6 must-read articles about Quality Control in fluorescence microscopy

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1. “Accuracy and precision in quantitative fluorescence microscopy”

Jennifer C. Waters – 2009 1

 

Read it here: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2712964/

A thorough work has been done to pinpoint the parameters that affect precision and accuracy, focusing on three main pains: signal, background, and noise. The article includes neat checklists (for optimizing images for quantitation) and protocols (for quantitation of fluorescence intensity values).

 

2. “Need for Standardization of Fluorescence Measurements from the Instrument Manufacturer’s View”

Andrew Dixon, Thomas Heinlein, Ralf Wolleschensky – 2008 2

 

Read it here: https://link.springer.com/chapter/10.1007/4243_2008_026

One of the authors works in R&D at Zeiss microscopy; this adds to the article an expertise and knowledge of the manufacturer’s constraints. Confrontation between manufacturer and users may arise when trying to find out if an issue comes from the system’s performance or from poor sample preparation/changes in the user’s imaging protocol. Thus, the need for an arbitrated tool for performance measurement.

 

3. “How to Improve Quality Assurance in Fluorometry: Fluorescence-Inherent Sources of Error and Suited Fluorescence Standards”

U. Resch-Genger, K. Hoffmann, et al.- 2005 3

 

Read it here: https://link.springer.com/article/10.1007/s10895-005-2630-3

The article discusses the variables that influence the information content of an image, and then goes into more detail about signal detection sensitivity and standardized tests. It depicts the use of a thin fluorescent film sample to determine the Signal-to-Noise Ratio in a confocal microscope.

   “Once the system is installed and commissioned, the user needs to maintain confidence that the instrument continues to deliver consistent performance.”   

4. “Quality Assessment of Confocal Microscopy Slide Based Systems: Performance”

 Robert M. Zucker – 2005 4

Read it here: https://onlinelibrary.wiley.com/doi/full/10.1002/cyto.a.20314

 In this article, a series of quality assurance tests have been performed on a confocal microscope to determine if the equipment is operating correctly, using Field Illumination-Fluorescent Slides, power meter, beads and biological test slides.

5. “Tutorial: guidance for quantitative confocal microscopy”

James Jonkman, Claire M. Brown, et al. – 2020 5

Read it here: https://www.nature.com/articles/s41596-020-0307-7.pdf

And the related visual summary : https://www.nature.com/articles/s41596-020-0307-7.pdf

This tutorial addresses several aspects of acquisition and use of quantitative confocal microscopy images, from sample preparation to analysis and presentation of the collected data.

   “Generating meaningful data using confocal microscopy requires careful planning and a thorough understanding of the technique.”  

6. “Editorial: Better research through metrology”

Editor of Nature Methods – 2018 6

Read it here:

https://doi.org/10.1038/s41592-018-0035-x

A direct approach to the a reality of metrology, that also mentions the historic origin of standards.

     “As has often been pointed out, if one does an experiment twice and gets a different result each time, the likelihood is strong […] that one has inadvertently done a different experiment.”    

There would be of course many more article to cite. Each year a new article, a new talk, conference session discusses the topic of Quality Control. Keep up the good work !  

Sources:

(1) “Accuracy and precision in quantitative fluorescence microscopy”. J. C. Waters, The Journal of cell biology ,185(7), 1135–1148. https://doi.org/10.1083/jcb.200903097

(2) ““Need for Standardization of Fluorescence Measurements from the Instrument Manufacturer’s View”. Andrew Dixon, Thomas Heinlein, Ralf Wolleschensky, from Standardization and Quality Assurance in Fluorescence Measurements II. Springer Series on Fluorescence, vol 6., (2008), https://doi.org/10.1007/4243_2008_026

(3) “How to Improve Quality Assurance in Fluorometry: Fluorescence-Inherent Sources of Error and Suited Fluorescence Standards”. U. Resch-Genger, K. Hoffmann, et al, Journal of Fluorescence, Vol. 15, No. 3, (2005) https://doi.org/10.1007/s10895-005-2630-3

(4) “Quality assessment of confocal microscopy slide based systems: Performance”. R. M. Zucker, Cytometry Part A 69A:659–676 (2006). https://doi.org/10.1002/cyto.a.20314

(5) “Tutorial: guidance for quantitative confocal microscopy”. J. Jonkman, C.M. Brown, G.D Wright. et al., Nature Protocol (2020). https://doi.org/10.1038/s41596-020-0313-9

(6) “Better research through metrology”. Nat Methods 15, 395 (2018). https://doi.org/10.1038/s41592-018-0035-x

Header photo by cheaousa on Unsplash
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