From bea2216b30f8167da9a4442a21ffb6450db0c352 Mon Sep 17 00:00:00 2001
From: Elisabeth Kugler <67630046+ElisabethKugler@users.noreply.github.com>
Date: Tue, 1 Mar 2022 15:28:21 +0000
Subject: [PATCH] keep largest

---
 AlternativeMacros/MGEndfeetCoverage.ijm |  6 ++++--
 Macros/step8_QuantificationTool.ijm     | 16 ++++++++++++++++
 2 files changed, 20 insertions(+), 2 deletions(-)

diff --git a/AlternativeMacros/MGEndfeetCoverage.ijm b/AlternativeMacros/MGEndfeetCoverage.ijm
index 11c7ea0..0ebd405 100644
--- a/AlternativeMacros/MGEndfeetCoverage.ijm
+++ b/AlternativeMacros/MGEndfeetCoverage.ijm
@@ -84,7 +84,9 @@ for (e=0; e< filelistMGs.length; e++) {
 		saveAs("Tiff", OutDir + "MAX_" + name);
 		
 		// smoothen and enhance before segmenting with Threshold
-		run("Median...", "radius=3");
+		run("Median...", "radius=2 stack");
+
+		
 		run("Enhance Local Contrast (CLAHE)", "blocksize=127 histogram=256 maximum=3 mask=*None* fast_(less_accurate)");
 		// segmentation with Otsu Threshold
 		run("Threshold...");
@@ -95,7 +97,7 @@ for (e=0; e< filelistMGs.length; e++) {
 	
 		run("Invert", "stack"); // it measures the white as MG endfeet
 		saveAs("Tiff", OutDir + "MAX_TH_" + name);
-		run("Invert", "stack"); 
+	//	run("Invert", "stack"); 
 		
 		// for timelapse - need to iterate over the individual timepoints
 		if (frames > 1){ // if more than one timeframe it is a timelapse
diff --git a/Macros/step8_QuantificationTool.ijm b/Macros/step8_QuantificationTool.ijm
index e1f6c24..deb88e6 100644
--- a/Macros/step8_QuantificationTool.ijm
+++ b/Macros/step8_QuantificationTool.ijm
@@ -61,6 +61,7 @@ run("Set Measurements...", "area mean standard min centroid center perimeter bou
 var imgName="";
 var shortTitle = "";
 var column_label = "";
+var halfPos = 10;
 
 ///// open images from inputfolder for overall quantification, EDM, and skeletonization
 for (i=0; i< sortedFilelist.length; i++) {   
@@ -71,7 +72,17 @@ for (i=0; i< sortedFilelist.length; i++) {
 	print("processing ... " + sortedFilelist[i]);
 
 	open(path + sortedFilelist[i]);
+
+	getDimensions(width, height, channels, slices, frames);
+
+	halfPos = round(slices / 2);
+	setSlice(halfPos);
+	
+	run("Make Binary", "method=Default background=Default");
 	run("Keep Largest Region"); // from MorpholibJ to keep largest region and remove anything unconnected
+	wait(3000);
+	
+	
 	Quantification(sortedFilelist[i]); // call function quantification
 	
 	close("*");
@@ -138,6 +149,8 @@ function Quantification(title){
 	getDimensions(width, height, channels, slices, frames);
 	getPixelSize(unit,pixelWidth,pixelHeight,voxelDepth);
 	volVox = pixelWidth * pixelHeight * voxelDepth;
+	setSlice(halfPos);
+
 
 	run("Histogram", "stack");
 	// [255] is MG Vox
@@ -153,10 +166,13 @@ function Quantification(title){
 	// surface smoothing
 	run("Median 3D...", "x=6 y=6 z=6");
 	wait(3000);
+
+	setSlice(halfPos);
 	run("Make Binary", "method=Default background=Default"); // binarize after smoothing
 
 	// 3D EDM for thickness
 	run("Duplicate...", "title=For3DEDM duplicate");
+	wait(2000);
 	selectWindow("For3DEDM");
 	run("Geometry to Distance Map", "threshold=1");
 	run("Green Fire Blue"); // assign LUT green-fire-blue