journal article·2023-04-05Open access

Tissue-Like 3D Standard and Protocols for Microscope Quality Management

Benjamin Abrams, Thomas Pengo, Tse-Luen Wee, Rebecca C Deagle, Nelly Vuillemin, Linda M Callahan, Megan A Smith, Kristopher E Kubow, Anne-Marie Girard, Joshua Z Rappoport, Carol J Bayles, Lisa A Cameron, Richard Cole, Claire M Brown

Microscopy and Microanalysis· Volume 29· Issue 2

Abstract

Abstract This article outlines a global study conducted by the Association of Biomedical Resource Facilities (ABRF) Light Microscopy Research Group (LMRG). The results present a novel 3D tissue-like biologically relevant standard sample that is affordable and straightforward to prepare. Detailed sample preparation, instrument-specific image acquisition protocols and image analysis methods are presented and made available to the community. The standard consists of sub-resolution and large well characterized relative intensity fluorescence microspheres embedded in a 120 µm thick 3D gel with a refractive index of 1.365. The standard allows the evaluation of several properties as a function of depth. These include the following: 1) microscope resolution with automated analysis of the point-spread function (PSF), 2) automated signal-to-noise ratio analysis, 3) calibration and correction of fluorescence intensity loss, and 4) quantitative relative intensity. Results demonstrate expected refractive index mismatch dependent losses in intensity and resolution with depth, but the relative intensities of different objects at similar depths are maintained. This is a robust standard showing reproducible results across laboratories, microscope manufacturers and objective lens types (e.g., magnification, immersion medium). Thus, these tools will be valuable for the global community to benchmark fluorescence microscopes and will contribute to improved scientific rigor and reproducibility.
Source: crossref · Quality: baseline

Research context

OpenAlex topics: Advanced Fluorescence Microscopy Techniques · Cell Image Analysis Techniques · Optical Imaging and Spectroscopy Techniques

SciVX discovery tags are terminology-based. OpenAlex topics are source-provided classifications.

Affiliations

Life Sciences Microscopy Center, University of California, Santa Cruz , 1156 High Street, 150 Sinsheimer Labs, Santa Cruz, CA 95064 , USA

Informatics Institute, University of Minnesota Twin Cities, Cancer and Cardiovascular Research Building , 2231 6th St SE, Minneapolis, MN 55449 , USA

Advanced BioImaging Facility (ABIF), McGill University , 3649 Prom, Sir William Osler, Bellini Building, Room 137, Montreal, QC H3G 0B1 , Canada

Department of Physiology, McGill University , 3655 Promenade Sir-William-Osler, Montreal, Quebec H3G 1Y6 , Canada

Department of Neuroscience, Del Monte Institute for Neuroscience, Univ. Rochester Medical Center , Rochester, NY 14642 , USA

Biology Department, James Madison University , Bioscience Building, 951 Carrier Drive, Harrisonburg, VA 22807 , USA

Center for Genome Research and Biocomputing, Oregon State University , 1500 SW Jefferson Way, Corvallis, OR 97331 , USA

Center for Advanced Microscopy and Nikon Imaging Center, Feinberg School of Medicine, Northwestern Medicine, Northwestern University , Chicago, IL 60611 , USA

Institute of Biotechnology, Cornell University , Cornell Institute of Biotechnology, Microscopy Facility, 130 Biotechnology Building, Cornell University Ithaca, NY 14853 , USA

Light Microscopy Core Facility, Duke University , 4215 French Family Science Center, 124 Science Drive, Durham, NC 27708 , USA

New York State Dept of Health/Wadsworth Center, Advanced Light Microscopy & Image Analysis Core Facility , 150 New Scotland Ave, Albany, NY 12208 , USA

References (45)

Show source-provided references
1. Methods to calibrate and scale axial distances in confocal microscopy as a function of refractive index10.1111/jmi.12194 ↗
2. QUAREP-LiMi: A community endeavor to advance quality assessment and reproducibility in light microscopy10.1038/s41592-021-01162-y ↗
3. A quantitative measure of field illumination10.7171/jbt.15-2602-001 ↗
4. Measuring and interpreting point spread functions to determine confocal microscope resolution and ensure quality control10.1038/nprot.2011.407 ↗
5. International test results for objective lens quality, resolution, spectral accuracy and spectral separation for confocal laser scanning microscopes10.1017/S1431927613013470 ↗
6. Policy: NIH plans to enhance reproducibility10.1038/505612a ↗
7. Microscope calibration using laser written fluorescence10.1364/OE.26.021887 ↗
8. Reproducibility in light microscopy: Maintenance, standards and SOPs10.1016/j.biocel.2017.06.008 ↗
9. Intensitycheck—The light measuring app for microscope performance checks and consistent fluorescence imaging10.1371/journal.pone.0214659 ↗
10. Using the NoiSee workflow to measure signal-to-noise ratios of confocal microscopes10.1038/s41598-018-37781-3 ↗
11. Reproducibility crisis in science or unrealistic expectations?10.15252/embr.201846008 ↗
12. Goodwin · 2007 · 39710.1016/S0091-679X(06)81018-6 ↗
13. Calibrating excitation light fluxes for quantitative light microscopy in cell biology10.1038/nprot.2008.180 ↗
14. An automated protocol for performance benchmarking a widefield fluorescence microscope10.1002/cyto.a.22519 ↗
15. Towards community-driven metadata standards for light microscopy: Tiered specifications extending the OME model10.1038/s41592-021-01327-9 ↗
16. Aberrations in confocal fluorescence microscopy induced by mismatches in refractive-Index10.1111/j.1365-2818.1993.tb03315.x ↗
17. Confocalcheck–a software tool for the automated monitoring of confocal microscope performance10.1371/journal.pone.0079879 ↗
18. Quantitative confocal microscopy: Beyond a pretty picture10.1016/B978-0-12-420138-5.00007-0 ↗
19. Tutorial: Guidance for quantitative confocal microscopy10.1038/s41596-020-0313-9 ↗
20. Method of calibration of a fluorescence microscope for quantitative studies10.1111/j.1365-2818.2011.03514.x ↗
21. Survey on scientific shared resource rigor and reproducibility10.7171/jbt.19-3003-001 ↗
22. What's in a sample? Increasing transparency in biospecimen procurement methods10.1038/nmeth.4684 ↗
23. Assessing camera performance for quantitative microscopy10.1016/B978-0-12-420138-5.00003-3 ↗
24. A beginner's Guide to rigor and reproducibility in fluorescence imaging experiments10.1091/mbc.E17-05-0276 ↗
25. Calibration of wide-field deconvolution microscopy for quantitative fluorescence imaging10.7171/jbt.14-2501-002 ↗
26. Imaging methods are vastly underreported in biomedical research10.7554/eLife.55133 ↗
27. A review of the scientific rigor, reproducibility, and transparency studies conducted by the ABRF research groups10.7171/jbt.20-3101-003 ↗
28. Best practices and tools for reporting reproducible fluorescence microscopy methods10.1038/s41592-021-01156-w ↗
29. Fluorescence microscope light source stability10.1007/s00418-019-01776-6 ↗
30. Practical aspects of quantitative confocal microscopy10.1016/B978-0-12-407761-4.00018-X ↗
31. Evaluating performance in three-dimensional fluorescence microscopy10.1111/j.1365-2818.2007.01861.x ↗
32. National Academies of Sciences, E., and Medicine · 2019
33. QUAREP-LiMi: A community-driven initiative to establish guidelines for quality assessment and reproducibility for instruments and images in light microscopy10.1111/jmi.13041 ↗
34. When light meets biology—How the specimen affects quantitative microscopy10.1242/jcs.259656 ↗
35. Effects of specimen refractive index on confocal imaging10.1046/j.1365-2818.1997.d01-627.x ↗
36. Quality assurance testing for modern optical imaging systems10.1017/S1431927611000237 ↗
37. A global view of standards for open image data formats and repositories10.1038/s41592-021-01113-7 ↗
38. PSFj: Know your fluorescence microscope10.1038/nmeth.3102 ↗
39. Volume measurements in three-dimensional microscopy10.1002/sca.4950160403 ↗
40. Accuracy and precision in quantitative fluorescence microscopy10.1083/jcb.200903097 ↗
41. Concepts in quantitative fluorescence microscopy10.1016/B978-0-12-420138-5.00001-X ↗
42. Wilhelm · 1997 · 1
43. Evaluation of confocal microscopy system performance10.1007/978-1-59259-993-6_5 ↗
44. Quality assessment of confocal microscopy slide based systems: Performance10.1002/cyto.a.20314 ↗
45. Evaluation of confocal microscopy system performance10.1002/1097-0320(20010801)44:4<273::AID-CYTO1120>3.0.CO;2-N ↗