WG 1 – Illumination power

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WG 1 – Objectives

Comparison of fluorescence intensities between images requires measurements of the illumination power and stability of the excitation light source. WG1 aims at establishing a recommended protocol for measuring the stability of a light source during both short- and long-term image acquisition sessions using calibrated external power sensors. This initial aim will be extended later to measure the absolute flux of light through the illumination path and irradiance of the sample. The initial protocol will be designed around lasers on confocal microscopy platforms (raster scanning and spinning disks). It will be later modified towards other microscopy techniques (widefield, TIRF, light-sheet, super-resolution).

WG 1 – Co-Chairs

WG 1 - Latest News

We welcome our first member from the United Arab Emirates. Rachid Rezgui of NYU Abu Dhabi joined the Working Groups 1-7, 9, 10 and 13. Change the Quality of Light Microscopy Imaging as member of QUAREP-LiMi join us here.

The WG 1 released its first protocol “Illumination Power and Illumination Stability“. The protocol is featured on

Beside step-by-step instructions, the protocol provides:

We welcome our first member from the Ecuador. William Narvaez joined the Working Groups 1, 3, 5, 9 and 11. Change the Quality of Light Microscopy Imaging as member of QUAREP-LiMi join us here.

 

Last meeting: 7th May 2025, 2:15 pm

Release of protocol “Illumination power and illumination stability measurements for contocal, widefield and multiphoton microscopes“. 

Beside step-by-step instructions, the protocol provides:

  1. Automation
    • Aims: The QUAREP-LiMi power and stability assessment protocol requires long and repetitive measurements for each light source as other parameters are tested. The aim of this subgroup is to provide feasible means for carrying out these processes, taking advantage of the automation capabilities of modern microscopes.
    • Co-leads: Mišo Mitkovski, Nasser Darwish
    • On-going tasks:
    • Cloud storage location (can be accessed as a member) here
  2. Environment
  3. Glossary
  4. Power Meter

31st meeting, 5th of March 2024

Meeting date

05/03/2024 @ 5:00 PM (CET/CEST)

Meeting summary

We summarized the By-Laws’ presentation since everyone had seen it at least once. The by-laws and its accompanying documents (Code of Conduct – CoC, Terms of Reference – ToR, and Best Practices – BP) aim to ensure QUAREP-LiMi provides a diverse, equitable, inclusive and transparent environment for its community to gather, address and efficiently make decision to critical challenges. You can find previous recordings of the By-Laws presentation from WG2 here, as well as the slides here. In the folder named Decision Making Discussion https://nextcloud.quarep.org/index.php/f/445127 you will find the other relevant documents related to the by-laws: •BP_BestPractices for QUAREP-LiMi Operations.docx •CoC_WorkGroups.docx •Bylaws Draft_04_a.docx Here are our action items: 1.      WG1 needs to appoint one representative for the SC, ERB, and OB (two-year terms). 2.      WG1 needs to elect its co-chairs for the next two-year term. Nathalie and Laurent are happy to continue on for another two year. We would love to have a third co-chair. 3.      We need candidates! – Let us know if you are interested in being our WG1 representative for SC (ideally one of the co-chairs), ERB, or OB, or if you want to become a WG1 co-chair before March 12th. We also briefly brainstormed on known ways to measure irradiance and decided that a path forward would be to review a proposed definition (by WG13) and present a manuscript (journal club style) at our next meeting. WG1 will meet again on the 3rd of April and will then continue expanding the Illumination power stability protocol to multi-photons.

Meeting documentation

30th meeting, 7th of February 2024

Meeting date

07/02/2024 @ 5:00 PM (CET/CEST)

Meeting summary

First, our collaboration with WG-13 has been defined. Philippe Laissue (co-chair of WG13) talked about the mission of WG13 and how a protocol on how to measure irradiance could fit within the guidance on phototoxicity. Our first goal will be to define irradiance and write a small paragraph to summarize existing methods and guidelines on irradiance. Please share all references you have in this folder or directly in Zotero. We will define irradiance and discuss papers found at our next WG1 meeting (30 min each month). We then briefly discussed Robert Lees very relevant preprint on bioRxiv. We also made headway with version 3 of our protocol and included multi-photons specific comments and recommendations to steps 4 to 6 when needed. We will continue with step 7 at our next meeting. Please go to the protocol online here and add your comments and suggestions to the first 6 steps of the protocol. We will discuss all new comments at our next meeting.

Meeting documentation

29th meeting, 6th of December 2023

Meeting date

06/12/2023 @ 5:00 PM (CET/CEST)

Meeting summary

During this meeting, we branched version 2 of our protocole to extrend it to Multi-photon systems. We discussed and modified staps 1 to 3 of the protocol.

Meeting documentation

28th meeting, 1st of November 2023

Meeting date

01/11/2023 @ 5:00 PM (CET/CEST)

Meeting summary

Lin Yangchen presented his protocol and tools for “Irradiance Measurement on a spinning-disk confocal and TIRF microscope”. We decided to extend next time our protocol to 2P microscopy modality, and we collected the names of people in WG1 who want to collaborate with WG13 to create protocols on irradiance measurement and assessment of phytotoxicity. If you want also to join this subgroup, please contact Natalie Gaudreault.

Meeting documentation

27th meeting, 4th of October 2023

Meeting date

04/10/2023 @ 5:00 PM (CET/CEST)

Meeting summary

Andrii Repula, Product Support Engineer at Lumencor, presented his white paper and how they measure light source stability at his company.

Meeting documentation

26th meeting, 1st of March 2023

Meeting date

01/03/2023 @ 5:00 PM (CET/CEST)

Meeting summary

The subgroup for automation has developed a new script for Zeiss instrument that makes the old one obsolete. It will be uploaded on the website soon. Other participants also inform that more scripts for Leica and Visitron are being developed. We also all agreed on the fact that the GitHub is a living repository and will evolve with time, even when the protocols are done. We then reviewed the latest comments on the protocol and made small improvements to it. We decide to keep the stability formula as it is, even though it might be very sensitive to one outlier and does not reflect always what is wrong. However, we agree that to know there is an outlier or something is wrong is already very important, and one has to plot the stability curve anyhow, and can thereby immediate see why the stability value might be low. Nathalie explains the process for co-authorship and endorsement. We will ask WG chairs to read and say is they want to co-sign or endorse “works for me” through protocol.io. QUAREP general assembly: Laurent will prepare a presentation. Survey: we will align with Roland.

Meeting documentation

25th meeting, 2nd of February 2023

Meeting date

02/02/2023 @ 5:00 PM (CET/CEST)

Meeting summary

Summary of last meeting by Nathalie.

Feed-back from subgroups:
  • Glossary subgroup only worked on camera part, not the illumination part.
  • CAD files for power meter sensor holders have been loaded on the GitHub.
Recently, a new holder for Thorlabs slides in a shape that fits into a multi-well plates holders has been made in Freiburg. Works only for inverted microscopes. Automation: Arne Fallish is working on a program analysing the data recorded with Thorlabs power meter, eventually feed data back to Zeiss Software. We should try to involve big companies more in the automation and encourage them to develop macros. Nathalie shows the new layout of the protocol, made by her and Laurent, to remove step cases and use collapsible notes: Britta discussed the issue with people from Protocol.io and they may investigate it, but right now step-cases are branching out and one cannot come back into the main common/root protocol. Roland indicates that the meeting with Emma has been recorded. One should be able to get in from our QUAREP Cloud. Britta suggests that appendices should be moved in an ad hoc section, if Protocol.io creates one, or we should have a dedicated box in the other sections, since the appendices are not steps and do not belong to the protocol. Another option is to create a pdf and add it to the description section. We will aslo ask protocols.io for an abbreviation box under the description tab. We will ask all WG1 participants, e.g., co-authors of version 1, to read the new protocol before we publish. If we do not here from them, it means that we can keep their names. Britta suggests asking feed-back from all QUAREP members to make sure we do not overlap and contradict part of their work. We agreed. Nathalie will investigate if in protocol.io we could have the endorsement separate from authorship. We screened the videos made by Laurent on power meter usage. We will add the logo of QUAREP on all slides. Also add the corresponding steps from the protocol. We will also consider a submission to PLOS ONE protocols. We finally made improvements to the protocol, based on WG1 members comments.

Meeting documentation

24th meeting, 7th of December 2022

Meeting date

07/12/2022 @ 5:00 PM (CET/CEST)

Meeting documentation

23rd meeting, 2nd of November 2022

Meeting date

02/11/2022 @ 5:00 PM (CET/CEST)

Meeting summary

We recapitulated the last changes in steps 1 to 4 of the protocol, and reviewed the new Question 19 about broad-band illumination sources in Appendix 1. We then further adapted steps 5 to 8 to cover new illumination/microscopy modalities.

Meeting documentation

Subgroup Automation, 4th meeting, 9th of September 2022

Meeting date

09/09/2022 @ 5:00 PM (CET/CEST)

Meeting documentation

Memberlist WG 1