Image stitcher

Purpose

The Image stitcher assembles multiple individual image-tiles acquired with the Tissue scan workflow into a single image that represents the entire area included in the scan.

Before you start

Complete a tissue scan with the option to Postpone stitching selected.

Compiling slide scans

1. Click Image stitcher on the Image ribbon.

2. A window titled Select the folder that contains the image tiles opens. Navigate to the folder that contains the unstitched image tiles and click Select Folder.

3. Next, a window titled, Choose the folder where the images will be saved opens. Navigate to the folder in which you want to save the stitched tissue scan and click Select Folder.

4. The Preview options window opens. Choose to run a preview of the stitched image before proceeding with the full stitching process or to skip the preview and start stitching, then click OK:

   • Preview images before stitching: Check to create a preview of the stitched tissue scan.

5. Depending on your choices in the previous step, you will generate either:

   • a final stitched image (or images if there were multiple tissue scans in the source-folder selected).

   • a preview image for review—the preview image opens in the Main window along with the Tissue Scan Preview dialog box.

     Review the image preview and indicate how you want to proceed (click to expand):

     • Filter Setup:

       Check or uncheck the box next to the filter name to see the results of applying or removing the filter in your image preview.

       Click Setup to change the filter parameters and preview the effect of the change(s).

     • Background subtraction: Minimize background in images by estimating and removing background illumination using an adaptive, local adjustment of image contrast.

     • Flatfield correction: Mitigates uneven illumination artifacts in fluorescence imaging using intelligent, image-data based correction algorithms—we recommend using it for fluorescence imaging. Check or uncheck the box to see the results of applying or removing the flatfield correction filter in your image preview.

     • Hot pixel filter: Minimize the appearance of hot (abnormally bright) pixels in images. Hot pixels are distinctive because they appear at the same XY position in every z-plane.

     • X,Y save resolution: Choose a resolution from the dropdown menu. Reducing the resolution combines signal from adjacent pixels in order to reduce the size of the stitched image. Note that reducing the resolution can negatively impact image quality. The balance between image size and quality needs to be determined based on your experimental needs for the image data.

     • Start Processing Select options, then click to generate an updated preview or to stitch your tissue scan:

       • Blending: Blending is designed to minimize the appearance of tiling in your stitched image by averaging the pixel intensities where image tiles meet. Confirm or specify the Blend width pixels; to change the blend width, enter a different number of pixels in the box.

       • Adv. Stitching XY and Adv. Stitching XYZ: Advanced stitching uses the image-tile overlap to visually align individual image tiles in the X, Y, and Z or X, Y orientations. It results in excellent image-tile alignment and clean stitched images—we recommend using it in most cases.

       • Stitch into a single image file: Stitch your tissue scan images into one large image.

         Leave the box unchecked and click Start Processing to generate an updated preview with Adv.Stitching XY or Adv. Stitching XYZ and/or Flatfield correction if selected.

     • Postpone Stitching Image-tile files are acquired and saved, but are not stitched into the full tissue scan image. When you're ready to stitch your tissue scan into a single image file, use the Image stitcher on the Image ribbon.

     • Delete Image Deletes the preview image.

     • Focus position: [displayed for 3D tissue scans only] Click the buttons corresponding to different Z-positions to quickly jump to that location for review of the preview image.

     • Subsampling: View a portion of the preview image to quickly determine if it meets your needs.

       Zoom in on the portion of the image that you want to evaluate and click Subsample Preview, to quickly load and view just that portion of the image.

       The subsampling feature can significantly speed up the generation of large preview images, especially if filtering has been applied.

See also Tissue scan workflow