Gel Intensity Normalization and Band Analysis

// Wait for the gel image to open
waitForImage();

// Display a message and prompt the user to draw a rectangle around the gel
waitForUser("Draw a rectangle around the gel");

// Get the ROI (Region of Interest) of the gel
getSelectionBounds(x, y, width, height);

// Check if a valid ROI has been selected
if (width > 0 && height > 0) {
  // Select the ROI
  makeRectangle(x, y, width, height);

  // Calculate the mean intensity value within the ROI
  run("Measure");

  // Get the mean intensity value
  var mean = getResult("Mean", 0);

  // Check if the mean intensity value is valid
  if (mean > 0) {
    // Normalize the image by dividing it by the mean intensity value
    run("Divide...", "value=" + mean);

    // Enhance the contrast and display the normalized image
    run("Enhance Contrast", "saturated=0.35");

    // Create a copy of the original gel image
    selectWindow("Original gel");
    run("Duplicate...", "title=Original_gel");

    // Display a message and prompt the user to select the bands of interest
    waitForUser("Use the 'Rectangular' tool to select the bands of interest");

    // Create a list to store the coordinates of the selected bands
    var bandCoords = [];

    // Loop for the user to select multiple bands of interest
    while (true) {
      // Wait for the user to select a band of interest
      waitForSelection("rectangle");

      // Get the coordinates of the selection
      getSelectionBounds(x_band, y_band, width_band, height_band);

      // Check if a valid band has been selected
      if (width_band > 0 && height_band > 0) {
        // Store the coordinates of the selected band in the list
        bandCoords.push([x_band, y_band, width_band, height_band]);

        // Draw the lines selected by the user on the graph
        setForegroundColor(255, 0, 0); // Red color
        drawLine(x_band, y_band + height_band / 2, x_band + width_band, y_band + height_band / 2);
        drawLine(x_band, y_band + height_band / 2 - 1, x_band + width_band, y_band + height_band / 2 - 1);
        drawLine(x_band, y_band + height_band / 2 + 1, x_band + width_band, y_band + height_band / 2 + 1);
      } else {
        // No valid band selected, end the selection process
        if (bandCoords.length === 0) {
          showError("Error: No bands of interest have been selected.");
        }
        break;
      }
    }

    // Check if bands of interest have been selected
    if (bandCoords.length > 0) {
      // Create a plot with the normalized intensity values
      newPlot("Normalized Intensity Profile", "Position", "Intensity");

      // Process each selected band
      for (var i = 0; i < bandCoords.length; i++) {
        var band = bandCoords[i];
        var x_band = band[0];
        var y_band = band[1];
        var width_band = band[2];
        var height_band = band[3];

        // Select the band of interest
        makeRectangle(x_band, y_band, width_band, height_band);

        // Calculate the normalized intensity profile in the selected band
        run("Plot Profile");

        // Get the normalized intensity values
        var intensityArray = newArray(width_band);
        getProfile(intensityArray);

        // Add the normalized intensity values to the plot
        addValues("Normalized Intensity Profile", newArray(width_band), intensityArray);
      }

      // Save the original gel image
      selectWindow("Original_gel");
      var originalGelPath = getPath("Save");
      if (originalGelPath) {
        saveAs("Tiff", originalGelPath);
      } else {
        showError("Error: No valid location has been selected to save the original gel image.");
      }

      // Save the plot
      selectWindow("Normalized Intensity Profile");
      var graphPath = getPath("Save");
      if (graphPath) {
        saveAs("PNG", graphPath);
      } else {
        showError("Error: No valid location has been selected to save the plot.");
      }

      // Save the normalized gel image
      selectWindow("Normalized gel");
      var normalizedGelPath = getPath("Save");
      if (normalizedGelPath) {
        saveAs("Tiff", normalizedGelPath);
      } else {
        showError("Error: No valid location has been selected to save the normalized gel image.");
      }

      // Display a success message with the saved files
      showSuccess("Process completed. The original gel image, normalized gel image, and plot have been saved.");

    } else {
      showError("Error: No valid bands of interest have been selected.");
    }
  } else {
    showError("Error: The mean intensity value is zero or negative. The image cannot be normalized.");
  }
} else {
  showError("Error: No valid region of interest has been selected.");
}

// Function to display an error message
function showError(message) {
  showMessage("Error", message, "Error");
}

// Function to display a success message
function showSuccess(message) {
  showMessage("Success", message, "Information");
}

// Generic function to display a message
function showMessage(title, message, type) {
  IJ.showMessage(title, message, type);
}

// Function to prompt the user for a location and file name to save
function getPath(action) {
  var dialog = FileDialog(action);
  dialog.setVisible(true);
  var selectedPath = dialog.getDirectory() + dialog.getFileName();
  return selectedPath !== "nullnull" ? selectedPath : null;
}