Microscopy Techniques….

Welcome to Small World! The 20 images in this exhibition represent the

winning entries in Nikon’s annual photomicrography competition. All of
these stunning creations have only one thing in common: they were taken
through some type of microscope. Beyond that, they are unique examples
of the wide variety of microscopy techniques available today.

As you explore the wonder and beauty of these images, take a moment to
read a little about how they were created. Explanations of the techniques
are placed throughout the exhibit and are numbered in ascending order
of complexity. Start here with #1 (Reflected Light Stereomicroscopy) and
work clockwise around the atrium until you reach #11 (2-Photon Confocal).
Each description tends to build a little more on the previous diagrams to
help you understand the technology required to make these images.

In addition to the equipment descriptions, we have included questions and

answers by the individual photomicrographers to help you appreciate the
additional elements of creativity and scientific content that it takes to create
these scientifically significant and eye catching images.
© James E. Hayden, RBC,FBCA
The Wistar Institute
© James E. Hayden, RBC,FBCA
The Wistar Institute
1
Reflected Light
Stereomicroscopy
The stereomicroscope, as shown in the previous diagram, is a relatively
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standards) specimens. When looking through the microscope, the viewer
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paths to the eyepieces, each from a slightly different angle. Imaging is
accomplished by diverting the light from one of those paths to the side-
mounted camera. The resulting image can only be seen in 2 dimensions, so
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Lighting techniques for the stereomicroscope are very similar to techniques

used for much larger subjects, like people portraits. The main difference
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simply means that the illumination on the subject is coming from above and
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used more than one light to accomplish their task; the 1st place entry needed
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the result, while the 15th place entry used four very small lights aimed at
different angles.
© James E. Hayden, RBC,FBCA
The Wistar Institute
2
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© James E. Hayden, RBC,FBCA
The Wistar Institute
3
Brightfield
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© James E. Hayden, RBC,FBCA
The Wistar Institute
4
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© James E. Hayden, RBC,FBCA
The Wistar Institute
5
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© James E. Hayden, RBC,FBCA
The Wistar Institute
6
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The lighting technique shown in the 19th place winning entry of a bird
feather is a simple expansion of the polarization described in the previous
(#5) technique. In this example, standard cross polarization creates colors
and contrast in the birefringent specimen, but additional color is added by
inserting a red/blue quadrant Rheinberg filter below the specimen. This
filter adds out of focus colored light to the background and also attenuates
the colors created by the cross polarization.
© James E. Hayden, RBC,FBCA
The Wistar Institute
7
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© James E. Hayden, RBC,FBCA
The Wistar Institute
8
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© James E. Hayden, RBC,FBCA
The Wistar Institute
9
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© James E. Hayden, RBC,FBCA
The Wistar Institute
10
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© James E. Hayden, RBC,FBCA
The Wistar Institute
11
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© James E. Hayden, RBC,FBCA
The Wistar Institute
 Credits
JAMES E. HAYDEN, RBP, FBCA
Manager, Microscopy Core Facility
jhayden@wistar.org

FREDERICK S. KEENEY
Assistant, Microscopy Core Facility

CHRISTINE DELAURENTIS
Graphic Artist

SOHEILA NIKPOUR
Web Developer

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3601 Spruce Street Philadelphia, PA 19104 www.wistar.org