Re: Macro to measure particles and save ROI on to original image

Posted by gnelson on May 19, 2017; 8:47am
URL: http://imagej.273.s1.nabble.com/Macro-to-measure-particles-and-save-ROI-on-to-original-image-tp5018747p5018757.html

Hi Eloiselockyer,

I think if you write

run("Duplicate...", "title=list[i]+_original");

it should work.  No spaces, and you can use the list of the directory images as you loop through them.  

Also, you can make it easier to navigate between images if you don't split the original image nad choose the channel to duplicate.  The example code below does a similar thing and works, but more simply (please excuse my crude coding!).  Rather than split the channels, it opens a multicahnnel image, and duplicates each channel one at a time (with the duplicate channels option).  It then thresholds the duplicate then counts particles (you can see I've commented out different thresholding methods to play around with the best one).  Found particles are then added to the ROI manager and the thresholded image renamed.  


//Assumes the image is a MIP'ed tif with PML in channel1, H2AX in channel2
//(although these can be the other way round and it doesn't matter too much except for labelling in results)
//and DAPI in channel3.  It looks for pure overlap and counts respective spots.
//All results get stored in a concatenating results file.
//Plus the mask images are stored (all 3 original images converted to masks plus the coloc maskin the 4th channel)
// so you can see what it did.

var tolerance = 30;
run("Set Measurements...", "area integrated limit display redirect=None decimal=3");

dir1 = getDirectory("Choose folder with tif files ");
list = getFileList(dir1);

setBatchMode(true);

// create folders for the tifs
        dir1parent = File.getParent(dir1);
        dir1name = File.getName(dir1);
        dir2 = dir1parent+File.separator+dir1name+"_masks";
        if (File.exists(dir2)==false) {
                                File.makeDirectory(dir2); // new directory for mask images
                }

for (i=0; i<list.length; i++) {
        showProgress(i+1, list.length);
        print("processing ... "+i+1+"/"+list.length+"\n         "+list[i]);
        filename = dir1 + list[i];
        if (endsWith(filename, "tif")) {
                path=dir1+list[i];
                run("Bio-Formats Importer", "open="+filename+" autoscale color_mode=Default view=Hyperstack stack_order=XYCZT");
                originalImage = getTitle();
                roiManager("Reset");
                selectWindow(originalImage);
                run("Duplicate...", "duplicate channels=1"); //Change this to the DAPI channel slice*******
// run("8-bit");
                setAutoThreshold("Moments dark");
                run("Threshold");
                run("Watershed");
// run("Entropy Threshold");
                run("Analyze Particles...", "size=120-2500 circularity=0.04-1.00 exclude add");
                rename("nuc");
//nuclei now added to ROI manager

//next find pml bodies
                selectWindow(originalImage);
                run("Duplicate...", "title=PML duplicate channels=3");
// run("8-bit");
                setAutoThreshold("Triangle dark");
                run("Threshold");
// rename("PML");

//next find h2ax foci
                selectWindow(originalImage);
                run("Duplicate...", "title=h2ax duplicate channels=2");
                wait(1000);
// run("8-bit");
                setAutoThreshold("MaxEntropy dark");
                run("Threshold");
// rename("h2ax");

//Next find colocalised PML-H2AX foci pixels
                imageCalculator("Multiply create", "h2ax","PML");
                selectWindow("Result of h2ax");
                rename(originalImage+":coloc");
                run("Grays");
                for(j=0; j<roiManager("count"); j++) {
                        roiManager("select", j);
                        run("Find Maxima...", "noise="+tolerance+" output=[Count]");
// run("Find Maxima...", "noise="+tolerance+" output=[Point Selection]");
                        run("Add Selection...");
                        }
// saveAs("Results", dir2+File.separator+originalImage+"_nuc_coloc.xls");

//next find H2AX foci per nuc
                selectWindow("h2ax");
                rename(originalImage+":h2ax");
                for(j=0; j<roiManager("count"); j++) {
                        roiManager("select", j);
                        run("Find Maxima...", "noise="+tolerance+" output=[Count]");
// run("Find Maxima...", "noise="+tolerance+" output=[Point Selection]");
                        run("Add Selection...");
                        }
                saveAs("Results", dir2+File.separator+originalImage+"_nuc_coloc_and_h2ax.xls"); //This is savign as it goes along in case it crashes
                selectWindow(originalImage+"h2ax");
                rename("h2ax");
                selectWindow(originalImage+"coloc");
                rename("coloc");
                selectWindow(originalImage);
                close();

                run("Merge Channels...", "c1=PML c2=h2ax c3=nuc c4=coloc create");
                rename("masks");

                saveAs("TIFF", dir2+File.separator+originalImage+"_colocmasks.tif");


        close();
        }
setBatchMode(false);
}

showMessage(" Well, I think we gave them a damn good thrashing there, what what??");

// macro