def segmentNuc(impc2): impdup = Duplicator().run(impc2) IJ.run(impdup, "8-bit", "") IJ.run(impdup, "Gaussian Blur...", "sigma=1.5 stack") # AutoThresholder().getThreshold(AutoThresholder.Method.valuOf('Otsu'), int[] histogram) IJ.setAutoThreshold(impdup, "Otsu dark") IJ.run(impdup, "Convert to Mask", "stack") #IJ.setAutoThreshold(impdup, "Otsu dark") #opt = PA.SHOW_MASKS + PA.SHOW_RESULTS + PA.EXCLUDE_EDGE_PARTICLES + PA.INCLUDE_HOLES # option for stack missing opt = PA.SHOW_MASKS + PA.EXCLUDE_EDGE_PARTICLES + PA.INCLUDE_HOLES # option for stack missing ##area mean centroid bounding integrated stack redirect=None decimal=4 meas = Meas.AREA + Meas.MEAN + Meas.CENTROID + Meas.RECT + Meas.INTEGRATED_DENSITY + Meas.STACK_POSITION rt = ResultsTable().getResultsTable() pa = PA(opt, meas, rt, 10.0, 300000.0, 0, 1.0) PA.processStack = True pa.setHideOutputImage(True) ##run("Analyze Particles...", "size=800-Infinity circularity=0.00-1.00 pixel show=Masks display exclude include stack"); outstack = ImageStack(impdup.getWidth(), impdup.getHeight()) for i in range(1,impdup.getStackSize()+1): impdup.setSlice(i) pa.analyze(impdup) impbin = pa.getOutputImage() outstack.addSlice(impbin.getProcessor()) impbin = ImagePlus("out", outstack) IJ.run(impbin, "Invert LUT", "") #IJ.run(impbin, "Fill Holes", "stack") return impbin, rt
def threshold(imPlus, edgeThreshold=2500): mask = Duplicator().run(imPlus) mask_stk = mask.getStack() # First, we threshold based on edges IJ.setThreshold(mask, edgeThreshold, 100000, "No Update") for i in range(mask.getImageStackSize()): mask_stk.getProcessor(i + 1).findEdges() IJ.run(mask, "Make Binary", "method=Default background=Default black") # Now, we need to clean up the binary images morphologically IJ.run(mask, "Dilate", "stack") IJ.run(mask, "Fill Holes", "stack") IJ.run(mask, "Erode", "stack") IJ.run(mask, "Erode", "stack") # Finally, remove the small particles stk = ImageStack(mask.getWidth(), mask.getHeight()) p = PA(PA.SHOW_MASKS, 0, None, 200, 100000) p.setHideOutputImage(True) for i in range(mask_stk.getSize()): mask.setSliceWithoutUpdate(i + 1) p.analyze(mask) mmap = p.getOutputImage() stk.addSlice(mmap.getProcessor()) mask.setStack(stk) mask.setSliceWithoutUpdate(1) mask.setTitle(mask_title(imPlus.getTitle())) mask.show() return mask
def generate_background_rois(input_mask_imp, params, membrane_edges, dilations=5, threshold_method=None, membrane_imp=None): """automatically identify background region based on auto-thresholded image, existing membrane edges and position of midpoint anchor""" if input_mask_imp is None and membrane_imp is not None: segmentation_imp = Duplicator().run(membrane_imp) # do thresholding using either previous method if threhsold_method is None or using (less conservative?) threshold method if (threshold_method is None or not (threshold_method in params.listThresholdMethods())): mask_imp = make_and_clean_binary(segmentation_imp, params.threshold_method) else: mask_imp = make_and_clean_binary(segmentation_imp, threshold_method) segmentation_imp.close() else: input_mask_imp.killRoi() mask_imp = Duplicator().run(input_mask_imp) rois = [] IJ.setForegroundColor(0, 0, 0) roim = RoiManager(True) rt = ResultsTable() for fridx in range(mask_imp.getNFrames()): mask_imp.setT(fridx + 1) # add extra bit to binary mask from loaded membrane in case user refined edges... # flip midpoint anchor across the line joining the two extremes of the membrane, # and fill in the triangle made by this new point and those extremes poly = membrane_edges[fridx].getPolygon() l1 = (poly.xpoints[0], poly.ypoints[0]) l2 = (poly.xpoints[-1], poly.ypoints[-1]) M = (0.5 * (l1[0] + l2[0]), 0.5 * (l1[1] + l2[1])) Mp1 = (params.manual_anchor_midpoint[0][0] - M[0], params.manual_anchor_midpoint[0][1] - M[1]) p2 = (M[0] - Mp1[0], M[1] - Mp1[1]) new_poly_x = list(poly.xpoints) new_poly_x.append(p2[0]) new_poly_y = list(poly.ypoints) new_poly_y.append(p2[1]) mask_imp.setRoi(PolygonRoi(new_poly_x, new_poly_y, PolygonRoi.POLYGON)) IJ.run(mask_imp, "Fill", "slice") mask_imp.killRoi() # now dilate the masked image and identify the unmasked region closest to the midpoint anchor ip = mask_imp.getProcessor() dilations = 5 for d in range(dilations): ip.dilate() ip.invert() mask_imp.setProcessor(ip) mxsz = mask_imp.getWidth() * mask_imp.getHeight() pa = ParticleAnalyzer( ParticleAnalyzer.ADD_TO_MANAGER | ParticleAnalyzer.SHOW_PROGRESS, ParticleAnalyzer.CENTROID, rt, 0, mxsz) pa.setRoiManager(roim) pa.analyze(mask_imp) ds_to_anchor = [ math.sqrt((x - params.manual_anchor_midpoint[0][0])**2 + (y - params.manual_anchor_midpoint[0][1])**2) for x, y in zip( rt.getColumn(rt.getColumnIndex("X")).tolist(), rt.getColumn(rt.getColumnIndex("Y")).tolist()) ] if len(ds_to_anchor) > 0: roi = roim.getRoi(ds_to_anchor.index(min(ds_to_anchor))) rois.append(roi) else: rois.append(None) roim.reset() rt.reset() roim.close() mask_imp.close() return rois
def __addroi(self, event) : if ( not self.__init) : IJ.showMessage("", "please start a new stack") return if ( not self.__initDIA) : IJ.showMessage("", "please select an image for DIA") return if ( not self.__initFLUO) : IJ.showMessage("", "please select an image for FLUO") return twres = TextWindow("measures-"+self.__name, "label\tname\tsol\tarea\tcirc\tAR\tFeret\taxis\traf\tdMajor\tdFeret\tdArea", "", 300, 450) tab="\t" self.__widthl = self.__display2.getText() IJ.selectWindow(self.__impF.getTitle()) self.__rm = RoiManager.getInstance() if (self.__rm==None): self.__rm = RoiManager() if self.__impF.getImageStackSize() > 1 : roisarray =[(roi, self.__rm.getSliceNumber(roi.getName())) for roi in self.__rm.getRoisAsArray()] else : roisarray =[(roi, 1) for roi in self.__rm.getRoisAsArray()] self.__rm.runCommand("reset") #self.__rm.runCommand("Delete") IJ.selectWindow(self.__impF.getTitle()) self.__maxraf=float(self.__display19.text) self.__minraf=float(self.__display20.text) count=1 for roielement in roisarray : roi = roielement[0] pos = roielement[1] lab = self.__impF.getImageStack().getShortSliceLabel(pos) if lab==None : lab=str(pos) if self.__conEllipses : IJ.selectWindow(self.__impF.getTitle()) self.__impF.setSlice(pos) self.__impF.setRoi(roi) self.__rm.runCommand("Add") IJ.run(self.__impF, "Fit Ellipse", "") ellipse=self.__impF.getRoi() params = ellipse.getParams() ferets = ellipse.getFeretValues() imp2 = Duplicator().run(self.__impF,pos,pos) IJ.run(imp2, "Rotate... ", "angle="+str(ferets[1])+" grid=0 interpolation=Bilinear enlarge slice") temproi=Roi((imp2.getWidth()-ferets[0])/2.0,(imp2.getHeight()-ferets[2])/2.0,ferets[0],ferets[2]) imp2.setRoi(temproi) imp3 = Duplicator().run(imp2,1,1) ip3=imp3.getProcessor() if int(self.__display5.text) < ip3.getWidth() < int(self.__display6.text) : self.__iplist.append(ip3) self.__display.text = self.__name + " cell " + str(len(self.__iplist)) fer=Line(params[0],params[1],params[2],params[3]) self.__cellsrois.append((fer, pos)) self.__labels.append(self.__isF.getShortSliceLabel(pos)) m=Morph(self.__impF, roi) twres.append(lab+tab+str(roi.getName())+tab+str(m.Solidity)+tab+str(m.Area)+tab+str(m.Circ)+tab+str(m.AR)+tab+str(m.MaxFeret)+tab+str(fer.getLength())+tab+str(1)+tab+str(0)+tab+str(0)+tab+str(0)) self.__dictCells[count]=(str(roi.getName()), lab, roi) count=count+1 continue if roi.getType() in [6,7] : self.__impF.setSlice(pos) self.__impF.setRoi(roi) self.__rm.runCommand("Add") elif roi.getType() in [2,4] : self.__impF.setSlice(pos) self.__impF.setRoi(roi) m=Morph(self.__impF, roi) m.setMidParams(10, 2) midroi=m.MidAxis if midroi == None : continue raf = m.MaxFeret/midroi.getLength() if (self.__maxraf < raf) or (raf < self.__minraf) : continue maxsol = float(self.__display7.text) minsol = float(self.__display8.text) maxarea = float(self.__display9.text) minarea = float(self.__display10.text) maxcirc = float(self.__display11.text) mincirc = float(self.__display12.text) maxar = float(self.__display13.text) minar = float(self.__display14.text) maxfer = float(self.__display15.text) minfer = float(self.__display16.text) maxmean = float(self.__display17.text) minmean = float(self.__display18.text) maxmferet = float(self.__display21.text) minmferet = float(self.__display22.text) testsol = (minsol<= m.Solidity <= maxsol) testarea = (minarea<= m.Area <= maxarea) testcirc = (mincirc<= m.Circ <= maxcirc) testar = (minar<= m.AR <= maxar) testfer = (minfer<= m.MaxFeret <= maxfer) testmean = (minmean <= m.Mean <= maxmean) testmferet = (minmferet <= m.MinFeret <= maxmferet) #print minmferet , m.MinFeret , maxmferet test = (testsol+testarea+testcirc+testar+testfer+testmean+testmferet)/7 if test : fmaj, ffmx, fa =[],[],[] for r in m.getMidSegments(10, 40, 0)[0] : if r == None : continue m2=Morph(self.__impF, r) fmaj.append(m2.Major) ffmx.append(m2.MaxFeret) fa.append(m2.Area) diffmajor, diffferet, diffarea = 0,0,0 if len(fa) > 4 : medfmaj = self.listmean(fmaj[1:-1]) medffmx = self.listmean(ffmx[1:-1]) medfa = self.listmean(fa[1:-1]) diffmajor = (max(fmaj[1:-1])-medfmaj)/medfmaj diffferet = (max(ffmx[1:-1])-medffmx)/medffmx diffarea = (max(fa[1:-1])-medfa)/medfa twres.append(lab+tab+str(roi.getName())+tab+str(m.Solidity)+tab+str(m.Area)+tab+str(m.Circ)+tab+str(m.AR)+tab+str(m.MaxFeret)+tab+str(midroi.getLength())+tab+str(m.MaxFeret/midroi.getLength())+tab+str(diffmajor)+tab+str(diffferet)+tab+str(diffarea)) #print lab+tab+str(roi.getName())+tab+str(m.Solidity)+tab+str(m.Area)+tab+str(m.Circ)+tab+str(m.AR)+tab+str(m.MaxFeret)+tab+str(midroi.getLength())+tab+str(m.MaxFeret/midroi.getLength())+tab+str(diffmajor)+tab+str(diffferet)+tab+str(diffarea) self.__impF.setRoi(roi) self.__rm.runCommand("Add") self.__impF.killRoi() self.__impF.setRoi(midroi) #self.__dictCells[str(roi.getName())]=(str(roi.getName()), lab, roi) self.__dictCells[count]=(str(roi.getName()), lab, roi) count=count+1 else : #print "test falls" continue else : print "out loop" continue straightener = Straightener() new_ip = straightener.straighten(self.__impF, midroi, int(self.__widthl)) if int(self.__display5.text) < new_ip.getWidth() < int(self.__display6.text) : self.__iplist.append(new_ip.convertToShort(False)) self.__display.text = self.__name + " cell " + str(len(self.__iplist)) #print "add", roi.getName(), roi.getType() self.__cellsrois.append((midroi, pos)) self.__labels.append(self.__isF.getShortSliceLabel(pos)) #roisarray=self.__rm.getRoisAsArray() #self.__rm.runCommand("reset") #self.__rm.runCommand("Delete") self.__impD.killRoi() self.__impF.killRoi() IJ.selectWindow(self.__impD.getTitle())