// Cellcounter3D is a macro for ImageJ. It counts the red and 
// green dots on the R or G images put in a folder (nothing else 
// should be there!). 
//First the .LSM stack files should be split into single channels "Red" and "Green". 
//all the images from one channel should be in one folder
//This can be done using for instance the macro "LSM_batch_withcolors.txt".
// It prints a summary of the results,i.e. cell number per image in 
// the Log window and saves it to an output file as CSV file when its done, which then can be read in any spread-sheet like program.
// Change the minimum/maximum cell size by changing the
// values after "smallest" and "largest" (pixel) in both the
// RED and GREEN section.

// Copyright (C) 2008 Ben Gantenbein/Laszlo Kupcsik

version = getVersion();

	MonthNames = newArray("Jan","Feb","Mar","Apr","May","Jun","Jul","Aug","Sep","Oct","Nov","Dec");
	DayNames = newArray("Sun", "Mon","Tue","Wed","Thu","Fri","Sat");
	getDateAndTime(year, month, dayOfWeek, dayOfMonth, hour, minute, second, msec);
	TimeString ="Date: "+DayNames[dayOfWeek]+" ";
	if (dayOfMonth<10) {TimeString = TimeString+"0";}
	TimeString = TimeString+dayOfMonth+"-"+MonthNames[month]+"-"+year+"\nTime: ";
	if (hour<10) {TimeString = TimeString+"0";}
	TimeString = TimeString+hour+":";
	if (minute<10) {TimeString = TimeString+"0";}
	TimeString = TimeString+minute+":";
	if (second<10) {TimeString = TimeString+"0";}
	TimeString = TimeString+second;


dir1 = getDirectory("Choose Source Directory ");
list = getFileList(dir1);
print("This is imageJ version: "+version+". The images were read from "+dir1+". Counting of cells using a constant threshold. "+TimeString);
for (i=0; i<list.length; i++) {
    
    open(dir1+list[i]);
    imgname = list[i];
    number=i+1;
    status=number/list.length*100;
    
    
// GREEN*     *THIS CAN BE CHANGED TO ANY CHANNEL
     selectImage(imgname);
     run("Duplicate...", "title=green");
     run("8-bit"); // Image>Type>8-bit command
// SPECIFIY AT WHICH LEVEL YOU WOULD LIKE TO THRESHOLD THE CHANNEL (IMG/TISSUE SPECIFIC)
    setThreshold(100, 255);  run("Threshold", "thresholded remaining black");
    run("Nucleus Counter", "smallest=15 largest=300 threshold=Current smooth=None watershed");
    print(status+" %: "+number+" out of "+list.length+", lower cut-off threshold applied: 100, "+imgname+" (green), "+nResults);
    selectImage("green");
    close();
    selectImage(imgname);
    close();
    selectImage("Result of Analysis");
// SPECIFY WHERE YOU WANT TO HAVE STORED THE BINARY IMGS
    saveAs("Tiff", "ENTER PATH HERE");
    close();
    }
selectWindow("Log");
// SPECIFIY WHERE YOU WANT TO HAVE STORED THE OUTPUT LOG FILE
saveAs("Text", "ENTER PATH HERE");
print("Output written onto output file");