NetNotes
Beanshell, Python or Java. Saving recorded or written macros with the analyzed data allows checking/showing later how the data were analyzed. Kees Straatman
krs5@leicester.ac.uk
Following up on this, I strongly endorse using a macro script
to standardize image processing for all images in an acquisition. Doing this has benefits that wildly exceed the time spent on learning the scripting process: 1) You save an incredible amount of time over modifying images manually. Te larger the image set the more time you save; 2) You can re-use a macro script or quickly modify it to meet your new needs, so you only pay the time cost of developing it once; 3) It ensures identical processing of all images in the data set. If one of the steps can’t be standardized (thresholding, for example), then you can put in a variable step that asks the user to set the value for each image; 4) It provides auditability, which IMO is extremely important in image analysis. If a problem comes up, you can skim through the protocol and identify the problem; 5) Once a problem is found, the script can be fixed and the whole data set re-processed with almost no time cost; 6) Te script can be placed in the supplemental data or sent to colleagues to ensure reproducibility; 7) Whatever the project, odds are good that a relevant script that’s already written can be found and adapted. Timothy Feinstein
tnf8@pitt.edu
I have a few things to add from the perspective of an image
processing developer who has worked on reproducible image processing workflows and algorithms. A couple of years ago I was part of a large group involved in a major reproducibility study: https://
pubmed.ncbi.nlm.nih.gov/31845647/. 1) In addition to MATLAB, Python, and ImageJ, I also
recommend a tool called KNIME (
https://www.knime.com/). It has a bit of a learning curve, but less so than MATLAB or Python. It is a GUI-based visual programming tool for data and image analysis. We used it for our reproducibility study and found it a nice way for developers and non-developers to collaborate on workflows; 2)
https://forum.image.sc/ The scientific community image forum (image sc forum) has become the go-to place for image processing discussions. Anyone doing image processing should take advantage of this resource; 3) MATLAB vs Python: I’ve used both and like both, and have not found a huge difference in learning curves. Python has a huge number of well-tested extensions, more than MATLAB in my experience, though I am not aware of an official count. Some Python tool kits like scikit-image, Napari (visualization), and the deep learning eco-system are extremely well supported on
https://forum.image.sc. For example, just today this thread started:
https://forum.image.sc/t/looking-for-life- scientists-to-collaborate-on-scikit-image-tutorials/49073; 4) For reproducible work, algorithms should be described with the same names across platforms (that is, Otsu Thresholding, Richardson Lucy Deconvolution, etc.). In our reproducibility study, it was hard for us to figure out the previous protocol as non-standard algorithm names were used in the description we received; 5) In my experience, if there is strong evidence in an image, you can often process the image and get results relatively easily and get the same result from multiple approaches. Tweaking super complicated image processing protocols sometimes just overfits the data; 6) Validation: I’ve heard a lot of talk over the years about tools being “validated” or “quantitative.” “Validation” of an algorithm isn’t trivial and “quantitative” is a vague term. The best “validation” test is one where an independent group publicly releases data and gives developers of different platforms a chance to run the test and show it meets a standard. Brian Northan
bnorthan@gmail.com
84
One other quick consideration for Python vs. MATLAB. If the
pipeline is in MATLAB, people will have to pay to verify and use your process. If it is in Python they can verify and use it for free. Having licensing fees as a barrier of entry to doing science feels less than ideal, especially if you feel science should be equally accessible to everyone. Ben Smith
benjamin.smith@
berkeley.edu
Tanks everyone. I am not trying to come up with my own rules
or even hunt for manipulations. I have a clear case of manipulation by someone who had no bad intention, but only a lack of knowledge. I remembered hearing about InspectJ but could not find the name. I would like to test it. As usual, thanks everyone for the quick help! Sylvie Le Guyader
sylvie.le.guyader@
ki.se
Zeiss Oil on a Leica Microscope Objective Confocal Listserver I was told recently by a Leica engineer that he encountered many
Leica objectives destroyed by Zeiss oil. I was not able to figure out from him if he was pulling my leg (which I think is the case) or was talking seriously. Tanks. Petro Khoroshyy
khoroshyy@gmail.com
We have been doing this for more than six years without any problems
on confocal and widefield systems. Eva Wegel
eva.wegel@jic.ac.uk If you read the Leica Immersion Fluid Safety Data Sheet the producer
is Carl Zeiss Jena Gmbh! I can send you the safety data sheet if you want to show it to the Leica engineer. Erwan Grandgirard
grandgie@igbmc.fr
We did this in both directions (using oil from Leica on a Zeiss
microscope, and vice versa). If the refractive index of the oil corresponds there seems, in our experience, no reason not to do this. I think it is more critical to not use oil for extended periods aſter opening, as it could degrade through oxidation. As a precaution, we try to avoid using an opened bottle for longer than six months. Christoph Ruediger Bauer
christoph.bauer@
unige.ch
If you check the Material Safety Data Sheet of the Leica oil you
will find that the producer is Zeiss. We are refilling the nice 10ml Leica bottles with plastic rods from a 500 ml Zeiss 518 F for years now with no problem at all. I do not think that this is the same for other oils though. We had some substantial chromatic aberrations in the deep red with Cargille HF. So, for a Leica microscope, I would buy either from Leica or Zeiss. Steffen Dietzel
lists@dietzellab.de
My microscope rep worked for Nikon and Zeiss with an independent
company, is very trustworthy, and he claimed the same thing. I have never seen it either, but I suspect that he really did have something happen. He never told me any sales nonsense at any other time. In his bad experience, the issue was mixing oils from two companies by not cleaning an objective before putting a different oil on it. I agree it seems unlikely there is a problem, but for safety, given the cost of objectives, I would just clean the objectives before switching oils and you will not have a problem. Dave Knecht
david.knecht@
uconn.edu
We have used Zeiss oil on Leica microscopes for 10 years without
objective damage and without the Leica service engineer complaining about it. Antonio Virgilio Failla
a.failla@
uke.de
While I think it is the same situation, can anyone comment
on the use of Immersol W with non-Zeiss objectives. Petro Khoroshyy
khoroshyy@gmail.com
www.microscopy-today.com • 2021 May
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