// cFOS Cell Segmentation roiManager("reset"); img = getTitle(); loc = getInfo("image.directory"); dir = loc; //File.getParent(loc); roiManager("show all"); roiManager("show none"); // Isolate Sample run("Morphological Filters", "operation=Opening element=Disk radius=10"); inner = getTitle() setAutoThreshold("Huang"); run("Convert to Mask"); run("Options...", "iterations=20 count=1 pad do=Nothing"); run("Erode"); inner = getTitle() roiManager("reset"); // Now get the cells selectWindow(img); run("Duplicate...", " "); smfilt = getTitle(); //run("Subtract Background...", "rolling=4 light"); run("Bandpass Filter...", "filter_large=2 filter_small=1 suppress=None tolerance=5 autoscale saturate"); run("Duplicate...", " "); cellmask = getTitle(); setAutoThreshold("Otsu"); run("Threshold..."); run("Convert to Mask"); run("Distance Transform Watershed", "distances=[Weights (5,7)] output=[16 bits] dynamic=1 connectivity=8"); run("Duplicate...", " "); roimask = getTitle(); setThreshold(1, 65535); run("Convert to Mask"); run("Analyze Particles...", "size=2-Infinity pixel clear add"); run("Set Measurements...", "area mean redirect=None decimal=3"); // Measure cell size and intensity selectWindow(img); roiManager("multi-measure measure_all"); // Save all cells saveAs("Results", dir+"/"+img+"_Results_all.csv"); roiManager("save", dir+"/"+img+"_rois_all.zip"); roiManager("reset"); // Just the inner tissue cells imageCalculator("AND create", roimask, inner); justinner = getTitle(); run("Analyze Particles...", "size=2-Infinity pixel clear add"); selectWindow(img); roiManager("multi-measure measure_all"); saveAs("Results", dir+"/"+img+"_Results_inner.csv"); roiManager("save", dir+"/"+img+"_rois_in.zip"); roiManager("reset"); // Just the outer tissue cells selectWindow(inner); run("Invert"); outer = getTitle(); imageCalculator("AND create", roimask, outer); run("Analyze Particles...", "size=2-Infinity pixel clear add"); selectWindow(img); roiManager("multi-measure measure_all"); saveAs("Results", dir+"/"+img+"_Results_outer.csv"); roiManager("save", dir+"/"+img+"_rois_out.zip"); roiManager("reset"); // Close everything out. Show inner ROIs selectWindow(img); close("\\Others"); roiManager("reset"); roiManager("open", dir+"/"+img+"_rois_in.zip"); /* run("Create Selection"); roiManager("Add"); */