Enhancing depth of field by combining images

by Wim van Egmond, The Netherlands

In two earlier Micscape articles I showed the possibilities of mouse-over images with 2 photomicrographs focussed in different focal planes. It is also possible to combine different images like that in one image. 

The pictures on this page show a very simple but useful example.


I have been working on radiolaria for some time now. Making mounted slides of the glass-like skeletons of these beautiful oceanic protozoa can be rewarding. In my case not that rewarding since I had terrible difficulty in getting mounts that lasted. But that is another story I will save for a later issue of Micscape :-).

These radiolaria are often too three-dimensional to get all parts of the structure in focus. The specimen on this page is almost spherical with protrusions only along the 'equator'. This made it possible to use just 2 images to create a sharp image. To enable this, it was important to make a mounted slide with the radiolaria in the right position so the protrusions were in the same focal plane. I mounted the radiolaria in Canada Balsam so I had a slide with a relatively high refractive index so the shell would get a good contrast. When you observe them in water these creatures will be almost invisible.

I scanned the images and made the final photo-montage in the computer. In Photoshop (and I am sure in many image editing programs) it is possible to make a selection with soft edges. I made a circular (feathered) selection from the center of the first image (left image below), copied it and pasted it at the right spot in the other image (right image below). When using soft edged selections you can hardly see where the border between the 2 images is.

It is a very simple trick that can be used on many subjects like Forams, Ostracods, plant pollen.

The two images used to create the montage at article head. Photographed with a 16X obj. (D.I.C.) 
Differential Interference Contrast works quite nice on these glass-like subjects.

All comments to the author Wim van Egmond are welcomed.

Visit Wim's home page for links to his many web pages on microscopy

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Published in the October 2001 edition of Micscape Magazine.

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