Centrifuge Polarizing Microscope

Abstract

<jats:p>We report here the successful development of a new instrument, the Centrifuge Polarizing Microscope (CPM). The CPM was developed to explore, by high-extinction polarized light microscopy, the dynamic alignment, linkage, and interaction of macromolecules, fine structure, and organelles in living cells and developing embryos being centrifugally stratified. It should also find applications in material sciences, e.g., for examining centrifugal stratification, compaction, and alignment of liquid crystals and emulsions; the CPM uniquely allows continuous observation and measurement of the weak birefringence exhibited by molecules and fine structures that are oriented, or become oriented, under centrifugal acceleration.</jats:p><jats:p>Centrifuge microscopes of various designs have been built since the 1930s but were not equipped for polarization optics. In the 8-cm-radius rotor of our prototype CPM, spinning between the objective and condenser lenses with numerical apertures of up to 0.55, the specimen chamber is illuminated stroboscopically by a 5- to 7-ns intense flash of 532-nm-wavelength light from a Nd- YAG laser, synchronized to fire exactly as the specimen intersects the axis of the objective lens (Fig. 1).</jats:p>

How well can an amoeba climb?

“…The centrifuge polarizing microscope was designed by S.I. and developed in collaboration with Hamamatsu Photonics (Hamamatsu City, Japan) and Olympus Optical Company (Tokyo) (11). Cells were suspended in standard buffer and allowed to settle on the strain-free glass cover of the centrifuge observation chamber.…”

“…In the present paper, we report the maximum ''apparent weight,'' or centrifugal force against which wild-type and myosin mutants of Dictyostelium discoideum amoebae were able to crawl ''upward.'' The small mass of the amoebae required the use of a recently developed centrifuge polarizing microscope capable of generating fields of greater than 11,465 ϫ g (Earth's gravitational acceleration), with image resolution of better than 1 m in differential interference or Nomarsky contrast microscopy (11).…”

Polarization Microscopy

“…(From Nicklas, .) For online version of time‐lapse series of this and other cells in mitosis, see http://www.molbiolcell.org and Inoué and Oldenbourg ().…”

Windows to dynamic fine structures, then and now

“…A special interference‐fringe‐free CCD camera captures the image. The output of the CCD camera is processed, combined with data signals, recorded, and displayed as seen in Figures 4 and 5 (21).…”

“…The part of the test target, made by electron lithography on a thin aluminum layer, shows the ruling with the 1.0‐µM period clearly resolved (upper right). The left and right numbers before ‘RPM’ are the momentary and designated speed for the rotor (21).…”