ClusterIndex is a Jython script for the quantiative evaluation of cell spatial clustering. To measure the degree of cell clustering, we used “nucleus-nucleus distance index (NND)”.
Fiji.
Note that it does not work with pure ImageJ as the script uses auto local threshold function.
Download the pakage from the URL linked below, unzip the file and open the script "ClusterIndex.py" in Fiji script editor. See "Workflow" section below for a detailed workflow.
http://dx.doi.org/10.5281/zenodo.18246
Images with Hoechst labeled nuclei are first binarized using intensity-based automatic image threshold using Otsu algorithm followed by binary fill-hole and open operations. Number of nucleus (
NND is computed by dividing the number of cell clusters (
NND is close to 1.0 when cells are sparsely distributed while it is close to 0 as cells become clustered.
Download the follwoing nucleus image files and open them in Fiji.
Click the image window "cluster.jpg" and bring the image to the front. Then run the script "ClusterIndex.py".
Click the image window "random.jpg" and bring the image to the front. Then run the script "ClusterIndex.py"
After these two actions, Log window should be showing follwoing measurement results.
==== cluster.jpg =====
Number of Nucleus : 237
Clusters at dilation 5: 86
Clusters/Nucleus 0.362869198312
==== random.jpg =====
Number of Nucleus : 346
Clusters at dilation 5: 280
Clusters/Nucleus 0.809248554913
The NND index ("Cluser/Nucleus") of "cluster.jpg" is 0.36 and that of "random.jpg" is 0.81. This suggest that nucleus in "cluster.jpg" is more clusterd.
To increase / decrease the width of dilation, change the number in line 22 ("SAMPLEITER = 5"). If the radius of cell is 10 pixels, then use that value and change it to "SAMPLEITER = 10".
To view intermediate images during processing, comment-out line 23 and uncommnet line 24 ("VERBOSE = True")
Kota Miura ( http://wiki.cmci.info )
This script was developed for the analysis reported in the paper below.
Fukuda, S., Nishida-Fukuda, H., Nanba, D., Nakashiro, K., Nakayama, H., Kubota, H., Higashiyama, S., 2016. Reversible interconversion and maintenance of mammary epithelial cell characteristics by the ligand-regulated EGFR system. Sci. Rep. 6, 20209.