Image stacking - ground beetle and microscope

Ground beetle, genus Agonum, shown with photoshop layers for image stacking
Most people associate the use of slides with microscopes, but you can also use anything that will fit under the objectives to view.  The issue is translucency, as thicker subject block the light which typically comes from underneath the specimen.  The key then is to use multiple light sources that shine directly upon the subject from acute positions to adequately illuminate it.

Image stacking, also called focus stacking, is a method of increasing depth of field.  Microscopes are notorious for having a very narrow depth of field, becoming increasingly narrow as magnification increases.  At low-power there may be as much as a millimeter but at high-power it can be measured in microns (also called micrometers; 1000 µm = 1 mm).  Taking any one photograph (inset, bottom right) will render a great deal of the specimen out of focus.  This is where image stacking helps.

Starting at the zenith of focus for the subject, take an image and refocus at a slightly lower point preferably that there is some focus overlap.  Take another picture and refocus again, repeating the procedure until all points have been covered.  Take the digital images to a computer and import them into a program that can merge them into a single unit.  I have used Zerene stacker in the past but recently discovered that it can be done with Photoshop CC.

I then began importing the seven photos into Photoshop using the "Load Files into Scripts" command found under Files>Scripts.  The inset (top left) shows the resulting layers.  Sometimes the images have to be adjusted if they do not line up but in this case the shots were perfectly synchronous.  Using the "Auto Blend Layers" command found under the Edit tab the program executed and all the images lined up nicely.  The software choose the parts of each shot that were in focus and brought them together nicely resulting in the picture you see above.  Although moderately time-consuming the process from beginning to end can yield startling outcomes.  I am looking forward to doing this again.

Thanks for reading.   www.ericspix.com   Eric Svendsen








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