4-colour merge?
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Hi All,
I'm a relatively new ImageJ user and would like to merge 4-colours into one composite image (CY3, CY5, Alexa488 and DAPI). Is this possible in ImageJ? How would I do this? Thank you, Shane |
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Dear Shane.
Your eyes have 3 color sensors, red green and blue, your computer screen had red green and blue pixels (and in printing its cyan, yellow, magenta and black) So, now if you want to merge 3 colours, thats fine, make each colour channel one of red green and blue, It will look very pretty (hopefully). Now with 4 colours you are faced with a big problem. Your eyes are not capable of picking out 4 different colours out of an RGB image. As far as your eyes are concerned, all colours are made of some mixture of red, green and blue. So if you already have a red, green and blue channel, then try to add 1 more, what colour would you pick? Magenta... no, because thats a mix of red and blue. Similar for cyan and yellow, and other colours. So, if you do mix red green blue and magenta, you will end up with a pretty image, but it will be impossible to tell which parts of the red part of the image come from the red channel or from the magenta channel. Some people use red, green blue and white, but you are faced with that same problem . There cold be a place with almost equal red green and blue intensity (= white) which you could not tell from a white pixel If you are just trying to make a pretty picture, and not say anything quantitative or too specifically qualitative, you might get away with RGB+white. If i reviewed it or saw it in talk, I would raise the points made above. If you are trying to look for colocalisation (or more specifically spatial intensity correlation) you can do this in pairwise way, using the imageJ colocalisation plugins (read the docs at http://www.macbiophotonics.ca/imagej/ ) Looking at a 2D intensity histogram / scatterplot of once channel vs a second gives a much better way of visualising the spatial intensity correlation between then than a colour merge image. In my opinion, colour merge images are hard to interpret, can be misleading, are a poor way of visualising "colocalisation" and this researcher would send back any paper to an author who has used colour merge images to try to show between channel spatial intensity correlations (colocalisation) In fact I would go as far as to say, wrong interpretation of colour merge images could ruin you career. 2D hisotgrams / scatter plots are a much better way to show it. cheers Dan On Nov 11, 2007, at 6:00 AM, IMAGEJ automatic digest system wrote: > Date: Sat, 10 Nov 2007 16:25:55 -0500 > From: Shane Harding <[hidden email]> > Subject: 4-colour merge? > > Hi All, > I'm a relatively new ImageJ user and would like to merge 4-colours > into one composite image (CY3, CY5, Alexa488 and DAPI). Is this > possible in ImageJ? How would I do this? > > Thank you, > Shane Dr. Daniel James White BSc. (Hons.) PhD Senior Microscopist / Image Processing and Analysis Light Microscopy Facility Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstrasse 108 01307 DRESDEN Germany +49 (0)176 61262128 (German Mobile) +49 (0)351 210 2627 (Work phone at MPI-CBG) +49 (0)351 3265048 (German Festnetz "landline" - cheaper) +49 (0)351 210 1078 (Fax MPI-CBG LMF) +358 (0) 468102840 (Finnish mobile, only when I'm in Finland) http://www.bioimagexd.net http://www.chalkie.org.uk [hidden email] ( [hidden email] ) |
Macro language exponential number format?
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Hi there,
working with my copy of ImageJ 1.38x i just discover that the macro language does not accept statements like a = 5e-3 ; or a = 5.e-3; or a = 5.0e-3; (a = 0.005 works, however) Am I totally confused here? Is that a "feature" that only recently crept in? Sincerely Joachim ______________________________________________________________________ This email has been scanned by the MessageLabs Email Security System. For more information please visit http://www.messagelabs.com/email ______________________________________________________________________ |
Re: Macro language exponential number format?
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On Nov 11, 2007, at 10:53 AM, Joachim Wesner wrote:
> Hi there, > > working with my copy of ImageJ 1.38x i just discover that the macro > language does not accept statements like > > a = 5e-3 ; > > or > a = 5.e-3; > or > a = 5.0e-3; > > (a = 0.005 works, however) > > Am I totally confused here? Is that a "feature" that only recently > crept > in? > Hi, You could use the pow() function. For example... print(5 * pow(10,-3)); results in 0.0050 Cheers, Ben Ben Tupper Bigelow Laboratory for Ocean Science 180 McKown Point Road POB 475 West Boothbay Harbor, ME 04575 |
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In reply to this post by Joachim Wesner
Hi,
On Sun, 11 Nov 2007, Joachim Wesner wrote: > working with my copy of ImageJ 1.38x i just discover that the macro > language does not accept statements like > > a = 5e-3 ; > > or > a = 5.e-3; > or > a = 5.0e-3; > > (a = 0.005 works, however) > > Am I totally confused here? Is that a "feature" that only recently crept > in? No, this is a long standing shortcoming. I just fixed it in ImageJA, and will send Wayne a patch. If you are interested in the patch, see http://repo.or.cz/w/imageja.git?a=commitdiff;h=2ff92a2c562bacce89f51a20171006914ba9d615. Ciao, Dscho |
Antwort: Re: Macro language exponential number format?
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Hi,
thanx a lot. It´s not that I could not live without it, it´s just a strange behaviour that I noted when working on macro. I think/thought that it should accept exponential format, as the macro language in all other respect is already very similar to basic Java or C. Wayne has already supplied me with a new IJ snapshot that has the patch built in, great. I wish other software would have such a quick bugfix cycle, excellent! Sincerely Joachim Johannes Schindelin <Johannes.Schinde An [hidden email]> [hidden email] Gesendet von: Kopie ImageJ Interest Group Thema <[hidden email]. Re: Macro language exponential GOV> number format? 11.11.2007 21:28 Bitte antworten an ImageJ Interest Group <[hidden email]. GOV> Hi, On Sun, 11 Nov 2007, Joachim Wesner wrote: > working with my copy of ImageJ 1.38x i just discover that the macro > language does not accept statements like > > a = 5e-3 ; > > or > a = 5.e-3; > or > a = 5.0e-3; > > (a = 0.005 works, however) > > Am I totally confused here? Is that a "feature" that only recently crept > in? No, this is a long standing shortcoming. I just fixed it in ImageJA, and will send Wayne a patch. If you are interested in the patch, see http://repo.or.cz/w/imageja.git?a=commitdiff;h=2ff92a2c562bacce89f51a20171006914ba9d615 . Ciao, Dscho ______________________________________________________________________ This email has been scanned by the MessageLabs Email Security System. For more information please visit http://www.messagelabs.com/email ______________________________________________________________________ |
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In reply to this post by Shane Harding-2
Joachim Walter wrote a nice plugin called Image5D. It's found at
http://rsb.info.nih.gov/ij/plugins/image5d.html . You can load your four channels and pick a color for each. I don't know much about colocalization or quantifying fluorescence, but I would only use this to make demonstration images. Justin On Nov 10, 2007 3:25 PM, Shane Harding <[hidden email]> wrote: > Hi All, > I'm a relatively new ImageJ user and would like to merge 4-colours > into one composite image (CY3, CY5, Alexa488 and DAPI). Is this > possible in ImageJ? How would I do this? > > Thank you, > Shane > |
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Shane,
there is a plugin called "Colour Merge" which can merge every colour, also violet plus yellow and so on. As already mentioned, such an extensive overlay doesn't make sense for quantitation. In my opinion one can use it if the structures are labeled with one colour at once only, i.e. nuclei, membrane, endosomes etc. separately - to get an overview of the situation in the cell/tissue. cheers, Michael Justin McGrath wrote: > Joachim Walter wrote a nice plugin called Image5D. It's found at > http://rsb.info.nih.gov/ij/plugins/image5d.html . You can load your > four channels and pick a color for each. I don't know much about > colocalization or quantifying fluorescence, but I would only use this > to make demonstration images. > > Justin > > > > On Nov 10, 2007 3:25 PM, Shane Harding <[hidden email]> wrote: >> Hi All, >> I'm a relatively new ImageJ user and would like to merge 4-colours >> into one composite image (CY3, CY5, Alexa488 and DAPI). Is this >> possible in ImageJ? How would I do this? >> >> Thank you, >> Shane |
import image sequence of raw images
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In reply to this post by Justin McGrath
Hi,
I would like to import a sequence of raw images (indicating the parameters like the x and y sizes, the Offset of the header to skip....). I am sure it is a common job... and then save as an another raw format (after removing the header). Thank you. Eric |
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In reply to this post by Michael Weber-4
Thank you all for your help.
I think the solution to my question is to use the DAPI image to create a boundary in which I can use the RGB labels. Is there an easy way to create an image with the DAPI regions outlined so that I may overlay my other 3 RGB coloured on this? I agree that pairwise will also work, I'm just searching for different, but acceptable options. Thanks Again, Shane Quoting Michael Weber <[hidden email]>: > Shane, > > there is a plugin called "Colour Merge" which can merge every colour, > also violet plus yellow and so on. > > As already mentioned, such an extensive overlay doesn't make sense for > quantitation. In my opinion one can use it if the structures are > labeled with one colour at once only, i.e. nuclei, membrane, endosomes > etc. separately - to get an overview of the situation in the > cell/tissue. > > cheers, > Michael > > > Justin McGrath wrote: >> Joachim Walter wrote a nice plugin called Image5D. It's found at >> http://rsb.info.nih.gov/ij/plugins/image5d.html . You can load your >> four channels and pick a color for each. I don't know much about >> colocalization or quantifying fluorescence, but I would only use this >> to make demonstration images. >> >> Justin >> >> >> >> On Nov 10, 2007 3:25 PM, Shane Harding <[hidden email]> wrote: >>> Hi All, >>> I'm a relatively new ImageJ user and would like to merge 4-colours >>> into one composite image (CY3, CY5, Alexa488 and DAPI). Is this >>> possible in ImageJ? How would I do this? >>> >>> Thank you, >>> Shane |
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In reply to this post by Justin McGrath
Justin McGrath a écrit :
> Joachim Walter wrote a nice plugin called Image5D. It's found at > http://rsb.info.nih.gov/ij/plugins/image5d.html . You can load your > four channels and pick a color for each. I don't know much about > colocalization or quantifying fluorescence, but I would only use this > to make demonstration images. > > Justin > > > > On Nov 10, 2007 3:25 PM, Shane Harding <[hidden email]> wrote: > >> Hi All, >> I'm a relatively new ImageJ user and would like to merge 4-colours >> into one composite image (CY3, CY5, Alexa488 and DAPI). Is this >> possible in ImageJ? How would I do this? >> >> Thank you, >> Shane >> >> to only make a demo image you could use aswell Photoshop or the freely available "the gimp" and work around with overlays. Sheers, Fabrice. -- Senger Fabrice |
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I have found that a quick way to merge images of non-RGB colors is to
use the Past Control and set it to Add ... John Senger Fabrice wrote: > Justin McGrath a écrit : >> Joachim Walter wrote a nice plugin called Image5D. It's found at >> http://rsb.info.nih.gov/ij/plugins/image5d.html . You can load your >> four channels and pick a color for each. I don't know much about >> colocalization or quantifying fluorescence, but I would only use this >> to make demonstration images. >> >> Justin >> >> >> >> On Nov 10, 2007 3:25 PM, Shane Harding <[hidden email]> wrote: >> >>> Hi All, >>> I'm a relatively new ImageJ user and would like to merge 4-colours >>> into one composite image (CY3, CY5, Alexa488 and DAPI). Is this >>> possible in ImageJ? How would I do this? >>> >>> Thank you, >>> Shane >>> >>> > Hi there, > > to only make a demo image you could use aswell Photoshop or the freely > available "the gimp" and work around with overlays. > > Sheers, > > Fabrice. > -- John K. Alexander, Ph.D. Post-Doctoral Fellow William Green Laboratory University of Chicago Dept. Neurobiology, Pharmacology, and Physiology 947 East 58th Street Abott Hall 402 Chicago, IL 60637 (off) 773-702-9386 (fax) 773-702-3774 [hidden email] |
Re: import image sequence of raw images
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In reply to this post by Eric BADEL
Hello,
I had the same problem so I created a new file format I name IJG for ImageJ Graphics. It is very simple. I don't know if there would be much interest in it. I have a program to convert any raw data file into ijg file type. It may drop any number of header bytes in the raw data file. It basically simply add the 16 header bytes in front of the raw data. I also heard of a small program that could add a DICOM header which is probably more interesting because you will be able to read the file with all programs supporting DICOM. I would be curious to compare complexity of the code and file changes. IJG requires only a 16byte header addition that specify the file type, the pixel type and the width, height and depth size. It would be nice if ImageJ could add support for a specific FileReaderPlugins. This plugin class would be derived from the Plugin class and simply add one method to tell if the reader can open the given file. The arguments would be the file name and a buffer holding the 1000 first bytes as is currently done in the HandleFileReader.java program. ImageJ could try all the plugins in InputOutput that are derived from the FileReaderPlugin class. Even those without an underscore in their names. The file open attempt would be performed in the lexicographic plugin name order. Adding a number in front of the name would then allow to control the order of plugin usage. The change required to support this would ensure backward compatibility since the HandleFileReader.java would try use FileReaderPlugins before the current code which would work as a fallback. |
Re: import image sequence of raw images
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In reply to this post by Eric BADEL
Here is the link to the programs to convert raw images into dicom. It's
for Linux http://www.dclunie.com/dicom3tools.html -- Bien cordialement, Ch. Meessen |
Re: import image sequence of raw images
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In reply to this post by Eric BADEL
You can import a sequence of raw images as a stack by using
File>Import>Raw and checking "Open All Files in Folder" in the dialog. You can save a stack as a raw image sequence by using File>Save As>Image Sequence and and choosing "Raw" as the format. -wayne On Nov 12, 2007, at 10:47 AM, Eric BADEL wrote: > Hi, > I would like to import a sequence of raw images (indicating the > parameters like the x and y sizes, the Offset of the header to > skip....). > I am sure it is a common job... > > and then save as an another raw format (after removing the header). > Thank you. > Eric > |
Re: import image sequence of raw images
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Very good. One more detail. I openned all the files as they are little
endian order. But when I save them, they are are not saved with Little Endian Order (I need to uncheck the option to read them again). It is bizarre. How could I force the little endian order using the Save As>Image Sequence and and choosing "Raw" as the format ? Thank you. Eric Wayne Rasband a e'crit : > You can import a sequence of raw images as a stack by using > File>Import>Raw and checking "Open All Files in Folder" in the dialog. > You can save a stack as a raw image sequence by using File>Save > As>Image Sequence and and choosing "Raw" as the format. > > -wayne > > On Nov 12, 2007, at 10:47 AM, Eric BADEL wrote: > >> Hi, >> I would like to import a sequence of raw images (indicating the >> parameters like the x and y sizes, the Offset of the header to >> skip....). >> I am sure it is a common job... >> >> and then save as an another raw format (after removing the header). >> Thank you. >> Eric >> > > |
Re: import image sequence of raw images
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In reply to this post by Eric BADEL
Check "Export Raw in Intel Byte Order" in the Edit>Options>Input/
Output dialog box. -wayne > Very good. One more detail. I openned all the files as they are little > endian order. But when I save them, they are are not saved with Little > Endian Order (I need to uncheck the option to read them again). It is > bizarre. How could I force the little endian order using the Save > As>Image Sequence and and choosing "Raw" as the format ? > Thank you. > Eric > > > Wayne Rasband a e'crit : > > You can import a sequence of raw images as a stack by using > > File>Import>Raw and checking "Open All Files in Folder" in the > dialog. > > You can save a stack as a raw image sequence by using File>Save > > As>Image Sequence and and choosing "Raw" as the format. > > > > -wayne > > > > On Nov 12, 2007, at 10:47 AM, Eric BADEL wrote: > > > >> Hi, > >> I would like to import a sequence of raw images (indicating the > >> parameters like the x and y sizes, the Offset of the header to > >> skip....). > >> I am sure it is a common job... > >> > >> and then save as an another raw format (after removing the header). > >> Thank you. > >> Eric > >> > > > > > > |
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