Quantification of non-colocalized signal
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Dear all,
Can you suggest a way to quantify an image after excluding co-localized signal? I have performed immunofluorescence experiment for two proteins and I would like to quantify everything that is not co-localized. Thank you for all your help in advance, Katerina -- Sent from: http://imagej.1557.x6.nabble.com/ -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
Re: Quantification of non-colocalized signal
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Dear Katerina,
I guess your request is quite weird which explains why nobody hasn't replied to it before. I'm also not very sure whether I understand correctly what you are trying to do. Nevertheless with the Colocalization_Finder plugin that you can download under the following link: http://questpharma.u-strasbg.fr/html/colocalization-finder.html you will get a composite picture composed of the color merged single channel pictures as well as a scatter_plot picture representing the pixels intensities correlation diagram. You can then define a ROI "on the fly" within the scatter_plot which will then colorize the corresponding selected pixels within the composite picture in white. By clicking within the ROI you will calculate the selected pictures correlation coefficients and by a double click within the ROI you will as well calculate the selected pictures correlation coefficients together of a "fixing" (i.e. they will be colorized in blue, green, red,... their color staying within the picture) of this ROI as well as the selected pixels within the composite picture. Thus by doing so you can figure out the pixels which are correlated and find out the one that aren't for further analysis. In parallel to the "on the fly" scatter_plot ROI selection, you can as well define "on the fly" a ROI within the composite picture which will then restrict the analysis only to the pixels within this ROI. The analysis being thus the cross selection of both the scatter_plot and composite pictures ROIs selections. Hoping my description helps you to move further I wish you a nice day. My best regards, Philippe Philippe CARL Laboratoire de Bioimagerie et Pathologies UMR 7021 CNRS - Université de Strasbourg Faculté de Pharmacie 74 route du Rhin 67401 ILLKIRCH Tel : +33(0)3 68 85 42 89 ----- Mail original ----- De: "Katerina" <[hidden email]> À: "imagej" <[hidden email]> Envoyé: Lundi 2 Novembre 2020 17:24:46 Objet: Quantification of non-colocalized signal Dear all, Can you suggest a way to quantify an image after excluding co-localized signal? I have performed immunofluorescence experiment for two proteins and I would like to quantify everything that is not co-localized. Thank you for all your help in advance, Katerina -- Sent from: http://imagej.1557.x6.nabble.com/ -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
Re: Quantification of non-colocalized signal
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The simple method of finding the non colocalized fraction derives immediately from the M1 and M2 coefficients - most colocalization software will report M1 and M2.
Each gives the fraction of one fluorophore found where the other is present i.e. in the area of colocalization. So, if M1 says that 0.22 of the fluorophore is found where the other is found, then 1 - 0.22 (M1) is found in the remaining area. Similarly with the other fluorophore use M2 You also want to consider which part of the image were used (the ROI) for the measurements and how the two image were thresholded. Jeremy Adler Uppsala U -----Original Message----- From: ImageJ Interest Group <[hidden email]> On Behalf Of CARL Philippe (LBP) Sent: Wednesday, November 4, 2020 5:14 PM To: [hidden email] Subject: Re: Quantification of non-colocalized signal Dear Katerina, I guess your request is quite weird which explains why nobody hasn't replied to it before. I'm also not very sure whether I understand correctly what you are trying to do. Nevertheless with the Colocalization_Finder plugin that you can download under the following link: http://questpharma.u-strasbg.fr/html/colocalization-finder.html you will get a composite picture composed of the color merged single channel pictures as well as a scatter_plot picture representing the pixels intensities correlation diagram. You can then define a ROI "on the fly" within the scatter_plot which will then colorize the corresponding selected pixels within the composite picture in white. By clicking within the ROI you will calculate the selected pictures correlation coefficients and by a double click within the ROI you will as well calculate the selected pictures correlation coefficients together of a "fixing" (i.e. they will be colorized in blue, green, red,... their color staying within the picture) of this ROI as well as the selected pixels within the composite picture. Thus by doing so you can figure out the pixels which are correlated and find out the one that aren't for further analysis. In parallel to the "on the fly" scatter_plot ROI selection, you can as well define "on the fly" a ROI within the composite picture which will then restrict the analysis only to the pixels within this ROI. The analysis being thus the cross selection of both the scatter_plot and composite pictures ROIs selections. Hoping my description helps you to move further I wish you a nice day. My best regards, Philippe Philippe CARL Laboratoire de Bioimagerie et Pathologies UMR 7021 CNRS - Université de Strasbourg Faculté de Pharmacie 74 route du Rhin 67401 ILLKIRCH Tel : +33(0)3 68 85 42 89 ----- Mail original ----- De: "Katerina" <[hidden email]> À: "imagej" <[hidden email]> Envoyé: Lundi 2 Novembre 2020 17:24:46 Objet: Quantification of non-colocalized signal Dear all, Can you suggest a way to quantify an image after excluding co-localized signal? I have performed immunofluorescence experiment for two proteins and I would like to quantify everything that is not co-localized. Thank you for all your help in advance, Katerina -- Sent from: http://imagej.1557.x6.nabble.com/ -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html När du har kontakt med oss på Uppsala universitet med e-post så innebär det att vi behandlar dina personuppgifter. För att läsa mer om hur vi gör det kan du läsa här: http://www.uu.se/om-uu/dataskydd-personuppgifter/ E-mailing Uppsala University means that we will process your personal data. For more information on how this is performed, please read here: http://www.uu.se/en/about-uu/data-protection-policy -- ImageJ mailing list: http://imagej.nih.gov/ij/list.html |
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