SarConfoCal - Simultaneous Fluorescence and Sarcomere Length Measurements from Laser Scanning Confocal Microscopy (LSCM) Images

Authors: Côme PASQUALIN - François GANNIER (gannier at univ-tours dot fr), University of Tours (France)
See also: Others productions for ImageJ from the authors
PCCV group main page
History: 2016/06/27: first version
Source: Link to GitHub.
Requires: tested with ImageJ 1.50i but should works on some older versions.
Limitations:Works on Mac OS, Linux and Windows
Description: SarConfoCal is used to analyse scanlines and multi-channel (fluorescence and transmission) from Laser Scanning Confocal Microscopy (LSCM) images of myocytes, then it plots the fluorescence signal and the sarcomere length of each line versus time.
Publication: "SarConfoCal: simultaneous sarcomere length and cytoplasmic calcium measurements for laser scanning confocal microscopy images. Bioinformatics." Côme Pasqualin, François Gannier, Angele Yu, Claire O Malecot, Pierre Bredeloux, Véronique Maupoil
Bioinformatics, DOI: 10.1093/bioinformatics/btw754
Installation: Download SarConfoCal.ijm and copy it into the "ImageJ\macros\toolsets" folder.
Start ImageJ or restart it if already opened.
In the "More tools" menu (>>) of the toolbar, select "SarConfoCal". A new set of buttons should now be present on the right side of the toolbar, as shown in the top figure below. SarConfoCal is now ready to use.
How to Use: Open in ImageJ a multi-channel image contening multiple lines from line-scanning confocal microscopy (x-scan horizontally, time vertically). For sarcomere length measurements, you need at least one channel with transmission image.
- Verify the vertical (temporal) calibration (button 1). This should correspond to the time of aqcuisition between two lines. Change it if needed.
- Verify the horizontal (spatial) calibration (button 2). This should correspond to the pixel width. Change it if needed.
N.B.: Most images from confocal microscopy can be imported with Bio-Formats, then spatial and temporal calibration should be correctly done.
- Eventually, you can visualize the FFT spectrum (button 3)
- Then, use button 4 to launch analysis.
Testing: To test SarConfoCal, a linescan file with both fluorescence and transmission images is provided: Fluo4_VG05_linescan_calib0.28.tif. This file was recorded on an Olympus Fluoview 500. Rat ventricular cardiomyocytes was loaded with fluo-4 AM. Channel 1: fluorescence and channel 2: transmission. File is calibrated and ImageJ must set it correctly (time: 0.002 seconds, SL: 0.2773 microns)
Contact Information: For inquiries regarding feature requests, bug reports, or anything else related to the SarConfoCal macro, please email us.

[Menu]

Toolbar providing icons to access SarConfoCal facilities.

[Toolbox 1][Toolbox 2]

Toolboxes for Analysis selection and Sarcomere length measurement.

[FFT Spectrum]

FFT spectrum (bottom right) of the grey profile (top right) of the highlighted line on the image. Red lines show the low and high frequency pass filters. The highest peak between the two red lines is analysed to compute the sarcomere length displayed on the status bar. You can change the analysed line with the 'UP' and DOWN' key or by dragging the line with the mouse.

[Complete analysis]

Complete SL and Fluorescence analysis.

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