Navigate to Plugins, Macros, Run and locate the fluorescenceintensity.igm macro file in the Autoflux folder.
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2
Select image folder for analysis
Choose the folder containing pre-processed images that have already undergone image processor and calibration macros. The demo uses the Flux vs. Time sample dataset.
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3
Set threshold limit value
Input the upper threshold limit value determined from the calibration macro. For this example, a value of 3 is selected.
▶ 00:57
4
Generate vacuole masks automatically
The macro automatically identifies vacuolar regions using the green fluorescent protein channel and saves the selected area masks as .tif files.
▶ 01:12
5
Quantify red and green fluorescence
The masks are applied to measure fluorescence intensities in both red and green channels of the corresponding image regions.
▶ 01:25
6
Save fluorescence intensity ratios
Ratios of vacuolar red to green fluorescence intensities are automatically calculated and exported to .csv files with matching filenames.
▶ 01:41
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