Polarization Microscopy - Biomedical Optical Phase Microscopy and Nanoscopy - page 313

Figure 15.7
LC-PolScope images of single MTs and of actin bundles. (A) Spontaneously assembled MTs,
stabilized with taxol, and adhered to the coverglass surface were imaged with the LC-PolScope
(retardance image). Most filaments are single MTs, with the exception of one bundle containing
one, two, and three MTs. The inset shows line scans across the filament axis at locations with one,
two, and three MTs. Note that at the top right end, the bundle sprays into three individual MTs.
(B) Living growth cone of an Aplysia bag cell neuron. The peripheral lamellar domain contains
radially aligned fibers composed of 15 40 actin filaments. A detailed description of the

birefringent fine structure including a time lapse movie of the growth cone dynamics is published in
[41] . Image brightness represents measured retardance between 0 nm (black) and 0.5 nm or larger
(white). Image width about 70 μ m. Source: Panel (A) Reproduced from Ref. [40] .

The polarization performance of the most highly corrected lenses, the so-called Plan
Apochromat objectives, is often compromised by large number of lens elements, special
antireflection coatings, and, in some cases, special types of glass used to construct the lenses
[21] . For some applications, these high-quality lenses, which provide a large, highly
corrected viewing field over a wide spectral range, might not be required. If the objective
lens is to be used only with the PolScope that requires monochromatic light and typically
acquires images from a region near the center of the viewing field, a less-stringent correction
might well suffice. To find out what works best for a particular imaging situation, several
lenses should be tested. It is also helpful to be able to select the best performing combination
of condenser and objective lens from a batch of the same lens types.

Whenever possible or practical, oil-immersion lenses should be used. This is because the
transition of a light ray between two media of different refractive indices ( n air 5 1.00,
n glass 5 1.52) introduces polarization distortions, especially for high NA lenses. The peripheral
rays leaving the condenser front lens are highly tilted to the slide surface and their polarization
is typically rotated when traversing the air glass interface. Oil, and to a lesser degree water

and other immersion liquids, greatly reduce polarization aberrations caused by air glass

interfaces between the specimen and the lenses. For a discussion of the origin of polarization
distortions in lenses and optical systems, and of ways to reduce them, see Ref. [22] , which also
discusses the various conoscopic images that can be observed in a polarizing microscope
equipped with crossed, or nearly crossed, polarizers and a Bertrand lens.