Page 152 - Fundamentals of Light Microscopy and Electronic Imaging

CHAPTER
9

POLARIZATION MICROSCOPY

OVERVIEW

Image formation in the polarizing microscope is based on the unique ability of polarized
light to interact with polarizable bonds of ordered molecules in a direction-sensitive
manner. Perturbations to waves of polarized light from aligned molecules in an object
result in phase retardations between sampling beams, which in turn allow interference-
dependent changes in amplitude in the image plane. Thus, image formation is based not
only on principles of diffraction and interference, but also on the existence of ordered
molecular arrangements. The degree of order encountered in objects ranges from near-
perfect crystals to loosely ordered associations of asymmetric molecules or molecular
assemblies. In the polarizing microscope, such structures generally appear bright
against a dark background (Fig. 9-1). Polarization microscopy has been used to study
the form and dynamics of many ordered cellular structures, including:

• Mitotic spindle fibers in dividing cells
• Actin filament bundles in a variety of cell types
• Actin and myosin filaments in the myofibrils of striated muscle cells
• Condensed DNA in certain sperm nuclei
• Kinetoplast DNA in the mitochondria of trypanosomes
• Helical strands of cellulose fibers in plant cell walls
• Condensates of starch and lignin in plant cells
• Virus crystalloids and crystals of organic compounds in the cytoplasm of plant cells
• Lipid bilayers of the cell plasma membrane and mitochondria

In many cases, polarization microscopy is the only available method for studying
the structure, formation, and dynamics of labile macromolecular assemblies or examin-
ing the effects of chemicals, drugs, or environmental conditions on cellular structures in
vivo. For additional examples of the application of polarized light in studies of mitosis,
see Inoué and Oldenbourg (1998) and Oldenbourg (1996, 1999).
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