocols, along with the calculation of robust metrics enable the monitoring of the microscope performance and therefore ensure better reproducibility of scientific experiments. During this workshop, we will show briefly how to prepare a bead slide, how to perform an acquisition, and finally how to analyze the PSF with open-source tools (1, 2). We will give some tips to troubleshooting PSFs that have an abnormal shape or are far from the theoretical expected size. We will also demonstrate the upload of the analysis results to OMERO, a database that will help monitor PSFs over time. We will use the protocols and metrics that are defined in the framework of the QUAREP-LiMi consortium’s WG5 (3).

Watch Presentation

1. Faklaris O., et al. “Quality Assessment in Light Microscopy for Routine Use through Simple Tools and Robust Metrics.” Journal of Cell Biology 221, no. 11 (2022): e202107093. https://doi.org/10.1083/jcb.202107093.

2. https://github.com/MontpellierRessourcesImagerie/MetroloJ_QC

3. Nelson G., et al. Protocolos.io, Monitoring the point spread function for quality control of confocal microscopes , dx.doi.org/10.17504/protocols.io.bp2l61ww1vqe/v1

WG1, WG2 and WG6 Present Workshop on Automation of QUAREP-LiMi Protocols at ELMI 2026
 
🎤 Britta Schroth-Diez, Kees van der Oord

a Nikon Europe B.V.

b Light Microscopy Facility, Max Planck Institute of Molecular Cell Biology and Genetics, Dresden Germany

The QUAREP-LiMi Tool Kit [1] was developed by the QUAREP-LiMi community to reduce the time needed to run periodic quality assessment protocols and organize the performance data acquired. During this workshop we will present its basic features and latest developments.

In accordance to the protocol defined by the QUAREP-LiMi WG 1 [2] the Tool Kit integrates user friendly interfaces for the assessment of illumination power and stability. These interfaces are already available for the Nikon and Zeiss acquistion software.

In addition, the Tool Kit includes support for the protocols for detector gain, dynamic range and noise assessment developed by the QUAREP-LiMi WG2 [2].

A highlight of the Tool Kit is its data browser. Thanks to the organized presentation of the performance metrics it is easier to diagnose problems – e.g. a small but steady power decrease over months– and take corrective and preventive actions before they become detrimental.

The Tool Kit functionality will be demonstrated during the workshop, where we will emphasize important details of the protocols.

Watch Presentation

[1] https://github.com/QUAREP-LiMi/QUAREP-LiMi-Tool-Kit

[2] https://www.protocols.io/workspaces/quarep-limi/publications

At ELMI 2026, WG2 hosted the workshop “Estimation of the Photon Conversion Factor, Noise and Dynamic Range of Light Microscope Detection Systems”
 
🎤 Britta Schroth-Diez, Kees van der Oord, Ruediger Bader

1. Light Microscopy Facility, Max Planck Institute of Molecular Cell Biology and Genetics, Dresden Germany

2. Nikon Europe BV

3. Hamamatsu Photonics Europe

QUAREP-LiMi Working Group 2 (WG2) developed a set of protocols [1] which focuses on the characterization and monitoring of the performance of light microscope detection systems, which collects light from the sample at the microscope and helps to convert the “arbitrary digital units” (ADU) provided by the microscope into “number of photons” detected.

The protocols collection is built around the photon transfer curve (PTC) method [2]. The goals of these protocols are to achieve 1) experiment quality control, 2) monitoring instrument quality over time, and 3) detailed detection system characterization. The protocols include both point and area detectors, allowing researchers to select appropriate methods based on their specific detection systems.

During the workshop, we will demonstrate how to use the protocols to acquire images and upload them in the software [3] to characterize the photon conversation factor (PCF photons/ADU), readnoise, background and dynamic range of a light microscopy detection system. The set includes protocols on sample preparation, data acquisition, and analysis. Furthermore, we will demonstrate its integration in the QUAREP-LiMi Tool Kit [4].

Watch Presentation

[1] Characterization of the Photon Conversion Factor, Noise, and Dynamic Range of Light Microscope Detection Systems – https://dx.doi.org/10.17504/protocols.io.14egn61pyl5d/v1

[2] Janesick JR. 2007. Photon Transfer. SPIE. https://dx.doi.org/10.1117/3.725073

[3] McFadden D. 2022. GUI Calibration Tool. https://github.com/bionanoimaging/NanoImagingPack/releases

[4] QUAREP-LiMi Tool Kit, https://github.com/QUAREP-LiMi/QUAREP-LiMi-Tool-Kit

Daniel Schröder Webinar (QUAREP-LiMi WG3): Universal Laser Engine with an Improved Homogeneous Illumination Profile

QUAREP-LiMi WG3 was pleased to host a webinar presented by Daniel Schröder on the topic of the "Universal Laser Engine" with an improved homogeneous illumination profile.
The webinar introduced a versatile laser excitation platform designed for advanced fluorescence microscopy and highlighted its applications across multiple imaging modalities.

Abstract

The Universal Laser Engine is a user-friendly laserbox designed to provide a versatile excitation for advanced fluorescence microscopy. In a single box, it holds up to 6 laser wavelengths operating in modes ranging from picosecond-pulsed to continuous-wave, enabling its use across a wide range of imaging modalities. The Universal Laser Engine has been successfully tested in widefield microscopy, super-resolution localization microscopy, total internal reflection fluorescence (TIRF), and fluorescence lifetime imaging microscopy (FLIM). In addition, the Universal Laser Engine incorporates a novel speckle-suppression technology that generates a highly homogeneous flat-top hat profile that is applicable for high-speed imaging at exposure times down to microseconds. The system provides various trigger inputs and two fiber outputs for seamless integration into both existing and new microscopes. By combining flexibility, compact design, and state-of-the-art illumination uniformity, the Universal Laser Engine offers a powerful and adaptable solution for simplifying and modernizing fluorescence microscopy instrumentation.

Watch the webinar recording

Join the QUAREP-LiMi Workshops at ELMI

We are pleased to announce that the QUAREP-LiMi consortium will be presenting five specialized workshops at the upcoming European Light Microscopy Institute (ELMI) meeting. Our sessions are designed to provide researchers with frameworks for improving image quality, ensuring data reproducibility, and implementing rigorous quality control standards in microscopy. The workshops will cover five critical pillars of instrument characterization:

  1. The QUAREP-LiMi Tool Kit (WGs 1 & 2):

    Discover how to streamline periodic quality assessments. We will present the latest developments in the Tool Kit, including user-friendly software tools for illumination power and stability (compatible with Nikon and Zeiss), as well as its data browser for long-term performance monitoring.

  2. Detector Performance & Photon Characterization (WG2):

    Learn how to transition from "arbitrary digital units" (ADU) to actual photon counts using the Photon Transfer Curve (PTC) method. We will demonstrate protocols for characterizing photon conversion factors, read noise, and dynamic range for both point and area detectors.

  3. Assessing the co-registration accuracy of a microscope (WG4):

    Chromatic aberration is a major source of spatial error in multi‑colour fluorescence microscopy. This workshop will show practical hints and tips to perform routine co‑registration measurement and analysis.

  4. Point Spread Function (PSF) Analysis (WG5):

    This workshop provides a practical guide to PSF measurement—from beads slide preparation and image acquisition to analysis using open-source tools and integration with OMERO for longitudinal monitoring.

  5. The Light-Microscopy Metadata Model (LiMi-Model, WG7):

    In collaboration with BioImaging North America, 4D Nucleome, and OME, we will introduce the LiMi-Model. This framework aims to harmonize the description of hardware and acquisition settings to ensure datasets are FAIR (Findable, Accessible, Interoperable, and Reusable) by design.
We look forward to engaging with you and hearing your feedback during these sessions!

New article about minimal reporting requirements published

A new article, under the guidance of QUAREP-LiMi WG 11, on the minimal reporting requirements for light microscopy data has been published in the Journal of Cell Biology. You find the article here.

More than 700 members have joined QUAREP-LiMi

QUAREP-LiMi has reached another milestone. By the end of 2025 more than 700 members have joined QUAREP-LiMi. When we launched this initiative, we never imagined the community would grow to include so many members from 46 different countries around the globe. A heartfelt thank‑you to all our members for your continued support and contributions.

Ensuring accurate co-registration measurement for quality control of Single Point Confocal Laser Scanning Microscopes – V1

We are thrilled that WG 4 - System chromatic aberration and co-registration Power published its first protocol "Ensuring accurate co-registration measurement for quality control of Single Point Confocal Laser Scanning Microscopes - V1". The protocol is featured on protocols.io

QUAREP Calendar

 

 

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