path="Z:\\Desktop\\Results\\Results 2010\\Niklas\\Image J analysis\\";
print(path);
sfmin=2;sfmax=10;lfmin=11;lfmax=6000;
Dialog.create("Number of images");
  Dialog.addString("Path:", path);
 Dialog.addNumber("images to be proccessed:", 10);
 Dialog.addNumber("Small foci min value in pixel:", 8);
 Dialog.addNumber("Small foci max value in pixel:", 75);
 Dialog.addNumber("Large foci min value in pixel:", 76);
 Dialog.addNumber("Large foci max value in pixel:", 500);
Dialog.addNumber("Small threshold :", 60);
Dialog.addNumber("Large threshold :", 60);
Dialog.show();
path= Dialog.getString();
nrbilder=Dialog.getNumber();
sfmin=Dialog.getNumber();
sfmax=Dialog.getNumber();
lfmin=Dialog.getNumber();
lfmax=Dialog.getNumber();
thsmall=Dialog.getNumber();
thlarge=Dialog.getNumber();
print(sfmin+": "+sfmax+": "+lfmin+": "+lfmax+": "+thsmall+": "+thlarge);
for (p=1; p<nrbilder+1; p++)    {
	open(path+p+".tif");
	run("RGB Stack");

	setSlice(3);
	run("Add Slice");
	run("Add Slice");
	run("Add Slice");
	run("Convert Stack to Images");
	selectWindow("Red");
	setThreshold(20, 255);
	run("Analyze Particles...", "size=200-200000 circularity=0.0-1.00 show=Outlines display exclude clear include record add slice");
	cellnumber = nResults;
	DNAarea=newArray(cellnumber);
	DNAmean=newArray(cellnumber);
	SmallFociMean=newArray(cellnumber);
	SmallFociSD=newArray(cellnumber);
	LargeFociMean=newArray(cellnumber);
	LargeFociSD=newArray(cellnumber);
	SmallFociArea=newArray(cellnumber);
	SmallFociIntensity=newArray(cellnumber);
	LargeFociArea=newArray(cellnumber);
	FOCIcolocarea=newArray(cellnumber);
	LargeFociIntensity=newArray(cellnumber);
	SmallFoci=newArray(cellnumber);
	LargeFoci=newArray(cellnumber);
	FOCIcoloc=newArray(cellnumber);
	selectWindow("Drawing of Red");
	run("Select All");
	run("Copy");
	selectWindow(p+"-0006");
	setPasteMode("Copy");
	run("Paste");
	run("Invert");
	run("Make Binary");
	run("Subtract...", "value=3");
	selectWindow("Drawing of Red");
	close();

	selectWindow("Green");
	run("Copy");
	selectWindow(p+"-0004");
	run("Paste");
            run("Subtract Background...", "rolling=5 sliding slice");
	run("Unsharp Mask...", "radius=4 mask=0.70 slice");
	setThreshold(thsmall, 255);
	run("Analyze Particles...", "size="+sfmin+"-"+sfmax+" circularity=0.0-1.00 show=Masks display slice");
	run("Select All");
	run("Copy");
	selectWindow(p+"-0004");
	run("Paste");
	run("Subtract...", "value=150");
	selectWindow("Mask of "+p+"-0004");
	close();

	selectWindow("Green");
	run("Copy");
	selectWindow(p+"-0005");
	run("Paste");
	 run("Subtract Background...", "rolling=5 sliding slice");
	run("Unsharp Mask...", "radius=4 mask=0.70 slice");
	setThreshold(thlarge, 255);
	run("Analyze Particles...", "size="+lfmin+"-"+lfmax+"circularity=0.0-1.00 show=Masks display slice");
	run("Select All");
	run("Copy");
	selectWindow(p+"-0005");
	run("Paste");
	run("Subtract...", "value=100");
	selectWindow("Mask of "+p+"-0005");
	close();

	run("Convert Images to Stack");
	setSlice(6);
	run("Add Slice");
	setSlice(4);
	run("Copy");
	setSlice(7);
	setPasteMode("Copy");
	run("Paste");
	setSlice(5);
	run("Copy");
	setSlice(7);
	setPasteMode("Add");
	run("Paste");
	run("Clear Results");
	for (i=0; i<cellnumber; i++)    {

		run("Set Measurements...", "area mean standard slice redirect=None decimal=1");
		roiManager("Select", i);
		setSlice(3);
		run("Measure");
		DNAarea[i]=getResult("Area");
		DNAmean[i]=getResult("Mean");
		setSlice(1);
		run("Measure");
		SmallFociMean[i]=getResult("Mean");
		SmallFociSD[i]=getResult("StdDev");
		setSlice(2);
		run("Measure");
		LargeFociMean[i]=getResult("Mean");
		LargeFociSD[i]=getResult("StdDev");
		setSlice(4);
		setThreshold(50, 255);
		run("Clear Results");
		run("Analyze Particles...", "size="+sfmin+"-"+sfmax+"circularity=0.0-1.00 show=Nothing display slice");
		SmallFoci[i]=nResults;
		fa=0;
		fI=0;
		for (j=0; j<nResults; j++)  {
			fa=getResult("Area",j)+fa;
			fI=getResult("Mean",j)*getResult("Area",j)+fI;
			}
		SmallFociArea[i]=fa;
		SmallFociIntensity[i]=fI;
		run("Clear Results");
		roiManager("Select", i);
		setSlice(5);
		setThreshold(50, 255);
		run("Analyze Particles...", "size="+lfmin+"-"+lfmax+"circularity=0.0-1.00 show=Nothing display slice");
		LargeFoci[i]=nResults;
		fa=0;
		fI=0;
		for (j=0; j<nResults; j++)  {
			fa=getResult("Area",j)+fa;
			fI=getResult("Mean",j)*getResult("Area",j)+fI;
			}
		LargeFociArea[i]=fa;
		LargeFociIntensity[i]=fI;

		run("Clear Results");
		roiManager("Select", i);
		setSlice(7);
		setThreshold(254, 255);
		run("Analyze Particles...", "size="+2+"-"+500+"circularity=0.0-1.00 show=Nothing display slice");
		nrcoloc=0;
		fc=0;
		fa=0;
		for (j=0; j<nResults; j++)  {
			if (getResult("Mean",j)>155)  {
				nrcoloc=nrcoloc+1;
				fa=getResult("Area",j);
				fi=getResult("Mean",j);
				fc=fc+(fa*fi-fa*155)/100;
				}
			}
		FOCIcolocarea[i]=fc;
		FOCIcoloc[i]=nrcoloc;
     	    }


for (i=0; i<cellnumber; i++) {
	j=i+1;
	 print(p+"	"+j+"	 "+DNAarea[i]+"	"+DNAmean[i]+"	"+SmallFociMean[i]+"	"+SmallFociSD[i]+"	"+LargeFociMean[i]+"	"+LargeFociSD[i]+"	"+SmallFociArea[i]+"	"+SmallFociIntensity[i]+"	"+SmallFoci[i]+"	"+LargeFociArea[i]+"	"+LargeFociIntensity[i]+"	"+LargeFoci[i]+"	"+FOCIcolocarea[i]+"	"+FOCIcoloc[i]);
	}
setSlice(1);
run("Subtract...", "slice value=4");
setSlice(2);
run("Subtract...", "slice value=4");
run("Copy");
setSlice(3);
setPasteMode("Copy");
run("Paste");
setSlice(4);
run("Multiply...", "slice value=5");
run("Subtract...", "slice value=2");
run("Copy");
setSlice(7);
setPasteMode("Copy");
run("Paste");

setSlice(5);
run("Multiply...", "slice value=5");
run("Subtract...", "slice value=1");
run("Copy");
setSlice(4);
setPasteMode("AND");
run("Paste");
run("Multiply...", "slice value=5");

setSlice(5);
run("Copy");
setSlice(7);
setPasteMode("XOR");
run("Paste");
setSlice(4);
run("Copy");
setSlice(7);
setPasteMode("Max");
run("Paste");
run("Copy");
setSlice(6);
setPasteMode("OR");
run("Paste");
run("Copy");
setSlice(2);
setPasteMode("Copy");
run("Paste");
setSlice(4);
run("Delete Slice");
setSlice(4);
run("Delete Slice");
setSlice(4);
run("Delete Slice");
setSlice(4);
run("Delete Slice");
run("Coloc");
saveAs("Tiff", path+p+"stack");


close();

}