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John Oreopoulos on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Advice-on-citations-for-homogenized-fluorescence-illumination-tp6854775p6855162.html
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Partial commercial response:
Hi Ricardo,
There are a number of fairly recent research articles which come to mind. I have collected a number of references below which you may find helpful.
In a related commercial response, I would like to add that the company that I work for, Spectral Applied research, developed a laser-based illumination technology named Borealis (patent-pending) for spinning disk confocal microscopes a few years ago. Although not the original intent of the technology, one of the benefits of Borealis is that it provides, to the best of our knowledge, the flattest and most homogenous illumination profile for any laser-based microscope illumination system. Since I joined the company a year ago, we've spent a great deal of time trying to quantitatively prove this in the R&D lab here at Spectral. Mike Model's concentrated dye solutions were particularly helpful in this regard (see references below). We measure illumination uniformity as the percent rolloff from the peak intensity in the middle to the lowest intensity at the ends of a line profile drawn across an image diagonal (as recommended in Pawley's confocal handbook). Spinning disk confocal systems out of the box are notoriously bad for uniformity and in some cases we measured a %rolloff as high as 60% in the red portions of the visible spectrum. With Borealis, we can bring this down 5% or less. When other optical system non-uniformities are included (camera lens field curvature and vignetting for example), this value is still on the order of only 8%. We are currently compiling all of this data into a publishable form. In addition, Borealis can be easily adapted for regular wide-field epfluorescence microscopes as well. We'll be demoing the wide-field version of Borealis at the upcoming Neuroscience and Cell Biology annual meetings.
The references I found useful over the years regarding microscope illumination uniformity are:
1. Waters, J.C., Accuracy and precision in quantitative fluorescence microscopy. Journal of Cell Biology, 2009. 185(7): p. 1135-1148.
2. Murray, J.M., et al., Evaluating performance in three-dimensional fluorescence microscopy. Journal of Microscopy-Oxford, 2007. 228(3): p. 390-405.
3. Wolf, D.E., C. Samarasekera, and J.R. Swedlow, Quantitative analysis of digital microscope images, in Digital Microscopy, 3rd Edition. 2007, Elsevier Academic Press Inc: San Diego. p. 365-396.
Wolf's chapter in Digital Microscopy talks about why there is a need for flat illumination in the context of an quantitative measurements with a fluorescence microscope, especially in cases where comparisons are made across different spectral channels (FRET, calcium imaging, etc.).
Next:
4. Zucker, R.M. and O.T. Price, Practical confocal microscopy and the evaluation of system performance. Methods-a Companion to Methods in Enzymology, 1999. 18(4): p. 447-458.
5. Zucker, R.M. and O. Price, Evaluation of confocal microscopy system performance. Cytometry, 2001. 44(4): p. 273-294.
Zucker talks about some alternative methods to assess microscope illumination Uniformity. You might also want to take a look at these references for some more ideas and methods:
6. Zwier, J.M., et al., Image calibration in fluorescence microscopy. Journal of Microscopy-Oxford, 2004. 216: p. 15-24.
7. Brakenhoff, G.J., et al., Characterization of sectioning fluorescence microscopy with thin uniform fluorescent layers: Sectioned Imaging Property or SIPcharts. Journal of Microscopy-Oxford, 2005. 219: p. 122-132.
8. Zwier, J.M., et al., Quantitative image correction and calibration for confocal fluorescence microscopy using thin reference layers and SIPchart-based calibration procedures. Journal of Microscopy-Oxford, 2008. 231(1): p. 59-69.
9. Model, M.A. and J.K. Burkhardt, A standard for calibration and shading correction of a fluorescence microscope. Cytometry, 2001. 44(4): p. 309-316.
10. Model, M.A., Intensity Calibration and Shading Correction for Fluorescence Microscopes. Current Protocols in Cytometry. 2006: John Wiley & Sons, Inc.
11. Model, M.A. and J.L. Blank, Intensity calibration of a laser scanning confocal microscope based on concentrated dyes. Analytical and Quantitative Cytology and Histology, 2006. 28(5): p. 253-261.
12. Model, M.A. and J.L. Blank, Concentrated dyes as a source of two-dimensional fluorescent field for characterization of a confocal microscope. Journal of Microscopy-Oxford, 2008. 229(1): p.12-16.
Mike Model's "concentrated fluorescent dye" solutions for measuring system uniformity are very easy to make and have some other useful properties, including intensity calibration. Finally, there are two publications from this year, one that actually tried to catalog the non-uniformities across different microscope systems (mainly confocal) during an international standardization study:
13. Stack, R.F., et al., Quality Assurance Testing for Modern Optical Imaging Systems. Microscopy and Microanalysis, 2011. 17(4): p. 598-606.
14. Michálek, J., M. Čapek, and L. Kubínová, Compensation of inhomogeneous fluorescence signal distribution in 2D images acquired by confocal microscopy. Microscopy Research and Technique, 2011. 74(9): p. 831-838.
John Oreopoulos
Research Assistant
Spectral Applied Research
Richmond Hill, Ontario
Canada
www.spectral.ca
On 2011-10-03, at 8:12 AM, Ricardo Henriques wrote:
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> To join, leave or search the confocal microscopy listserv, go to:
>
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> *****
>
> Hello everyone,
>
> I'm preparing a small manuscript that in part covers the concept that most fluorescence microscopes (SD, WF, TIRF, ...) have a non-homogenous illumination of the field of view - typically gaussian-shaped, less intensity on the image extremities. I was wondering if you could advise me on what you consider the most interesting publications tackling or approaching this subject.
>
> Thank you in advanced.
> Best regards,
> Ricardo Henriques
>
> Ricardo Henriques
> Instituto de Medicina Molecular (Lisbon, Portugal).
> For contact information see:
https://sites.google.com/site/paxcalpt/