Hello,
I want to write a macro allowing me to characterize inclusions in the cell nucleus (number / nucleus, size and intensity of fluorescence). I think at first to segment the nuclei and then look inside this mask for the inclusions of interest. My problem is that I do not know how to make a new segmentation in the segmentation of the nuclei. Thank you in advance for your advice and ideas so that I can write this macro. I attach an image of cores (DAPI) and an image with the marker of interest (TxRed). A big thank you to all, Patricia _______________________________________ [http://www.cea.fr/var/cea/signatures/cea_logo.jpg]<http://www.cea.fr/> Patricia OBEÏD Ingénieur de Recherche /Animateur Informatique Biomics Institut de Recherche Interdisciplinaire de Grenoble DRF / IRIG / DS /BGE U1038/ Biomics / Bât 4020 Commissariat à l'énergie atomique et aux énergies alternatives Email : [hidden email]<mailto:[hidden email]> Tél : (+33)4 38 78 47 12 Fax : (+33)4 38 78 59 17 Toute notre actualité sur www.cea.fr et sur http://www.cea.fr/drf/big/bge/biomics<http://www.cea.fr> <http://www.cea.fr>Suivez-nous également sur Twitter : @CEA_Recherche<https://twitter.com/#!/CEA_Recherche> et sur https://twitter.com/BiomicsLab -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html image001.jpg (11K) Download Attachment s1_TxRed.tif (700K) Download Attachment s1_DAPI.tif (700K) Download Attachment |
This kind of problem can be resolved using RCC
https://blog.bham.ac.uk/intellimic/spatial-reasoning-with-imagej-using-the-region-connection-calculus/ Once you segment your objects as binary, you could look for the relation of regions in one image with the regions in the other image. See "Computing relations between multiple regions" section in that page. RCC5 should be enough to detect the PP relations between the intranuclear regions and the nuclei. Once you have the RCC table generated, then it is a matter of searching the table across the "nuclear regions axis" and count how many PP relations they hold with the "intranuclear regions" axis. There are several papers with examples here: https://blog.bham.ac.uk/intellimic/references/ Hope it helps. Gabriel On Thursday, 12 December 2019 05:31:49 GMT [hidden email] wrote: > Hello, > I want to write a macro allowing me to characterize inclusions in the cell > nucleus (number / nucleus, size and intensity of fluorescence). I think at > first to segment the nuclei and then look inside this mask for the > inclusions of interest. My problem is that I do not know how to make a new > segmentation in the segmentation of the nuclei. Thank you in advance for > your advice and ideas so that I can write this macro. I attach an image of > cores (DAPI) and an image with the marker of interest (TxRed). A big thank > you to all, > Patricia > > _______________________________________ > [http://www.cea.fr/var/cea/signatures/cea_logo.jpg]<http://www.cea.fr/> > > Patricia OBEÏD > Ingénieur de Recherche /Animateur Informatique Biomics > Institut de Recherche Interdisciplinaire de Grenoble > DRF / IRIG / DS /BGE U1038/ Biomics / Bât 4020 > Commissariat à l'énergie atomique et aux énergies alternatives > Email : [hidden email]<mailto:[hidden email]> > Tél : (+33)4 38 78 47 12 Fax : (+33)4 38 78 59 17 > > Toute notre actualité sur www.cea.fr et sur > http://www.cea.fr/drf/big/bge/biomics<http://www.cea.fr> > > <http://www.cea.fr>Suivez-nous également sur Twitter : > @CEA_Recherche<https://twitter.com/#!/CEA_Recherche> et sur > https://twitter.com/BiomicsLab > > > > > -- > ImageJ mailing list: http://imagej.nih.gov/ij/list.html -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
In reply to this post by OBEID Patricia 154904
That may be overkill if I understand the original question. Do you just need to count the objects inside the ROIs? If that is the case, then you can cycle through each ROI in the ROI manager and find local maxima in each ROI on the channel of interest. The manual code below will let you do it. Obviously, you can automate the whole thing. If you need/want other data for the spots, then obviously this won't work, and I would consider Gabriel's option, or you could try cycling through each ROI and duplicating that ROI as a new image to allow analysis of the spots in that- I have considered doing this before- you need to delete data outside the ROI after duplicating, since it duplicates a rectangle around the ROI. I abandoned this since I was more interested in counts, and it was getting a bit complicated for my abilities!
------ //collect ROIS (e.g. nuclei) from one image,then swap to object of interest (e.g. foci) channel and count the spots. //Assumes you have both images open (the one for finding the ROIs, eg nuclear counterstain and the one for the spots inside it. var tolerance = 30; originalImage = getTitle(); run("8-bit"); run("Overlay Options...", "stroke=red width=1 fill=none set"); run("Median...", "radius=6"); run("Subtract Background...", "rolling=125 sliding"); run("Threshold", "method=Default white"); roiManager("Reset"); run("Analyze Particles...", "pixel exclude clear add"); waitForUser("Select the image for spot counting"); run("8-bit"); run("Gaussian Blur...", "sigma=1"); run("Subtract Background...", "rolling=25"); for(i=0; i<roiManager("count"); i++) { roiManager("select", i); run("Find Maxima...", "noise="+tolerance+" output=[Count]"); run("Find Maxima...", "noise="+tolerance+" output=[Point Selection]"); run("Add Selection..."); } ------ Glyn -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
Hi Glyn,
I'm actually interested in the intensity and size of each object contained in the nucleus. So I'm trying to adapt Gabriel's solution but I admit that I'm getting lost ... Thank you very much for your feedback. Regards, Patricia -----Message d'origine----- De : ImageJ Interest Group <[hidden email]> De la part de Glyn Nelson Envoyé : vendredi 13 décembre 2019 12:52 À : [hidden email] Objet : Re: Help to create macro That may be overkill if I understand the original question. Do you just need to count the objects inside the ROIs? If that is the case, then you can cycle through each ROI in the ROI manager and find local maxima in each ROI on the channel of interest. The manual code below will let you do it. Obviously, you can automate the whole thing. If you need/want other data for the spots, then obviously this won't work, and I would consider Gabriel's option, or you could try cycling through each ROI and duplicating that ROI as a new image to allow analysis of the spots in that- I have considered doing this before- you need to delete data outside the ROI after duplicating, since it duplicates a rectangle around the ROI. I abandoned this since I was more interested in counts, and it was getting a bit complicated for my abilities! ------ //collect ROIS (e.g. nuclei) from one image,then swap to object of interest (e.g. foci) channel and count the spots. //Assumes you have both images open (the one for finding the ROIs, eg nuclear counterstain and the one for the spots inside it. var tolerance = 30; originalImage = getTitle(); run("8-bit"); run("Overlay Options...", "stroke=red width=1 fill=none set"); run("Median...", "radius=6"); run("Subtract Background...", "rolling=125 sliding"); run("Threshold", "method=Default white"); roiManager("Reset"); run("Analyze Particles...", "pixel exclude clear add"); waitForUser("Select the image for spot counting"); run("8-bit"); run("Gaussian Blur...", "sigma=1"); run("Subtract Background...", "rolling=25"); for(i=0; i<roiManager("count"); i++) { roiManager("select", i); run("Find Maxima...", "noise="+tolerance+" output=[Count]"); run("Find Maxima...", "noise="+tolerance+" output=[Point Selection]"); run("Add Selection..."); } ------ Glyn -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
On Tuesday, 17 December 2019 13:44:41 GMT [hidden email] wrote:
> Hi Glyn, > I'm actually interested in the intensity and size of each object contained > in the nucleus. So I'm trying to adapt Gabriel's solution but I admit that > I'm getting lost ... Thank you very much for your feedback. > Regards, > Patricia If you get stuck with the RCC analysis, please binarise (and clean) the images of the nuclei and the bright dots in the nuclei and post them here. I can then attempt to show you how I think this can be resolved using macros calling the RCC plugin(s). Cheers Gabriel -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
Good evening Gabriel,
as proposed, here are the binarized and cleaned images (as best as I could). Thank you very much for your help. Cordially Patricia -----Message d'origine----- De : ImageJ Interest Group <[hidden email]> De la part de Gabriel Landini Envoyé : mardi 17 décembre 2019 15:38 À : [hidden email] Objet : Re: Help to create macro On Tuesday, 17 December 2019 13:44:41 GMT [hidden email] wrote: > Hi Glyn, > I'm actually interested in the intensity and size of each object > contained in the nucleus. So I'm trying to adapt Gabriel's solution > but I admit that I'm getting lost ... Thank you very much for your feedback. > Regards, > Patricia If you get stuck with the RCC analysis, please binarise (and clean) the images of the nuclei and the bright dots in the nuclei and post them here. I can then attempt to show you how I think this can be resolved using macros calling the RCC plugin(s). Cheers Gabriel -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
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