/*
* Cell_Proliferation
* Authors: Pau Carrillo-Barberà, José M. Morante-Redolat, José F. Pertusa
* Department of Cellular & Functional Biology
* University of Valencia (Valencia, Spain)
*/
/*
* This macro is a high-throughput screening tool for cell proliferation assays.
* It is based on nucleoside analogue pulse alone or in combination with up to
* two additional nuclear markers.
*/
macro "Cell_Proliferation" {
//choose a macro mode and a directory
#@ String (label=" ", value="High Content Screening
Cell Proliferation", visibility=MESSAGE, persist=false) heading
#@ String(label="Select mode:", choices={"Analysis", "Pre-Analysis (parameter tweaking)"}, style="radioButtonVertical") mode
#@ File(label="Select a directory:", style="directory") dir
#@ String (label="Load pre-established
parameter dataset?", choices={"No", "Yes"}, persist=true, style="radioButtonHorizontal") importPD
#@ String (label="Load llumination
correction reference
image?", choices={"No", "Yes"}, value="Yes", persist=true, style="radioButtonHorizontal") illumCorr
#@ String (label="Save ROIs?", choices={"No", "Yes"}, value="Yes", persist=true, style="radioButtonHorizontal") saveROIs
#@ String (label=" ", value="![](\"https://live.staticflickr.com/65535/48557333566_d2a51be746_o.png\")
", visibility=MESSAGE, persist=false) logo
#@ String (label=" ", value="Neuromolecular Biology Lab
ERI BIOTECMED - Universitat de València (Spain)", visibility=MESSAGE, persist=false) message
//set options
setOption("ExpandableArrays", true);
setOption("BlackBackground", false);
setOption("ScaleConversions", true);
roiManager("reset");
print("\\Clear");
roiManager("reset");
close("*");
//File management
//Identification of the TIF files
//create an array containing the names of the files in the directory path
list = getFileList(dir);
Array.sort(list);
tifFiles=0;
//count the number of TIF files
for (i=0; i count+1) {
index1=indexOf(tifArray[count], "wv ");
index2=lastIndexOf(tifArray[count], " - ");
channels[count]=substring(tifArray[count], index1+3, index2);
count++;
}
//add an 'Empty' option into the channels name array
for (i=0; i=imagesxfield) {
channels[i]="Empty";
}
}
//create a channel array without the 'Empty' option
channelsSlice=Array.slice(channels, 0, channels.length-1);
//set some parameter menu arrays
threshold=getList("threshold.methods");
pattern=newArray(4);
flat_field=newArray(4);
//create a flat-field array with a 'None' option
if(illumCorr=="Yes") {
illumCorrPath=getDirectory("Choose the folder containing the flat-field images");
illumCorrList=getFileList(illumCorrPath);
concatNone=newArray("None");
illumCorrList=Array.concat(concatNone,illumCorrList);
} else {
illumCorrList=newArray("None");
}
//Extract values from a parameter dataset file
if(importPD=="Yes") {
parametersDatasetPath=File.openDialog("Choose the parameter dataset file:");
//parameter selection (pre-established)
parametersString=File.openAsString(parametersDatasetPath);
parameterRows=split(parametersString, "\n");
parameters=newArray(parameterRows.length);
for(i=0; idocumentation"
+" for help";
Dialog.addHelp(html);
Dialog.show()
projectName=Dialog.getString();
pattern[0]=Dialog.getChoice();
pattern[1]=Dialog.getChoice();
pattern[2]=Dialog.getChoice();
pattern[3]=Dialog.getChoice();
normalize=Dialog.getCheckbox();
gaussianNuclei=Dialog.getNumber();
thresholdNuclei=Dialog.getChoice();
erodeNuclei=Dialog.getNumber();
openNuclei=Dialog.getNumber();
watershedNuclei=Dialog.getCheckbox();
size=Dialog.getString();
flat_field[0]=Dialog.getChoice();
flat_field[1]=Dialog.getChoice();
flat_field[2]=Dialog.getChoice();
flat_field[3]=Dialog.getChoice();
//check the channel selection
if(pattern[0]==pattern[1]) {
beep();
exit("Nuclei ["+pattern[0]+"] and nucleoside analogue ["+pattern[1]+"] channels can not be the same")
} else if (pattern[0]==pattern[2] || pattern[0]==pattern[3]) {
beep();
exit("Nuclei ["+pattern[0]+"] and addititional marker ["+pattern[2]+"]/["+pattern[3]+"] channels can not be the same")
} else if (pattern[1]==pattern[2] || pattern[1]==pattern[3]) {
beep();
exit("Nucleoside analogue ["+pattern[1]+"] and addititional marker ["+pattern[2]+"]/["+pattern[3]+"] channels can not be the same")
} else if (pattern[2]==pattern[3] && pattern[2]!="Empty") {
beep();
exit("Addititional marker ["+pattern[2]+"]/["+pattern[3]+"] channels can not be the same")
}
//get min and max sizes
size=min_max_size(size);
//Create a parameter dataset file
title1 = "Parameter dataset";
title2 = "["+title1+"]";
f = title2;
run("Table...", "name="+title2+" width=500 height=500");
print(f, "Project\t" + projectName);
print(f, "Nuclei\t" + pattern[0]);
print(f, "Nucleoside analogue\t" + pattern[1]);
print(f, "Marker1\t" + pattern[2]);
print(f, "Marker2\t" + pattern[3]);
print(f, "Enhance (nuclei)\t" + normalize);
print(f, "Gaussian (nuclei)\t" + gaussianNuclei);
print(f, "Threshold (nuclei)\t" + thresholdNuclei);
print(f, "Erode (nuclei)\t" + erodeNuclei);
print(f, "Open (nuclei)\t" + openNuclei);
print(f, "Watershed (nuclei)\t" + watershedNuclei);
print(f, "Size\t" + size[0]+"-"+size[1]);
print(f, "Flat-field (nuclei)\t" + flat_field[0]);
print(f, "Flat-field (nucleoside analogue)\t" + flat_field[1]);
print(f, "Flat-field (marker1)\t" + flat_field[2]);
print(f, "Flat-field (marker2)\t" + flat_field[3]);
//save as TXT
saveAs("txt", dir+File.separator+projectName);
selectWindow(title1);
run("Close");
//'Well Selection' dialog box
selectionOptions=newArray("Select All", "Include", "Exclude");
fileCheckbox=newArray(nWells);
selection=newArray(nWells);
title = "Select Wells";
Dialog.create(title);
Dialog.addRadioButtonGroup("", selectionOptions, 3, 1, selectionOptions[0]);
Dialog.addCheckboxGroup(sqrt(nWells) + 1, sqrt(nWells) + 1, wellName, selection);
if(mode=="Pre-Analysis (parameter tweaking)") {
if(fieldsxwell>=10) {
maxRandomFields=10;
} else {
maxRandomFields=fieldsxwell;
}
Dialog.addMessage("Random fields per well:");
Dialog.addSlider("", 1, maxRandomFields, maxRandomFields);
measurements=newArray("Area", "Circ.", "AR", "Solidity", "Round", "Mean", "IntDen");
Dialog.addChoice("Measure", measurements, "Mean");
Dialog.addNumber("Split threshold", 500);
}
Dialog.show();
selectionMode=Dialog.getRadioButton();
if(mode=="Pre-Analysis (parameter tweaking)") {
maxRandomFields=Dialog.getNumber();
measure_test=Dialog.getChoice();
split_test=Dialog.getNumber();
}
for (i=0; i=0; i--) {
if (number==randomArray[i]) {
recurrent=true;
}
}
if(recurrent==false || count==0) {
// open images
channels_test=newArray(2);
for (i=0; i threshold) {
roiManager("select", a);
roiManager("Set Color", "orange");
roiManager("draw");
} else {
roiManager("select", a);
roiManager("Set Color", "cyan");
roiManager("draw");
}
}
roiManager("reset");
run("Clear Results");
}
}