diff --git a/Coloc_2.java b/Coloc_2.java
index c8a334c..1651a0b 100644
--- a/Coloc_2.java
+++ b/Coloc_2.java
@@ -1,684 +1,688 @@
import gadgets.DataContainer;
import ij.IJ;
import ij.ImagePlus;
import ij.Prefs;
import ij.WindowManager;
import ij.gui.GenericDialog;
import ij.gui.Roi;
import ij.gui.ShapeRoi;
import ij.plugin.PlugIn;
import ij.plugin.frame.RoiManager;
import ij.process.Blitter;
import ij.process.ImageProcessor;
import java.awt.Checkbox;
import java.awt.Frame;
import java.awt.Rectangle;
import java.awt.event.ItemEvent;
import java.awt.event.ItemListener;
import java.awt.event.WindowAdapter;
import java.awt.event.WindowEvent;
import java.awt.event.WindowListener;
import java.util.ArrayList;
import java.util.Arrays;
import java.util.List;
import mpicbg.imglib.container.array.ArrayContainerFactory;
import mpicbg.imglib.cursor.LocalizableByDimCursor;
import mpicbg.imglib.cursor.LocalizableCursor;
import mpicbg.imglib.cursor.special.TwinCursor;
import mpicbg.imglib.image.Image;
import mpicbg.imglib.image.ImageFactory;
import mpicbg.imglib.image.ImagePlusAdapter;
import mpicbg.imglib.type.logic.BitType;
import mpicbg.imglib.type.numeric.RealType;
import results.PDFWriter;
import results.ResultHandler;
import results.SingleWindowDisplay;
import results.Warning;
import algorithms.Algorithm;
import algorithms.AutoThresholdRegression;
import algorithms.CostesSignificanceTest;
import algorithms.Histogram2D;
import algorithms.InputCheck;
import algorithms.LiHistogram2D;
import algorithms.LiICQ;
import algorithms.MandersColocalization;
import algorithms.MissingPreconditionException;
import algorithms.PearsonsCorrelation;
/**
Copyright 2010, 2011 Daniel J. White, Tom Kazimiers, Johannes Schindelin
and the Fiji project. Fiji is just imageJ - batteries included.
This program is free software: you can redistribute it and/or modify
it under the terms of the GNU General Public License as published by
the Free Software Foundation, either version 3 of the License, or
(at your option) any later version.
This program is distributed in the hope that it will be useful,
but WITHOUT ANY WARRANTY; without even the implied warranty of
MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the
GNU General Public License for more details.
You should have received a copy of the GNU General Public License
along with this program. If not, see http://www.gnu.org/licenses/ .
*/
/**
* A plugin which does analysis colocalisation on a pair of images.
*
* @param <T>
*/
public class Coloc_2<T extends RealType<T>> implements PlugIn {
// a small bounding box container
protected class BoundingBox {
public int[] offset;
public int[] size;
public BoundingBox(int [] offset, int[] size) {
this.offset = offset.clone();
this.size = size.clone();
}
}
// a storage class for ROI information
protected class MaskInfo {
BoundingBox roi;
public Image<T> mask;
// constructors
public MaskInfo(BoundingBox roi, Image<T> mask) {
this.roi = roi;
this.mask = mask;
}
public MaskInfo() { }
}
// the storage key for Fiji preferences
protected final static String PREF_KEY = "Coloc_2.";
// Allowed types of ROI configuration
protected enum RoiConfiguration {
None,
Img1,
Img2,
Mask,
RoiManager
};
// the ROI configuration to use
protected RoiConfiguration roiConfig = RoiConfiguration.Img1;
// A list of all ROIs/masks found
protected ArrayList<MaskInfo> masks = new ArrayList<MaskInfo>();
// default indices of image, mask and ROI choices
protected static int index1 = 0;
protected static int index2 = 1;
protected static int indexMask = 0;
protected static int indexRoi = 0;
// the images to work on
protected Image<T> img1, img2;
// the channels of the images to use
protected int img1Channel = 1, img2Channel = 1;
/* The different algorithms this plug-in provides.
* If a reference is null it will not get run.
*/
protected PearsonsCorrelation<T> pearsonsCorrelation;
protected LiHistogram2D<T> liHistogramCh1;
protected LiHistogram2D<T> liHistogramCh2;
protected LiICQ<T> liICQ;
protected MandersColocalization<T> mandersCorrelation;
protected Histogram2D<T> histogram2D;
protected CostesSignificanceTest<T> costesSignificance;
// indicates if images should be printed in result
protected boolean displayImages;
// indicates if a PDF should be saved automatically
protected boolean autoSavePdf;
public void run(String arg0) {
if (showDialog()) {
try {
for (MaskInfo mi : masks) {
colocalise(img1, img2, mi.roi, mi.mask);
}
} catch (MissingPreconditionException e) {
IJ.handleException(e);
IJ.showMessage("An error occured, could not colocalize!");
return;
}
}
}
public boolean showDialog() {
// get IDs of open windows
int[] windowList = WindowManager.getIDList();
// if there are less than 2 windows open, cancel
if (windowList == null || windowList.length < 2) {
IJ.showMessage("At least 2 images must be open!");
return false;
}
/* create a new generic dialog for the
* display of various options.
*/
final GenericDialog gd
= new GenericDialog("Coloc 2");
String[] titles = new String[windowList.length];
/* the masks and ROIs array needs three more entries than
* windows to contain "none", "ROI ch 1" and "ROI ch 2"
*/
String[] roisAndMasks= new String[windowList.length + 4];
roisAndMasks[0]="<None>";
roisAndMasks[1]="ROI(s) in channel 1";
roisAndMasks[2]="ROI(s) in channel 2";
roisAndMasks[3]="ROI Manager";
// go through all open images and add them to GUI
for (int i=0; i < windowList.length; i++) {
ImagePlus imp = WindowManager.getImage(windowList[i]);
if (imp != null) {
titles[i] = imp.getTitle();
roisAndMasks[i + 4] =imp.getTitle();
} else {
titles[i] = "";
}
}
// set up the users preferences
displayImages = Prefs.get(PREF_KEY+"displayImages", false);
autoSavePdf = Prefs.get(PREF_KEY+"autoSavePdf", true);
boolean displayShuffledCostes = Prefs.get(PREF_KEY+"displayShuffledCostes", false);
boolean useLiCh1 = Prefs.get(PREF_KEY+"useLiCh1", true);
boolean useLiCh2 = Prefs.get(PREF_KEY+"useLiCh2", true);
boolean useLiICQ = Prefs.get(PREF_KEY+"useLiICQ", true);
boolean useManders = Prefs.get(PREF_KEY+"useManders", true);
boolean useScatterplot = Prefs.get(PREF_KEY+"useScatterplot", true);
boolean useCostes = Prefs.get(PREF_KEY+"useCostes", true);
int psf = (int) Prefs.get(PREF_KEY+"psf", 3);
int nrCostesRandomisations = (int) Prefs.get(PREF_KEY+"nrCostesRandomisations", 10);
/* make sure the default indices are no bigger
* than the amount of images we have
*/
index1 = clip( index1, 0, titles.length );
index2 = clip( index2, 0, titles.length );
indexMask = clip( indexMask, 0, roisAndMasks.length - 1);
gd.addChoice("Channel_1", titles, titles[index1]);
gd.addChoice("Channel_2", titles, titles[index2]);
gd.addChoice("ROI_or_mask", roisAndMasks, roisAndMasks[indexMask]);
//gd.addChoice("Use ROI", roiLabels, roiLabels[indexRoi]);
gd.addCheckbox("Show_\"Save_PDF\"_Dialog", autoSavePdf);
gd.addCheckbox("Display_Images_in_Result", displayImages);
gd.addCheckbox("Display_Shuffled_Images", displayShuffledCostes);
final Checkbox shuffleCb = (Checkbox) gd.getCheckboxes().lastElement();
// Add algorithm options
gd.addMessage("Algorithms:");
gd.addCheckbox("Li_Histogram_Channel_1", useLiCh1);
gd.addCheckbox("Li_Histogram_Channel_2", useLiCh2);
gd.addCheckbox("Li_ICQ", useLiICQ);
gd.addCheckbox("Manders'_Correlation", useManders);
gd.addCheckbox("2D_Instensity_Histogram", useScatterplot);
gd.addCheckbox("Costes'_Significance_Test", useCostes);
final Checkbox costesCb = (Checkbox) gd.getCheckboxes().lastElement();
gd.addNumericField("PSF", psf, 1);
gd.addNumericField("Costes_randomisations", nrCostesRandomisations, 0);
// disable shuffle checkbox if costes checkbox is set to "off"
shuffleCb.setEnabled(useCostes);
costesCb.addItemListener(new ItemListener() {
@Override
public void itemStateChanged(ItemEvent e) {
shuffleCb.setEnabled(costesCb.getState());
}
});
// show the dialog, finally
gd.showDialog();
// do nothing if dialog has been canceled
if (gd.wasCanceled())
return false;
ImagePlus imp1 = WindowManager.getImage(gd.getNextChoiceIndex() + 1);
ImagePlus imp2 = WindowManager.getImage(gd.getNextChoiceIndex() + 1);
// get information about the mask/ROI to use
indexMask = gd.getNextChoiceIndex();
if (indexMask == 0)
roiConfig = RoiConfiguration.None;
else if (indexMask == 1)
roiConfig = RoiConfiguration.Img1;
else if (indexMask == 2)
roiConfig = RoiConfiguration.Img2;
else if (indexMask == 3)
roiConfig = RoiConfiguration.RoiManager;
else {
roiConfig = RoiConfiguration.Mask;
/* Make indexMask the reference to the mask image to use.
* To do this we reduce it by three for the first three
* entries in the combo box.
*/
indexMask = indexMask - 4;
}
// save the ImgLib wrapped images as members
img1 = ImagePlusAdapter.wrap(imp1);
img2 = ImagePlusAdapter.wrap(imp2);
/* check if we have a valid ROI for the selected configuration
* and if so, get the ROI's bounds. Alternatively, a mask can
* be selected (that is basically all, but a rectangle).
*/
if (roiConfig == RoiConfiguration.Img1 && hasValidRoi(imp1)) {
createMasksFromImage(imp1);
} else if (roiConfig == RoiConfiguration.Img2 && hasValidRoi(imp2)) {
createMasksFromImage(imp2);
} else if (roiConfig == RoiConfiguration.RoiManager) {
- createMasksFromRoiManager(imp1.getWidth(), imp1.getHeight());
+ if (!createMasksFromRoiManager(imp1.getWidth(), imp1.getHeight()))
+ return false;
} else if (roiConfig == RoiConfiguration.Mask) {
// get the image to be used as mask
ImagePlus maskImp = WindowManager.getImage(windowList[indexMask]);
Image<T> maskImg = ImagePlusAdapter.<T>wrap( maskImp );
// get a valid mask info for the image
MaskInfo mi = getBoundingBoxOfMask(maskImg);
masks.add( mi ) ;
} else {
/* if no ROI/mask is selected, just add an empty MaskInfo
* to colocalise both images without constraints.
*/
masks.add(new MaskInfo(null, null));
}
// read out GUI data
autoSavePdf = gd.getNextBoolean();
displayImages = gd.getNextBoolean();
displayShuffledCostes = gd.getNextBoolean();
useLiCh1 = gd.getNextBoolean();
useLiCh2 = gd.getNextBoolean();
useLiICQ = gd.getNextBoolean();
useManders = gd.getNextBoolean();
useScatterplot = gd.getNextBoolean();
useCostes = gd.getNextBoolean();
psf = (int) gd.getNextNumber();
nrCostesRandomisations = (int) gd.getNextNumber();
// save user preferences
Prefs.set(PREF_KEY+"autoSavePdf", autoSavePdf);
Prefs.set(PREF_KEY+"displayImages", displayImages);
Prefs.set(PREF_KEY+"displayShuffledCostes", displayShuffledCostes);
Prefs.set(PREF_KEY+"useLiCh1", useLiCh1);
Prefs.set(PREF_KEY+"useLiCh2", useLiCh2);
Prefs.set(PREF_KEY+"useLiICQ", useLiICQ);
Prefs.set(PREF_KEY+"useManders", useManders);
Prefs.set(PREF_KEY+"useScatterplot", useScatterplot);
Prefs.set(PREF_KEY+"useCostes", useCostes);
Prefs.set(PREF_KEY+"psf", psf);
Prefs.set(PREF_KEY+"nrCostesRandomisations", nrCostesRandomisations);
// Parse algorithm options
pearsonsCorrelation = new PearsonsCorrelation<T>(PearsonsCorrelation.Implementation.Fast);
if (useLiCh1)
liHistogramCh1 = new LiHistogram2D<T>("Li - Ch1", true);
if (useLiCh2)
liHistogramCh2 = new LiHistogram2D<T>("Li - Ch2", false);
if (useLiICQ)
liICQ = new LiICQ<T>();
if (useManders)
mandersCorrelation = new MandersColocalization<T>();
if (useScatterplot)
histogram2D = new Histogram2D<T>("2D intensity histogram");
if (useCostes) {
costesSignificance = new CostesSignificanceTest<T>(pearsonsCorrelation,
psf, nrCostesRandomisations, displayShuffledCostes);
}
return true;
}
/**
* Call this method to run a whole colocalisation configuration,
* all selected algorithms get run on the supplied images. You
* can specify the data further by supplying appropriate
* information in the mask structure.
*
* @param img1
* @param img2
* @param roi
* @param mask
* @param maskBB
* @throws MissingPreconditionException
*/
public void colocalise(Image<T> img1, Image<T> img2, BoundingBox roi,
Image<T> mask) throws MissingPreconditionException {
// create a new container for the selected images and channels
DataContainer<T> container;
if (mask != null) {
container = new DataContainer<T>(img1, img2,
img1Channel, img2Channel, mask, roi.offset, roi.size);
} else if (roi != null) {
// we have no mask, but a regular ROI in use
container = new DataContainer<T>(img1, img2,
img1Channel, img2Channel, roi.offset, roi.size);
} else {
// no mask and no ROI is present
container = new DataContainer<T>(img1, img2,
img1Channel, img2Channel);
}
// create a results handler
final List<ResultHandler<T>> listOfResultHandlers = new ArrayList<ResultHandler<T>>();
final PDFWriter<T> pdfWriter = new PDFWriter<T>(container);
final SingleWindowDisplay<T> swDisplay = new SingleWindowDisplay<T>(container, pdfWriter);
listOfResultHandlers.add(swDisplay);
listOfResultHandlers.add(pdfWriter);
//ResultHandler<T> resultHandler = new EasyDisplay<T>(container);
// this list contains the algorithms that will be run when the user clicks ok
List<Algorithm<T>> userSelectedJobs = new ArrayList<Algorithm<T>>();
// add some pre-processing jobs:
userSelectedJobs.add( container.setInputCheck(
new InputCheck<T>()) );
userSelectedJobs.add( container.setAutoThreshold(
new AutoThresholdRegression<T>(pearsonsCorrelation)) );
// add user selected algorithms
addIfValid(pearsonsCorrelation, userSelectedJobs);
addIfValid(liHistogramCh1, userSelectedJobs);
addIfValid(liHistogramCh2, userSelectedJobs);
addIfValid(liICQ, userSelectedJobs);
addIfValid(mandersCorrelation, userSelectedJobs);
addIfValid(histogram2D, userSelectedJobs);
addIfValid(costesSignificance, userSelectedJobs);
// execute all algorithms
int count = 0;
int jobs = userSelectedJobs.size();
for (Algorithm<T> a : userSelectedJobs){
try {
count++;
IJ.showStatus(count + "/" + jobs + ": Running " + a.getName());
a.execute(container);
}
catch (MissingPreconditionException e){
for (ResultHandler<T> r : listOfResultHandlers){
r.handleWarning(
new Warning( "Probem with input data", a.getName() + ": " + e.getMessage() ) );
}
}
}
// clear status
IJ.showStatus("");
// let the algorithms feed their results to the handler
for (Algorithm<T> a : userSelectedJobs){
for (ResultHandler<T> r : listOfResultHandlers)
a.processResults(r);
}
// if we have ROIs/masks, add them to results
if (displayImages) {
Image<T> channel1, channel2;
if (mask != null || roi != null) {
int[] offset = container.getMaskBBOffset();
int[] size = container.getMaskBBSize();
channel1 = createMaskImage( container.getSourceImage1(),
container.getMask(), offset, size, "Channel 1" );
channel2 = createMaskImage( container.getSourceImage2(),
container.getMask(), offset, size, "Channel 2" );
} else {
channel1 = container.getSourceImage1();
channel2 = container.getSourceImage2();
channel1.setName("Channel 1");
channel2.setName("Channel 2");
}
for (ResultHandler<T> r : listOfResultHandlers) {
r.handleImage (channel1);
r.handleImage (channel2);
}
}
// do the actual results processing
swDisplay.process();
// add window to the IJ window manager
swDisplay.addWindowListener(new WindowAdapter() {
@Override
public void windowClosed(WindowEvent e) {
WindowManager.removeWindow((Frame) swDisplay);
}
});
WindowManager.addWindow(swDisplay);
// show PDF saving dialog if requested
if (autoSavePdf)
pdfWriter.process();
}
/**
* A method to get the bounding box from the data in the given
* image that is above zero. Those values are interpreted as a
* mask. It will return null if no mask information was found.
*
* @param mask The image to look for "on" values in
* @return a new MaskInfo object or null
*/
protected MaskInfo getBoundingBoxOfMask(Image<T> mask) {
LocalizableCursor<T> cursor = mask.createLocalizableCursor();
int numMaskDims = mask.getNumDimensions();
// the "off type" of the mask
T offType = mask.createType();
offType.setZero();
// the corners of the bounding box
int[] min = null;
int[] max = null;
// indicates if mask data has been found
boolean maskFound = false;
// a container for temporary position information
int[] pos = new int[numMaskDims];
// walk over the mask
while (cursor.hasNext() ) {
cursor.fwd();
T data = cursor.getType();
// test if the current mask data represents on or off
if (data.compareTo(offType) > 0) {
// get current position
cursor.getPosition(pos);
if (!maskFound) {
// we found mask data, first time
maskFound = true;
// init min and max with the current position
min = Arrays.copyOf(pos, numMaskDims);
max = Arrays.copyOf(pos, numMaskDims);
} else {
/* Is is at least second hit, compare if it
* has new "extreme" positions, i.e. does
* is make the BB bigger?
*/
for (int d=0; d<numMaskDims; d++) {
if (pos[d] < min[d]) {
// is it smaller than min
min[d] = pos[d];
} else if (pos[d] > max[d]) {
// is it larger than max
max[d] = pos[d];
}
}
}
}
}
cursor.close();
if (!maskFound) {
return null;
} else {
// calculate size
int[] size = new int[numMaskDims];
for (int d=0; d<numMaskDims; d++)
size[d] = max[d] - min[d] + 1;
// create and add bounding box
BoundingBox bb = new BoundingBox(min, size);
return new MaskInfo(bb, mask);
}
}
/**
* Adds the provided Algorithm to the list if it is not null.
*/
protected void addIfValid(Algorithm<T> a, List<Algorithm<T>> list) {
if (a != null)
list.add(a);
}
/**
* Returns true if a custom ROI has been selected, i.e if the current
* ROI does not have the extent of the whole image.
* @return true if custom ROI selected, false otherwise
*/
protected boolean hasValidRoi(ImagePlus imp) {
Roi roi = imp.getRoi();
if (roi == null)
return false;
Rectangle theROI = roi.getBounds();
// if the ROI is the same size as the image (default ROI), return false
return (theROI.height != imp.getHeight()
|| theROI.width != imp.getWidth());
}
/**
* Clips a value to the specified bounds.
*/
protected static int clip(int val, int min, int max) {
return Math.max( Math.min( val, max ), min );
}
/**
* This method checks if the given ImagePlus contains any
* masks or ROIs. If so, the appropriate date structures
* are created and filled.
*/
protected void createMasksFromImage(ImagePlus imp) {
// get ROIs from current image in Fiji
Roi[] impRois = split(imp.getRoi());
// create the ROIs
createMasksAndRois(impRois, imp.getWidth(), imp.getHeight());
}
/**
* A method to fill the masks array with data based on the ROI manager.
*/
- protected void createMasksFromRoiManager(int width, int height) {
+ protected boolean createMasksFromRoiManager(int width, int height) {
RoiManager roiManager = RoiManager.getInstance();
- if (roiManager == null)
+ if (roiManager == null) {
IJ.error("Could not get ROI Manager instance.");
+ return false;
+ }
Roi[] selectedRois = roiManager.getSelectedRoisAsArray();
// create the ROIs
createMasksAndRois(selectedRois, width, height);
+ return true;
}
/**
* Creates appropriate data structures from the ROI information
* passed. If an irregular ROI is found, it will be put into a
* frame of its bounding box size and put into an Image<T>.
*
* In the end the members ROIs, masks and maskBBs will be
* filled if ROIs or masks were found. They will be null
* otherwise.
*/
protected void createMasksAndRois(Roi[] rois, int width, int height) {
// create empty list
masks.clear();
for (Roi r : rois ){
MaskInfo mi = new MaskInfo();
// add it to the list of masks/ROIs
masks.add(mi);
// get the ROIs/masks bounding box
Rectangle rect = r.getBounds();
mi.roi = new BoundingBox(
new int[] {rect.x, rect.y} ,
new int[] {rect.width, rect.height});
ImageProcessor ipMask = r.getMask();
// check if we got a regular ROI and return if so
if (ipMask == null) {
continue;
}
// create a mask processor of the same size as a slice
ImageProcessor ipSlice = ipMask.createProcessor(width, height);
// fill the new slice with black
ipSlice.setValue(0.0);
ipSlice.fill();
// position the mask on the new mask processor
ipSlice.copyBits(ipMask, mi.roi.offset[0], mi.roi.offset[1], Blitter.COPY);
// create an Image<T> out of it
ImagePlus maskImp = new ImagePlus("Mask", ipSlice);
// and remember it and the masks bounding box
mi.mask = ImagePlusAdapter.<T>wrap( maskImp );
}
}
/**
* This method duplicates the given images, but respects
* ROIs if present. Meaning, a sub-picture will be created when
* source images are ROI/MaskImages.
* @throws MissingPreconditionException
*/
protected Image<T> createMaskImage(Image<T> image, Image<BitType> mask,
int[] offset, int[] size, String name) throws MissingPreconditionException {
int[] pos = image.createPositionArray();
// sanity check
if (pos.length != offset.length || pos.length != size.length) {
throw new MissingPreconditionException("Mask offset and size must be of same dimensionality like image.");
}
// use twin cursor for only one image
TwinCursor<T> cursor = new TwinCursor<T>(
image.createLocalizableByDimCursor(),
image.createLocalizableByDimCursor(),
mask.createLocalizableCursor());
// prepare output image
ImageFactory<T> maskFactory = new ImageFactory<T>(
image.createType(), new ArrayContainerFactory());
Image<T> maskImage = maskFactory.createImage( size, name );
LocalizableByDimCursor<T> maskCursor =
maskImage.createLocalizableByDimCursor();
// go through the visible data and copy it to the output
while (cursor.hasNext()) {
cursor.fwd();
cursor.getPosition(pos);
// shift coordinates by offset
for (int i=0; i < pos.length; ++i) {
pos[i] = pos[i] - offset[i];
}
// write out to correct position
maskCursor.setPosition( pos );
maskCursor.getType().set( cursor.getChannel1() );
}
cursor.close();
maskCursor.close();
return maskImage;
}
/**
* Splits a non overlapping composite ROI into its sub ROIs.
*
* @param roi The ROI to split
* @return A list of one or more ROIs
*/
public static Roi[] split(Roi roi) {
if (roi instanceof ShapeRoi)
return ((ShapeRoi)roi).getRois();
return new Roi[] { roi };
}
}