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Electron Microscopy Sciences

Technical Data Sheets

Focus Stacking for Improved Depth of Field

EMS Catalog #SFA Nightsea

Some kinds of photography have shallow depth of field, making it difficult to impossible to have as much of the subject in focus as you would like. This is especially true for macro photography and for photography under the microscope. The technique of focus stacking combines multiple images, each taken with a slightly different plane of focus, into a single in-focus image. Google ‘focus stacking’ to find plenty of posts with explanations and examples, and Wikipedia has an explanation and a list of focus stacking software.

Focus stacking works great with fluorescence photography. An ant had the audacity to crawl around the office one day so of course we put it under the stereo microscope to see what we could see. Using the GFP excitation/emission set of the NIGHTSEA Stereo Microscope Fluorescence Adapter there was a striking yellow fluorescence from hairs on the abdomen. Only a small portion of the curved surface was sharp with any one focus setting of the microscope, so we collected a series of images and combined them using Helicon Focus. The first two images show very different parts of the scene in focus, while the third shows the result of the focus stacking.


  • Stereo microscope fluorescence photograph of ant abdomen. © Charles Mazel
  • Stereo microscope fluorescence photograph of ant abdomen. © Charles Mazel
  • Stereo microscope fluorescence photograph of ant abdomen, with focus stacking. © Charles Mazel

For another example we took photographs of a burr that had caught in clothes while out mowing the lawn. This clothes-grabbing property burrs is what led to the invention of velcro®.


  • Fluorescence photograph of a burr. © Charles Mazel
  • Fluorescence photograph of a burr. © Charles Mazel
  • Fluorescence photograph of a burr, with focus stacking. © Charles Mazel

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