This page describes in a simplified way the first steps required
to start using MicrobeTracker and
MicrobeTracker tools. See also more detailed
tutorials based on the included datasets. Overall, the tutorials are listed in
the order of increasing complexity:
Additionally, the help pages for some tools have extended
examples of their usage:
SpotFinderZ (finding round / diffraction-limited spots in cells),
SpotFinderF (finding round / diffraction-limited spots in an image),
MicrobeTracker requires MATLAB with Image Processing
Toolbox to run, though it can be compiled and will run stand-alone. However,
MATLAB will still be needed to view or process the output files and run the
tools. Here are the instructions how to install the MATLAB-based version:
Unpack or copy the content of MicrobeTracker package into
Add this folder to the list of default paths in MATLAB (in
MATLAB select File>Set Path>Add folder, then select the MicrobeTracker
folder and click Save)
Now you can run MicrobeTracker by typing microbeTracker in
MATLAB workspace window and the rest of the functions by typing the function's
Optionally, you can add support for images in other formats
than series of single-page TIFF files and multi-page TIFF files by installing
(the images in this case have to be in separate stack files for each filter
block / color). To do so, exit MicrobeTracker, download
file, and place it to the MicrobeTracker folder.
For program and functions description see help either by
clicking Help button in MicrobeTracker (for the program itself) or opening help
(htm) file from the MicrobeTracker folder (for the rest of the functions).
Sometimes MicrobeTrackers's compiled functions will not run
because of some libraries missing on your operating system. To test, type in
MATLAB command window a=intxy2C(0,0,0,0), which
should return an empty matrix, not an error. If it returns an error, install the
compatible C++ compiler, which can be found on
MathWorks website (e.g.
for version 2011a; make sure you select your operating system).
Prepare the image files to load. If you are using the
program for the first use, select images with very clearly visible and well
separated cells. This description will focus on using phase contrast microscopy
(called simply phase below) images, see the detailed help for
fluorescence or DIC. The files must be in
TIFF format placed in a separate folders for phase and corresponding
fluorescence images. If the files have to be in a Make sure that the particular
order (e.g. timelapse), nake sure that they are sorted correctly by the
operating system. Usually the files have the same base name followed by a
number. In this case the numbers have to be x01, x02, ... , x09, x10, x11, ...
as opposed to x1, x2, ... , x10, ...
Load the phase images by clicking 'Load phase data' button
on the top panel and selecting the folder with the images. If fluorescence
images are used as well, load them clicking 'Load signal 1' button.
Load parameter set by clicking 'Load parameters' button
from the Parameters panel. Typically select 'alg4.set' from the MicrobeTracker
folder. It is based on algorithm 4, designed to work with any rod-shaped cells,
any curvature and length. See
the detailed help to figure out what algorithm to use for specific cases.
Adjust the most critical parameters by modifying the text
in the 'Parameter' control on the bottom right:
- set areaMin and areaMax so that the area of all good cells falls into the
range between these two values. If not known precisely, set areaMin much smaller
and areaMax much larger, and then adjust them more presizely (the area of a
detected cell can be seen on the 'Information' panel on the bottom left once the
cell is selected).
- set cellwidth to the width of the cell in pixels (approximately).
- for filamentous cells increase fsmooth, to measure
variations in cell width reduce wspringconstant, for
curved cells reduce rigidityB, for timelapses with
dividing cells set splitThreshold to ~ 0.4, see the
detailed help for other parameters.
If fluorescence needs to be added, subtract the background
by selecting 'All' and clicking 'Subtract bgrnd' button on the 'Background
subtraction' panel (bottom center) and select 'Compute signal 1 profile' on
the 'Detection & analysis' panel.
Select 'Time lapse' or 'Independent frames' mode on the
'Detection & analysis' panel.
Click on 'This frame' button to see the performance on one
frame. Wait for up to a few minutes to process the frame.
If the detection was good, click 'All frames' to process
the whole image set. This may take up to several hours, depending on the number
and size of the images.
After the program finishes working, save the results by
clicking 'Save analysis' button.
This section describes how to use the functions for cell dimensions and
intensity statistics. This description is primarily for those who have no MATLAB
experience in order to start using the functions. If you are familiar with basic
MATLAB concepts, refer directly to the description of the
- Load the cellList variabe into MATLAB workspace. To do so, drag and drop
the meshes (results) file into the workspace or type
<filename> with the name of the file). You only need the variable called
- Now run a function according to its description
. For example, to plot a histogram of cell lengths, type:
here the command figure produces a new figure.
Otherwise the histogram will replot the existing figure or will be plotted
inside the MicrobeTracker window.
In order to compare data from two experiments on the same plot you will need
cellList from both loaded at the same time. For this rename this variable after
loading, for example:
- Load the first file the same way as above. Then rename the cellList:
list1 = cellList;
- Now load the second file and rename the cellList:
list2 = cellList;
- Now run the function with two arguments:
In order to specify additional parameters, put them after cellList. Most of
the other parameters are optional and the order is not important. If the order is
important (when the function cannot distinguish the parameters based on the
variable type), place an empty array  to omit a parameter. For example, to show
an overlapping histogram (i.e. plotting the overlaps of bars with another color),
To specify the x-axis (an array of lengths of
the cell in which the bin centers will be placed), type:
The record 5:10:100 means that the
centers will be places in positions from 5 to 100 pixels with the step of 10.
Usually specify the position of the first bin center at one half of the bin
width, so that the bins start from zero.