文件列表:

trackmovie_wrapped.m (1132, 2003-05-06)
trackstats.m (2245, 2003-09-26)
velocityjdj.m (1362, 2003-05-06)
ExampleInput.txt (678, 2004-01-21)
track_stuff_queued.m (1982, 2003-05-20)
track_stuff_queued_movie.m (1981, 2003-05-20)
trackbeads.m (3924, 2003-04-29)

Particle/Bacterial Tracking Software Readme File ================================================================================ Contents: -------------------- I. Copyright notice II. Introduction III. Requirements IV. Installation V. File Formats VI. Syntax and Examples VII. Notes VIII. Included F-iles -------------------- I. Copyright notice This software is copyright (C)2003 Nicholas Darnton, Jacob D. Jaffe, and the President and Fellows of Harvard University. It may be redistributed freely providing that this copyright notice is included and the software remains in its original, unmodified form. The end user may modify the software for their own needs but ANY MODIFIED VERSION OF THE SOFTWARE MAY NOT BE REDISTRIBUTED. The software is provided "as is" with no guarantees of performance, functionality, compatibility, or technical support. Use of this software is at your own risk. -------------------- II. Introduction This software was designed by Nicholas Darnton for purposes of tracking small fluorescent beads in a microscope field over time. It does so by using a "nearest neighbor" approach comparing adjacent frames in a movie file. This version is modified for tracking gliding bacteria in a microscope field. In general, it should be able to track any moving object(s) in a movie provided that there is sufficient contrast between the object(s) being tracked and the background of the image. Particular applications may require the end user to modify various parameters. This software is a series of scripts to be run in MATLAB and is not a stand-alone software package. This version is designed to accept a "queue" file of movie analysis tasks to make automated measurements easier. Operation of the queueing system will be described later in (V.) File Formats. The basic input to the program is a movie file and the program will output the average velocity and standard deviation of the objects tracked in the analysis period. It will also report the number of objects tracked as well as the number that are considered to be moving (with velocity above a user defined threshold). The user also supplies various ancillary data such as scale and contrast thresholding value. The program does not generally differentiate between types of objects being tracked - i.e., big objects will be lumped together with small ones. Again, this may be modified with some tweaks to the scripts at the user's discretion and peril. Optionally, the program can output a movie with the tracks it drew overlaid onto the original frames. -------------------- III. Requirements This software requires a functional copy of MATLAB with the MATLAB Image Analysis Toolbox installed. It was developed and tested with version 6.5 release 13 but may work on other versions. MATLAB may be obtained from The Mathworks (www.mathworks.com). This software has been successfully executed in both Windows and MacOS environments. It may work in other environments as well, provided that MATLAB is also functional in that environment. These instructions and examples are written from the Windows PC point of view. You may need to modify accordingly for other platforms. Movie files are required to be AVI format with INDEO Video 5.10 com- pression. You may wish to use a program such as Adobe Premier to convert any other movie file format into the required format. This version assumes that you will be tracking DARK OBJECTS on a LIGHT BACKGROUND. -------------------- IV. Installation Simply unzip the files into a directory. Then run your MATLAB program and make the "Current Directory" the one where you unzipped the files. This archive also includes an example movie and an example queue file to help get you started. Move the files "ExampleMovie-Indeo.avi" and "ExampleInput.txt" to the directory C:\TEMP, creating it if it does not already exist. -------------------- V. File Formats INPUT FILES: Movie Files: Must be in AVI format with Indeo video 5.10 compression. - use Adobe Premier or equivalent to interconvert file formats. - This version assumes that youwill be tracking DARK OBJECTS on a LIGHT BACKGROUND. Queue File: The queue file is a tab-delimited text file where each row is a separate analysis task and the columns are defined as follows (see "ExampleInput.txt" for an example): Column 1: Output directory (full path, with trailing \) (string) Column 2: Analysis task label (string) Column 3: Full path and file name of the movie input file (string) Column 4: Analysis task label 2 (should be duplicate of column 2) (string) Column 5: Threshold value (a decimal from 0.0 to 1.0) - this number scales onto the 256 possible gray levels and determines the thresholding cutoff for objects to include as the image is converted to a binary equivalent. 0.5 is generally OK. Column 6: First frame to analyze (w/r/t movie file in Col. 3) (integer) Column 7: Last frame to analyze (w/r/t movie file in Col. 3) (integer) Column 8: Frame rate of movie (frames / sec) (decimal or integer) Column 9: Scale of movie (pixels / unit length) (decimal) Column 10: Search radius (How many unit lengths to search from one frame to the next for the same objects. This can be a decimal. Small for slow objects, larger for fast objects.) (decimal) Column 11: Minium number of contiguous frames that an object must be detected in to be considered "real." (integer) Column 12: Velocity threshold (in unit lengths / sec) that an object must be going to be considered "moving." (decimal) Column 13: Velocity thresholding state (0 for off, 1 for on) - sets whether velocity thresholding is performed. If it is "off," even stationary objects will be reported in the average speed. If it is on, only objects with velocity greater than that specified in Col. 12 will be included in average speed calculations. There is no limit to the number of analysis lines possible in the queue file. OUTPUT FILES: For each analysis task (i.e., row of the queue file), two output files are generated. Both can be found in the output directory specified in the queue file and will have the base name specified in columns 2 and 4 of the queue file. One is a tab-delimited text file with the following format: Column 1: Average speed Column 2: Standard deviation of average speed Column 3: Number of objects counted Column 4: Number of objects considered to be moving Column 5: Percentage of objects moving Note: "-stats.txt" is appended to the queue file analysis task label. The second is a MatLab datafile that simply contains a list of velocities for the objects tracked in the analysis. MOVIE OUTPUT: If track_stuff_queued_movie is invoked, an AVI movie file is generated and placed in the output directory, again with the base name specified in the queue file. Generating output movies takes a really long time. -------------------- VI. Syntax and Examples Assuming that MATLAB is loaded and you have changed the "Current Directory" to the one that contains the MATLAB scripts, it is very easy to run. You just need to make sure that you have followed all the requirements concerning file formats and the queue file as stated above. There are two ver- sions that may be invoked. At the MATLAB prompt, type: track_stuff_queued(''); - or - track_stuff_queued_movie(''); where is the full path and filename of the queue file. Note the single quotes (') surrounding the file name. The second version outputs a movie which is basically the same as the input movie but with the tracks generated by the program overlaid in multiple colors. This option will take a very long time to run and you can't touch the computer while it's runing or else the output movie gets corrupted. It does make nice movies, though. Assuming that you have moved the ExampleMovie-Indeo.avi file and ExampleQueue.txt files to "C:\TEMP", you can invoke the following example: track_stuff_queued('C:\TEMP\ExampleQueue.txt'); You will see some dots go across the screen as it processes each analysis line in the queue file, and then a bunch of numbers will be output from the program. These numbers are all saved in permenant files as specified in the queue file (see the File Formats section above). For the example, you should see files called: TP00-stats.txt TP00.m TP10-stats.txt TP10.m ...etc. appearing in C:\TEMP as the program processes. You are encouraged to look at the example queue file so that you can better understand its structure. Using a spreadsheet program such as Excel is extremely useful for setting up queue files. Just be sure to save as a tab-delimited text file. If you invoke the _movie version, additional .AVI files will also appear in C:\TEMP. Again, this will take a really long time and you should not use the _movie version for routine use. -------------------- VII. Notes - It is not recommended to routinely generate track movies for every analysis unless you don't mind locking up your computer for long periods of time. - Unfortunately we do not have the resources to provide technical support on the programs, but if you find it to be useful, feel free to drop us a line. jdjaffe#fas.harvard.edu ndarnton#rowland.harvard.edu (convert the # to a @ - this is an effort to reduce spam) Hackers guide: - Most of the code is commented. Feel free to inspect it. - To analyze light objects on a dark background, comment out (with a %) line 33 (bw=~bw) in trackstats.m. - To alter the minimum object size, edit value in line 35 of trackstats.m (default is set to 9 pixels). - To see what is going on as you run it, it is useful to put in breakpoints and then checking the frames as they are processed. This happens mostly in trackstats.m. You can adjust the thresholding and size parameters by do this, and then using the imshow command as needed. Just "step through" the program as it executes.

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