class MandersPlugin(ImageListener, WindowAdapter):
def __init__(self):
self.imp = None
self.preview = None
self.createMainWindow()
self.cells = None
self.files = []
self.results = ResultsTable()
ImagePlus.addImageListener(self)
self.selectInputDir()
self.selectOutputDir()
self.pairs = []
self.methods = []
self.processNextFile()
def selectInputDir(self):
inputDialog = DirectoryChooser("Please select a directory contaning your images")
inputDir = inputDialog.getDirectory()
for imageFile in os.listdir(inputDir):
self.files.append(inputDir + imageFile)
def selectOutputDir(self):
outputDialog = DirectoryChooser("Please select a directory to save your results")
self.outputDir = outputDialog.getDirectory()
def closeImage(self):
if self.imp is not None:
self.imp.close()
self.imp = None
if self.preview is not None:
self.preview.close()
self.preview = None
def openImage(self, imageFile):
try:
images = BF.openImagePlus(imageFile)
self.imp = images[0]
except UnknownFormatException:
return None
if self.imp.getNChannels() < 2:
IJ.error("Bad image format", "Image must contain at lease 2 channels!")
return None
if not self.pairs or \
not self.methods:
self.getOptionsDialog(self.imp)
title = self.imp.title
self.imp.title = title[:title.rfind('.')]
return self.imp
def getOptionsDialog(self, imp):
thr_methods = ["None", "Default", "Huang", "Intermodes", "IsoData", "Li", "MaxEntropy","Mean", "MinError(I)", "Minimum", "Moments", "Otsu", "Percentile", "RenyiEntropy", "Shanbhag" , "Triangle", "Yen"]
gd = GenericDialog("Please select channels to collocalize")
for i in range(1, imp.getNChannels() + 1):
gd.addChoice("Threshold method for channel %i" % i, thr_methods, "None")
gd.showDialog()
if gd.wasCanceled():
self.exit()
channels = []
for i in range(1, imp.getNChannels() + 1):
method = gd.getNextChoice()
self.methods.append(method)
if method != "None":
channels.append(i)
for x in channels:
for y in channels:
if x < y:
self.pairs.append((x, y))
def processNextFile(self):
if self.files:
imageFile = self.files.pop(0)
return self.processFile(imageFile)
else:
return False
def processFile(self, imageFile):
imp = self.openImage(imageFile)
if imp is not None:
cell = Cell(imp.NSlices, 1)
self.cells = DelegateListModel([])
self.cells.append(cell)
self.showMainWindow(self.cells)
if self.checkbox3D.isSelected():
self.displayImage(imp)
else:
self.displayImage(imp, False)
self.preview = self.previewImage(imp)
self.displayImage(self.preview)
return True
else:
return self.processNextFile()
def displayImage(self, imp, show = True):
imp.setDisplayMode(IJ.COMPOSITE)
enhancer = ContrastEnhancer()
enhancer.setUseStackHistogram(True)
splitter = ChannelSplitter()
for c in range(1, imp.getNChannels() + 1):
imp.c = c
enhancer.stretchHistogram(imp, 0.35)
if show:
imp.show()
def previewImage(self, imp):
roi = imp.getRoi()
splitter = ChannelSplitter()
channels = []
for c in range(1, imp.getNChannels() + 1):
channel = ImagePlus("Channel %i" % c, splitter.getChannel(imp, c))
projector = ZProjector(channel)
projector.setMethod(ZProjector.MAX_METHOD)
projector.doProjection()
channels.append(projector.getProjection())
image = RGBStackMerge.mergeChannels(channels, False)
image.title = imp.title + " MAX Intensity"
image.luts = imp.luts
imp.setRoi(roi)
return image
def getCroppedChannels(self, imp, cell):
splitter = ChannelSplitter()
imp.setRoi(None)
if cell.mode3D:
cropRoi = cell.getCropRoi()
else:
cropRoi = cell.roi
if cropRoi is None:
return None
crop = cropRoi.getBounds()
channels = []
for c in range(1, imp.getNChannels() + 1):
slices = ImageStack(crop.width, crop.height)
channel = splitter.getChannel(imp, c)
for z in range(1, channel.getSize() + 1):
zslice = channel.getProcessor(z)
zslice.setRoi(cropRoi)
nslice = zslice.crop()
if cell.mode3D:
oroi = cell.slices[z - 1].roi
else:
oroi = cell.roi
if oroi is not None:
roi = oroi.clone()
bounds = roi.getBounds()
roi.setLocation(bounds.x - crop.x, bounds.y - crop.y)
nslice.setColor(Color.black)
nslice.fillOutside(roi)
slices.addSlice(nslice)
channels.append(ImagePlus("Channel %i" % c, slices))
return channels
def getThreshold(self, imp, method):
thresholder = Auto_Threshold()
duplicator = Duplicator()
tmp = duplicator.run(imp)
return thresholder.exec(tmp, method, False, False, True, False, False, True)
def getContainer(self, impA, impB):
imgA = ImagePlusAdapter.wrap(impA)
imgB = ImagePlusAdapter.wrap(impB)
return DataContainer(imgA, imgB, 1, 1, "imageA", "imageB")
def getManders(self, imp, cell):
### Crop channels according to cell mask
channels = self.getCroppedChannels(imp, cell)
if channels is None:
return None
### Calculate channel thresholds
thrs = []
thrimps = []
for c, method in enumerate(self.methods):
if method != "None":
thr, thrimp = self.getThreshold(channels[c], method)
else:
thr, thrimp = None, None
thrs.append(thr)
thrimps.append(thrimp)
### Calculate manders colocalization
manders = MandersColocalization()
raws = []
thrds = []
for chA, chB in self.pairs:
container = self.getContainer(channels[chA - 1], channels[chB - 1])
img1 = container.getSourceImage1()
img2 = container.getSourceImage2()
mask = container.getMask()
cursor = TwinCursor(img1.randomAccess(), img2.randomAccess(), Views.iterable(mask).localizingCursor())
rtype = img1.randomAccess().get().createVariable()
raw = manders.calculateMandersCorrelation(cursor, rtype)
rthr1 = rtype.copy()
rthr2 = rtype.copy()
rthr1.set(thrs[chA - 1])
rthr2.set(thrs[chB - 1])
cursor.reset()
thrd = manders.calculateMandersCorrelation(cursor, rthr1, rthr2, ThresholdMode.Above)
raws.append(raw)
thrds.append(thrd)
return (channels, thrimps, thrs, raws, thrds)
def saveMultichannelImage(self, title, channels, luts):
tmp = RGBStackMerge.mergeChannels(channels, False)
tmp.luts = luts
saver = FileSaver(tmp)
saver.saveAsTiffStack(self.outputDir + title + ".tif")
tmp.close()
def createMainWindow(self):
self.frame = JFrame('Select cells and ROIs',
defaultCloseOperation = JFrame.DISPOSE_ON_CLOSE
)
self.frame.setLayout(GridBagLayout())
self.frame.addWindowListener(self)
self.frame.add(JLabel("Cells"),
GridBagConstraints(0, 0, 1, 1, 0, 0,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(5, 2, 2, 0), 0, 0
))
self.cellList = JList(DelegateListModel([]),
selectionMode = ListSelectionModel.SINGLE_SELECTION,
cellRenderer = MyRenderer(),
selectedIndex = 0,
valueChanged = self.selectCell
)
self.frame.add(JScrollPane(self.cellList),
GridBagConstraints(0, 1, 1, 5, .5, 1,
GridBagConstraints.CENTER, GridBagConstraints.BOTH,
Insets(0, 2, 2, 0), 0, 0
))
self.frame.add(JButton('Add cell', actionPerformed = self.addCell),
GridBagConstraints(1, 2, 1, 2, 0, .25,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(0, 0, 0, 0), 0, 0
))
self.frame.add(JButton('Remove cell', actionPerformed = self.removeCell),
GridBagConstraints(1, 4, 1, 2, 0, .25,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(0, 5, 0, 5), 0, 0
))
self.frame.add(JLabel("Slices"),
GridBagConstraints(0, 6, 1, 1, 0, 0,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(5, 2, 2, 0), 0, 0
))
self.sliceList = JList(DelegateListModel([]),
selectionMode = ListSelectionModel.SINGLE_SELECTION,
cellRenderer = MyRenderer(),
selectedIndex = 0,
valueChanged = self.selectSlice
)
self.frame.add(JScrollPane(self.sliceList),
GridBagConstraints(0, 7, 1, 5, .5, 1,
GridBagConstraints.CENTER, GridBagConstraints.BOTH,
Insets(0, 2, 2, 0), 0, 0
))
self.frame.add(JButton('Update ROI', actionPerformed = self.updateSlice),
GridBagConstraints(1, 8, 1, 2, 0, .25,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(0, 0, 0, 0), 0, 0
))
self.frame.add(JButton('Done', actionPerformed = self.doneSelecting),
GridBagConstraints(1, 10, 1, 2, 0, .25,
GridBagConstraints.CENTER, GridBagConstraints.NONE,
Insets(0, 0, 0, 0), 0, 0
))
self.checkbox3D = JCheckBox('3D selection mode', True, actionPerformed=self.toggle3D)
self.frame.add(self.checkbox3D,
GridBagConstraints(0, 13, 2, 1, 0, 1,
GridBagConstraints.WEST, GridBagConstraints.NONE,
Insets(0, 0, 0, 0), 0, 0
))
def showMainWindow(self, cells = None):
if cells is not None:
self.cellList.model = cells
if cells:
self.cellList.selectedIndex = 0
self.frame.pack()
self.frame.visible = True
def hideMainWindow(self):
self.frame.visible = False
def closeMainWindow(self):
self.frame.dispose()
def toggle3D(self, event):
mode3D = self.checkbox3D.isSelected()
if mode3D:
self.sliceList.enabled = True
if self.imp is not None:
self.imp.show()
if self.preview is not None:
self.preview.hide()
else:
self.sliceList.enabled = False
if self.preview is None:
self.preview = self.previewImage(self.imp)
self.displayImage(self.preview)
else:
self.preview.show()
if self.imp is not None:
self.imp.hide()
selectedCell = self.cellList.selectedIndex
if selectedCell >= 0:
cell = self.cells[selectedCell]
self.sliceList.model = cell.slices
self.sliceList.selectedIndex = 0
def addCell(self, event):
size = len(self.cells)
if (size > 0):
last = self.cells[size - 1]
n = last.n + 1
else:
n = 1
self.cells.append(Cell(self.imp.NSlices, n))
self.cellList.selectedIndex = size
def removeCell(self, event):
selected = self.cellList.selectedIndex
if selected >= 0:
self.cells.remove(self.cells[selected])
if (selected >= 1):
self.cellList.selectedIndex = selected - 1
else:
self.cellList.selectedIndex = 0
def selectCell(self, event):
selected = self.cellList.selectedIndex
if selected >= 0:
cell = self.cells[selected]
self.sliceList.model = cell.slices
self.sliceList.selectedIndex = 0
else:
self.sliceList.model = DelegateListModel([])
if self.preview is not None:
self.preview.setRoi(cell.roi)
def selectSlice(self, event):
selectedCell = self.cellList.selectedIndex
selectedSlice = self.sliceList.selectedIndex
if selectedCell >= 0 and selectedSlice >= 0:
cell = self.cells[selectedCell]
image = self.imp
mode3D = self.checkbox3D.isSelected()
if image is not None and cell is not None and mode3D:
roi = cell.slices[selectedSlice].roi
if (image.z - 1 != selectedSlice):
image.z = selectedSlice + 1
image.setRoi(roi, True)
if self.preview is not None and not mode3D:
self.preview.setRoi(cell.roi, True)
def updateSlice(self, event):
if self.checkbox3D.isSelected():
self.updateSlice3D(self.imp)
else:
self.updateSlice2D(self.preview)
def updateSlice3D(self, imp):
selectedCell = self.cellList.selectedIndex
selectedSlice = self.sliceList.selectedIndex
if selectedCell >= 0 and selectedSlice >= 0 and imp is not None:
cell = self.cells[selectedCell]
impRoi = imp.getRoi()
if cell is not None and impRoi is not None:
index = selectedSlice + 1
roi = ShapeRoi(impRoi, position = index)
cell.mode3D = True
cell.name = "Cell %i (3D)" % cell.n
cell.slices[selectedSlice].roi = roi
if (index + 1 <= len(cell.slices)):
imp.z = index + 1
self.cellList.repaint(self.cellList.getCellBounds(selectedCell, selectedCell))
self.sliceList.repaint(self.sliceList.getCellBounds(selectedSlice, selectedSlice))
def updateSlice2D(self, imp):
selectedCell = self.cellList.selectedIndex
if selectedCell >= 0 and imp is not None:
cell = self.cells[selectedCell]
impRoi = imp.getRoi()
if cell is not None and impRoi is not None:
roi = ShapeRoi(impRoi, position = 1)
cell.mode3D = False
cell.name = "Cell %i (2D)" % cell.n
cell.roi = roi
self.cellList.repaint(self.cellList.getCellBounds(selectedCell, selectedCell))
def imageOpened(self, imp):
pass
def imageClosed(self, imp):
pass
def imageUpdated(self, imp):
if self.checkbox3D.isSelected():
if imp is not None:
selectedCell = self.cellList.selectedIndex
selectedSlice = imp.z - 1
if imp == self.imp and selectedSlice != self.sliceList.selectedIndex:
self.sliceList.selectedIndex = selectedSlice
def doneSelecting(self, event):
oluts = self.imp.luts
luts = []
channels = []
for c, method in enumerate(self.methods):
if method != "None":
luts.append(oluts[c])
channels.append(c)
for cell in self.cells:
manders = self.getManders(self.imp, cell)
if manders is not None:
chimps, thrimps, thrs, raws, thrds = manders
index = self.cells.index(cell) + 1
title = "Cell_%i-" % index + self.imp.title
self.saveMultichannelImage(title, chimps, oluts)
title = "Cell_%i_thrd-" % index + self.imp.title
self.saveMultichannelImage(title, thrimps, luts)
self.results.incrementCounter()
row = self.results.getCounter() - 1
for i, thr in enumerate(thrs):
if thr is not None:
self.results.setValue("Threshold %i" % (i + 1), row, int(thr))
for i, pair in enumerate(self.pairs):
self.results.setValue("%i-%i M1 raw" % pair, row, float(raws[i].m1))
self.results.setValue("%i-%i M2 raw" % pair, row, float(raws[i].m2))
self.results.setValue("%i-%i M1 thrd" % pair, row, float(thrds[i].m1))
self.results.setValue("%i-%i M2 thrd" % pair, row, float(thrds[i].m2))
self.closeImage()
if not self.processNextFile():
print "All done - happy analysis!"
self.results.show("Manders collocalization results")
self.exit()
def windowClosing(self, e):
print "Closing plugin - BYE!!!"
self.exit()
def exit(self):
ImagePlus.removeImageListener(self)
self.closeImage()
self.closeMainWindow()
