I have been looking at colocalization of two proteins in cells with fluoresence microscope. I take three channels of pictures for each slide ( DAPI, green and red). All original files are tiff and 16 bit. They appear to be black and white when I open them with regular photo viewer.
I use imageJ to merge the channels for each slide. Then I convert the tiff to RGB or 8 bit one by one so that I can open them in original color with photoviewer.
After merging, I also have to go back to each slide and split the channels and save each channel as JPEG so that I have pictures of individual channels in color.
I wonder if there's an efficient way to process a batch of slide pictures to merge and split so that I can show "colorful" pictures for each channel and overlay. I stain more than 20 slides a time. The data processing takes me over 3 hours. I just wish there's a smarter to to do this. Thanks.
hi there,
work with tiff files please. because all journals accept your photos in tiff format.
for colouring images follow this:
Image=>look up tables(at the end)=> pseudocolor your pics=> save as them (not save)
for merging pics:
image=> color=> merge channels (make sure the box to composite below it is anticked otherwise it wont save as colored overlay)
hope this helps
vahid
If you are performing a repetitive task on a large number of image files, it is probably easier to use GraphicsMagick or ImageMagick (GM is based on IM but forked some years ago - they offer similar functionality, although I personally prefer GM). These are command-line tools which are designed for batch processing of images. It should be possible to do what you require, perhaps with the aid of a little shell scripting, but you'll need to spend some time reading through the extensive documentation. Easiest to do this with Linux or Mac OS X, but of course there are versions for Windows as well.
ImageJ: A convenient way for doing the same task on a set of images is to import them as an image sequence (File>Import>Image sequence). This will give you a stack of images. Many image operations supprt stack, and you can also save it as a stack of tiffs (single file), or export to individual images. Also, the color split/merge operations can be performed on stack very easily.
Thank you all. Very interesting answers. I will try to explore the different way to simplify my life. thanks again
Hi Jan,
I followed the instructions you suggested. I am able to import a stack of merged files. They are imported as single channels. i.e. I have 5 merged images, each of which has 3 channels. I see 15 images in sequence. They are 16 bit and black/white. I wonder how to make them become colorful like blue, red green. I tried to set to 8 bit, or save as JPEG. They are still black and white.
Have you tried writing a macro for ImageJ? You start by recording all your manual steps Plugins->Macros->Record and then with a little bit of adjusting the code recorded you can run it in a loop.
Hi Zhenning,
you should really look in writing macros, see the attached link. It's very easy. Or Java programming, but that's another story.
To your question:
- if you have RGB files, just import them as a sequence. If you need to work on individual channels, Image>Color>Split Channels
- if you have individual channels, specify the file template for each channel (part of the file name, that all files in this channel have in common), and import each channel separately.
- assign color to the channel: Image>Color>Channels tool>More, select color.
Definitely go through the documentation section at imagej.org, there's lot of useful information there ;-)
http://imagej.nih.gov/ij/developer/macro/macros.html
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