information should not be used for analysis since the units will be
arbitrary.
3.5.2 Position in μm (and
Other Specified Units)
This is the same information as described above but corrected for
the dimensions as detailed in the properties of the original file. That
information was adjusted in step 1 of Subheading 3.2. It is detailed
in columns J, K, and L, named “X [μm],” “Y [μm],” and “X [μm],”
respectively. If the units are different to μm, the names will display
those.
3.5.3 Time
There are two columns named “Time [time],” column G and
N. They both display the same information: the time point
corresponding to the values for X, Y, and Z in that row. For cells
tracked from the beginning of the movie, this list will start at
0. Otherwise it will start at the time point where the track was
initiated.
3.5.4 List Name
The last column on the file, column X, is titled “ListName” and it
tells us which cell the track corresponds to. If you have taken note
of their identities during the tracking (see Subheading 3.4, step 7),
you can identify each cell by the number in this column.
Calculation of speed, velocity, and persistence is performed
using simple Euclidian formulas or as per Gorelik and
Gautreau [12].
3.5.5 Visualization of 3D
Tracks in 2D
It is usually easier to generate 2D representations of View5D tracks
to visualize and qualitatively compare several tracks at a time. We
perform this step by uploading View5D tracks onto the “Manual
Tracking” plugin of Fiji. This can be performed manually, if you
wish to select precisely which tracks to visualize, for example only
cells of a certain type; or automatically if you wish to have a
representation of all the tracks.
This step requires to generate an Excel sheet that contains
tracking (X, Y position and time) and cell identity information in
a manner that Manual Tracking can read.
For manual conversion, follow steps 1–9:
1. Create a new workbook on Microsoft Excel and copy the
information in the following columns for each of the tracks to
be visualized.
2. Name the columns as follows: “Track n” on cell B1, “Slice n”
on C1, “X” on D1, and “Y” on E1 (leave column A free, see
below).
3. Under “Track n,” on column B, add the number of the track.
Start with the track with the lowest number; it is easier to
maintain the numbers View5D assigned to each track.
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Zain Alhashem et al.
arbitrary.
3.5.2 Position in μm (and
Other Specified Units)
This is the same information as described above but corrected for
the dimensions as detailed in the properties of the original file. That
information was adjusted in step 1 of Subheading 3.2. It is detailed
in columns J, K, and L, named “X [μm],” “Y [μm],” and “X [μm],”
respectively. If the units are different to μm, the names will display
those.
3.5.3 Time
There are two columns named “Time [time],” column G and
N. They both display the same information: the time point
corresponding to the values for X, Y, and Z in that row. For cells
tracked from the beginning of the movie, this list will start at
0. Otherwise it will start at the time point where the track was
initiated.
3.5.4 List Name
The last column on the file, column X, is titled “ListName” and it
tells us which cell the track corresponds to. If you have taken note
of their identities during the tracking (see Subheading 3.4, step 7),
you can identify each cell by the number in this column.
Calculation of speed, velocity, and persistence is performed
using simple Euclidian formulas or as per Gorelik and
Gautreau [12].
3.5.5 Visualization of 3D
Tracks in 2D
It is usually easier to generate 2D representations of View5D tracks
to visualize and qualitatively compare several tracks at a time. We
perform this step by uploading View5D tracks onto the “Manual
Tracking” plugin of Fiji. This can be performed manually, if you
wish to select precisely which tracks to visualize, for example only
cells of a certain type; or automatically if you wish to have a
representation of all the tracks.
This step requires to generate an Excel sheet that contains
tracking (X, Y position and time) and cell identity information in
a manner that Manual Tracking can read.
For manual conversion, follow steps 1–9:
1. Create a new workbook on Microsoft Excel and copy the
information in the following columns for each of the tracks to
be visualized.
2. Name the columns as follows: “Track n” on cell B1, “Slice n”
on C1, “X” on D1, and “Y” on E1 (leave column A free, see
below).
3. Under “Track n,” on column B, add the number of the track.
Start with the track with the lowest number; it is easier to
maintain the numbers View5D assigned to each track.
98
Zain Alhashem et al.