Batch deconvolution software for ImageJ - by me (Jeff Kuhn). Both Windows, Macintosh (Lion), and Linux-64bit version are included. I have tested the mac version on Lion, the Linux version on Ubunty 12.04 64-bit and the Windows Version on Windows 7 (32 and 64 bit).
Last updated 2012-07-26
The software relies on fftw3 for 3D Fourier transforms. It must be installed either with "apt-get install fftw3*" on linux, "port install fftw3*" on OSX, or by downloading the appropriate windows DLL files from fftw.org.
The software was originialy written as a client-server platform. The server did the actual work of the deconvolution while the client ran on ImageJ and fed stacks to the server. The client-server model was reworked using JNA to allow Java and C code to talk to each other. This sped the transfer of stacks back-and-forth, but precluded the eventual port of the code to a true client-server model with the deconvolution server running on a separate machine or cluster.
The ImageJ portion of the code was written before ImageJ had hyperstacks or virtual stacks. Thus, the code treats all stacks as XYZ-WL-T. Thus, it reads through each stack and splits it into temporary files containing one 3D scan each. It chooses the PSF to use for deconvolution based on the order you set in the options file. If there are still stack planes after iterating through the list of wavelengths, it assumes that each is now a new timepoint and starts the process of writing temporary 3D files again. Once the stack has been split, the ImageJ client feeds the stacks one-by-one to the deconvolution server. I would love to rewrite the client to take advantage of hyperstacks, but I just don't have the time. If anyone else wants to tackle this, please feel free.
Batch processing is handled by the options editor and batch processor plugin menu items in ImageJ. Crop your stacks down to the size you want to deconvolve and place the set of files in a subfolder. Then use the options editor to create a file called "allstacks.dop" in that subfolder. The options file tells the batch processor where to find the measured PSF files for each wavelength. See below for more instructions.
FFTW (http://fftw.org/) is a fantastic platform for doing deconvolution. Not only can it handle sizes that are powers of 2 (2^n), but it does prime-factor algorithms as well. In other words, image dimensions can be any multiple of primes or powers of 2, 3, 5, and 7. For example, an image can be 7x5x8 = 280 pixels on a side.
Here is a list of some prime factors of 2, 3, 5, and 7 that can be used as x, y, and z dimensions for deconvolution:
2 3 5 6 7 9 10 12 14 15 18 20 21 24 25 27 28 30 35 36 40 42 45 48 49 50 54 56 60 63 70 72 75 80 84 90 96 98 100 105 108 112 120 125 126 135 140 144 147 150 160 168 175 180 189 192 196 200 210 216 224 225 240 245 250 252 270 280 288 294 300 315 320 336 343 350 360 375 378 384 392 400 420 432 441 448 450 480 490 500 504 525 540 560 576 588 600 630 640 672 675 686 700 720 735 750 756 768 784 800 840 864 875 882 896 900 945 960 980 1000 1008 1029 1050 1080 1120 1125 1152 1176 1200 1225 1260 1280 1323 1344 1350 1372 1400 1440 1470 1500 1512 1536 1568 1575 1600 1680 1715 1728 1750 1764 1792 1800 1890 1920 1960 2000 2016 2058 2100 2160 2205 2240 2250 2304 2352 2400 2450 2520 2560 2625 2646 2688 2700 2744 2800 2880 2940 3000 3024 3087 3136 3150 3200 3360 3375 3430 3456 3500 3528 3584 3600 3675 3780 3840 3920 4000 4032 4116 4200 4320 4410 4480 4500 4608 4704 4725 4800 4900 5040 5120 5145 5250 …
- Download and install ImageJ
- Download the DecoJ-plugin.zip (3,3 MB) package and unzip it.
- Place the contents of this folder into the ImageJ plugins folder. For example, your folder hierarchy should be: ImageJ/plugins/DecoJ_Client.jar ImageJ/plugins/DecoJNA.dll ImageJ/plugins/lib/... ImageJ/plugins/libDecoJNA.dylib ...
- Run ImageJ.
- Select "Plugins > DecoJ > Server Information" to test the installation.
NOTE: The windows versions of the fftw DLLs are included with DecoJ package.
- Open a terminal window to get a command prompt
- Use the package manager to install libfftw3 sudo apt-get install libfftw3
- Install MacPorts ().
- Open a terminal window to get a command prompt
- Use the package manager to install fftw-3 sudo port install fftw-3
- Take a stack of bead images.
- Draw a small box around a bead.
- Choose Plugins > DecoJ > Auto Shift and Crop PSF.
- Do this for a number of PSFs.
- Add several PSFs together via ImageJ's "Process > Image Calculator..."
- Divide by the number of PSFs you added together via "Process > Math > Divide..."
- Save this PSF
- Create an empty folder to contain the images to deconvolve.
- Use "Image > Duplicate" to crop out a portion of your image stack to deconvolve.
- Save the image to that folder. NOTE: DecoJ expects the image stacks to be sorted by Z-plane, then Color, then Time.
- Run "Plugins > DecoJ > Options Editor..."
- Type in the number of itterations.
- Type in the number of Z planes in your stack.
- Select the number of wavelengths. DecoJ uses the number of Z planes and number of wavelengths to parse large stacks into individual components for deconvolution.
- For each wavelength, point to the PSF file. NOTE: The directory will likely be different for each computer you run DecoJ on, so make sure and point to the correct PSF file.
- Save the "allstacks.dop" file to the folder with the images you selected for processing above. Close the options editor. DecoJ will try to deconvolve every file in a folder containing the "allstacks.dop" file. Make sure the folder just contains image stacks and the ".dop" file.
- Run "Plugins > DecoJ > Deconvolve Batch" and select the "allstacks.dop" file you just created.
- DecoJ will show you a list of files to deconvolve.
- Approve the operation, sit back, and wait. NOTE: If you quit ImageJ or close the ImageJ main window, deconvolution will STOP. Be careful.
- DecoJ creates a subfolder named "out" to store the deconvolved stacks. You can peek at them in ImageJ as they come out, but do not do too much image processing while DecoJ is running in the background.
- I suggest a test run with 10 or so iterations to try it out.
NOTE: I have included some Test Data (95.4 MB) of a large bead and a PSF to make sure everything is working. You will have to change the location of the PSF file in the "allstacks.dop" example file. You can load it with the options editor, modify it, and save it. You can also edit options file with a simple text editor.
- Let me know if you have any problems.
- The source code is available upon request if you want to try compiling it or modifying it. Note that I have made several changes over the years and the source by now has become somewhat "cobbled together". The whole thing needs a major rewrite if I ever have the time.