Enhance Images

Once the image is captured, it is a digital file (matrix of pixels). Cytogenetic software provides powerful tools to enhance the visibility of bands and correct physical preparation defects. However, Enhancement is distinct from Manipulation. Enhancement makes existing data clearer; manipulation creates false data. Clinical laboratories must strictly adhere to ethical guidelines regarding image processing

Standard Enhancement Tools

These tools are applied routinely to facilitate karyotyping

• Contrast and Brightness (The Histogram)
  • Goal: To maximize the dynamic range of the image
  • Action: The “Black Point” is set to the darkest pixel in the chromosomes, and the “White Point” is set to the background. This stretches the grey values, making faint bands darker and the background cleaner
  • Caution: Excessive contrast (“Thresholding”) turns the chromosome into a black silhouette, erasing the G-bands entirely
• Background Flattening
  • Goal: To remove debris, stain precipitate, or uneven lighting from the empty space around the chromosomes
  • Action: The software identifies pixels that are not part of a chromosome and sets them to uniform white. This produces the clean “textbook” look of a clinical karyotype
• Sharpening (Filters)
  • Goal: To define the edges of bands
  • Action: Algorithms (like Laplacian or Unsharp Mask) increase the contrast at boundaries between light and dark pixels
  • Caution: Over-sharpening creates “salt and pepper” noise and ragged chromosome edges

Geometric Corrections

Chromosomes are flexible biological structures. They often land on the slide bent, crossed, or touching. The software allows the laboratory scientist to “fix” these physical problems digitally

• Separation (Cutting)
  • Problem: Two chromosomes are touching or slightly overlapping
  • Action: The laboratory scientist draws a line between them. The software splits the object into two separate distinct images
• Straightening
  • Problem: A long chromosome (e.g., Chromosome 1) is curved like a “C” or “S”. This makes it hard to compare to its straight homolog
  • Action: The laboratory scientist draws a guideline down the center axis of the curved chromosome. The software maps the pixels to a straight line
  • Rule: Straightening should only be used if the banding pattern remains undistorted. If straightening stretches or compresses bands artificially, the chromosome should be left curved
• Resolving Overlaps
  • Problem: Chromosome A is lying on top of Chromosome B (a “cross”)
  • Action: This is the most dangerous enhancement. Modern software can attempt to mathematically “subtract” the top chromosome to reveal the bottom one, but this is prone to error
  • Best Practice: Find a better metaphase. Do not rely on heavily overlapped cells for diagnosis

Ethics of Enhancement

• Permissible: Adjusting contrast to verify bands; Straightening curved chromosomes; Removing background dirt
• Forbidden
  • Deleting Signals: Using the “eraser” tool to remove a FISH signal that “looks like background.”
  • Copy/Paste: Copying a normal chromosome from another cell to replace a blurry one in the current cell
  • Band Drawing: Using a paintbrush tool to darken a band that “should be there.”
• Audit Trail: Clinical software preserves the Original Raw Image. Any changes made are saved as a separate layer or file. An auditor can always revert to the raw capture to verify the diagnosis