Table Of ContentANRV356-CB24-01 ARI 27 September 2008 2:16
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
ANNUAL Microtubule Dynamics in
REVIEWS Further
Click here for quick links to
Annual Reviews content online, Cell Division: Exploring
including:
• Other articles in this volume Living Cells with Polarized
• Top cited articles
• Top downloaded articles
• Our comprehensive search Light Microscopy
Shinya Inoue´
Marine Biological Laboratory, Woods Hole, Massachusetts 02543;
email:
ANRV356-CB24-01 ARI 27 September 2008 2:16
the 1980s brought about by electronic imaging
Contents and striking advances in molecular biology.
Although I was able to participate in both
INTRODUCTION . . . . . . . . . . . . . . . . . . 2
recent transitions in microscopy, this Perspec-
ENCOUNTER WITH KATIE AND
tive covers my experience during the earlier part
JEAN DAN: INTRODUCTION
of those events. By following the birefringence
TO LIVING CELLS . . . . . . . . . . . . . . 3
in live dividing cells with an improved polariz-
THE “SHINYA-SCOPE”. . . . . . . . . . . . . 5
ing microscope, we learned about the reality of
TO PRINCETON (1948–51) . . . . . . . . . 7
spindle fibers, the dynamic organization of their
THE PRINCETON
filaments, and their labile assembly/disassembly
MICROSCOPE . . . . . . . . . . . . . . . . . . . 9
and force-generating properties. The dynamic
FINALLY TO WOODS HOLE:
behavior of molecules making up the birefrin-
REALITY AND BEHAVIOR
gent spindle filaments could now be followed
OF SPINDLE FIBERS
directly in actively dividing cells.
AND FIBRILS . . . . . . . . . . . . . . . . . . . . 10
Our studies on live cells were followed by
CONVINCING THE SKEPTICS:
the isolation of a colchicine-binding protein,
SPINDLE FIBERS IN
identified as the microtubule protein, and by
TIME-LAPSE MOVIES OF
the discovery that microtubules could be dis-
DIVIDING CELLS . . . . . . . . . . . . . . . 11
assembled or assembled in vitro. Those studies
THE LABILE NATURE OF
verified our analysis of the birefringence ob-
SPINDLE FIBERS: REVERSIBLE
served in living cells and opened up vast new
DEPOLYMERIZATION AND
avenues for exploring the molecules and mech-
ASSOCIATED CHROMOSOME
anisms involved in mitosis and a wide range of
MOVEMENTS . . . . . . . . . . . . . . . . . . . 12
related cellular events.
ORIENTING CENTERS AND
As may be apparent from my publications,
ULTRAVIOLET MICROBEAM
including this essay, I have been interested
EXPERIMENTS . . . . . . . . . . . . . . . . . . 14
in improving the capabilities of the light
SPINDLE BIREFRINGENCE,
microscope and exploring its uses as much as in
MICROTUBE DYNAMICS, AND
uncovering the submicroscopic structures and
FORCE GENERATION FOR
dynamic events taking place in the living cell.
CHROMOSOME MOVEMENT. . 17
Recently many have contributed immensely
EARLY BIOCHEMISTRY. . . . . . . . . . . . 18
to and made unbelievable advances in both
EARLY ELECTRON
of these fields (see, e.g., Howard & Hyman
MICROSCOPY . . . . . . . . . . . . . . . . . . . 20
2007, Maiato et al. 2004, Pawley 2006, Sluder
CONCLUDING REMARKS
& Wolf 2003, Wittmann et al. 2001). The
AND UPDATE . . . . . . . . . . . . . . . . . . . 21
current essay, thus, focuses on events relating
to some early developments to which I had the
1
good fortune to contribute.
INTRODUCTION
1
This Perspective emphasizes our early studies on the mitotic
Classically the light microscope was used to
spindle and microtubules but does not cover other topics that
examine preserved, thin-sectioned tissues and my colleagues and I have explored using polarized light and
other advanced modes of microscopy. Articles reporting on
cells. More recently it has become an impor-
a number of such biological studies and our contributions to
tant tool for exploring the molecular basis of
advances in microscopy are assembled in Collected Works of
physiological functions directly in active living Shinya Inoue´: Living Cells, Light Microscopy, and Molecular Dy-
namics, just published by World Scientific Publishing (Inoue´
cells. The major transition to its modern use
2008a). This book also includes a DVD disk featuring many
was prompted by optical advances in the mid-
cine´- and video-micrographs of active living cells and nar-
twentieth century, followed by another spurt in rated explanations on polarized light microscopy.
2 Inoue´
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
ENCOUNTER WITH KATIE AND
JEAN DAN: INTRODUCTION TO
LIVING CELLS
Born 1921 in London, England, as the eldest
child of a Japanese diplomat, I was brought up
together with my sisters in several countries.
I enjoyed my early school years in Portland,
Oregon, and Sydney, Australia, but from 1932
I remained in Japan to enter a municipal high
school in Tokyo.
From my early days, I was interested in fig-
uring out how to build electric motors and tiny
portable radios that actually worked, but was
not so interested in biology. My mother gave me
a small microscope, but it was so disappointing;
Figure 1
nothing in the prepared slides was doing any-
Katsuma Dan (1905–1996) at the Centennial
thing. Still, at our home, I did raise silkworms
Celebration of Misaki Marine Biological Station in
and later even collected and mounted butterflies
1987. From Inoue´ (1994).
on occasion. But in high school, the only thing
that really impressed me in biology was the be- told us about how Karl von Frisch took advan-
havior of a bird’s feather that our teacher let us tage of the sugar rationing in Germany during
examine under a loupe. The tiny, barbed hooks World War I to explore how honeybees found
allowed the feather to be ruffled yet be zippered their way back home by using polarization of
neatly back together. The image seen through a the sky light to navigate, then dance and sig-
magnifier finally explained how something ac- nal to their hive mates how to reach the nectar
tually worked! source.
My deeper interest in biology was aroused In the lab, Katie let his students try experi-
in 1941 while I was a student at Musashi Ko¯ to¯ ments that might or might not work, rather than
Gakko, a junior college in Tokyo. There I met have us follow pretested procedures. I still can-
Professor Katsuma Dan (frivolously nicknamed not forget the excitement of having been able to
Katie by himself ) in the first class that he show how Lillie’s iron wire model of nerve con-
taught in his home country (Figure 1). Katie duction worked by successive electrical depolar-
had returned to Japan in 1937 with his Amer- ization (of a passivated layer on a steel wire im-
ican wife, Jean Clark Dan, a fellow graduate mersed in concentrated nitric acid) rather than
student who had also worked with L.V. Heil- by propagation of a chemical change, as argued
brunn at the University of Pennsylvania and by my classmates. And I found that even the
with whom he had spent summers at the Marine conduction speed could be enhanced by mak-
Biological Laboratory (MBL) in Woods Hole, ing the current jump past a locally insulated seg-
Massachusetts. ment of the model [ just as Ichiji Tasaki demon-
As a student who had been unhappy with the strated the same year for saltatory conduction
high school classes in militaristic Japan, I was in myelinated nerve fibers (Tasaki & Takeuchi
shocked but delighted by Katie’s different atti- 1941)]!
tude and approach. Instead of promoting rote But that was the year the Japanese Navy at-
learning, he told us about how he and his friends tacked Pearl Harbor, and Japan and the United
were figuring out how cells divided by tracing States became embroiled in World War II.
the movement of kaolin particles placed on the Still, unlike many of my former high school
surface of developing sea urchin eggs. And he classmates, especially the A students who had
www.annualreviews.org • Microtubule Dynamics in Mitosis 3
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
become navy officers and soon perished at sea, was generally not visible under the microscope
I was deferred from military service as a science in living cells.
major (until four months before the end of the A notable exception, as Katie noted, was W.J.
war, when everybody was conscripted). Thus, I Schmidt’s 1937 observation of developing sea
was able to enter Tokyo Imperial University in urchin eggs, made with a polarizing microscope
1942 and finish the curtailed 2.5-year curricu- (Figure 2). As reinterpreted by Schmidt him-
lum with a major in zoology. self in 1939, those pictures showed the football-
One evening in 1943, Katie invited me to shaped spindles whose contrast depended on
his home in Kudan, Tokyo, to try visualizing the birefringence produced by aligned protein
the spindle during cell division. Imaging the molecules (Schmidt 1939; see Figure 5 and as-
mitotic spindle in living cells was of particu- sociated caption, below, for an explanation of
lar interest to Katie; he posited that egg cells birefringence).
divided by an elongating spindle pushing apart That evening in Tokyo, Katie fertilized clear
the two astrospheres attached to its poles (Dan eggs of sea urchins that he had brought home
1943). The problem was that the spindle itself from the marine lab in Misaki. Under air-raid
Figure 2
(Left) W.J. Schmidt’s 1937 monograph. (Right) In this monograph, on p. 89, figure 31, Schmidt shows live,
fertilized (flattened) sea urchin eggs observed between crossed Nicols with a polarizing microscope.
(a) Four-cell stage in metaphase. (b) Four-cell stage in anaphase. (c) Anaphase sperm nuclei in polyspermic
egg. (d ) Spindle and chromosomes in a blastomere at the two-cell stage. (e) Spindles and chromosomes in a
blastomere at the eight-cell stage. At the time of publication of his monograph, Schmidt interpreted the
football-shaped white and dark birefringent structures as being chromosomes and only the asters (seen in
panels d and e) as the nuclear spindle. Reproduced from Schmidt (1937) with kind permission of Gebru¨der
Borntra¨ger (http://www.borntraeger-cramer.de).
4 Inoue´
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
blackout curtains, we spent several hours trying Scope with analyzer attached
to bottom of draw tube
to see the spindle birefringence, using a polar-
izing microscope that Katie had borrowed from
his colleague in Geology. But alas, the evening Mica compensator
ended with inconclusive results.
Calcite prism polarizer
THE “SHINYA-SCOPE”
Five years later we resumed these studies, this
time at the Misaki Marine Biological Station, Tea can housing
which Katie had recovered in 1945 from the al- AH-4 Hg lamp
lied occupation forces, using his message enti-
tled “The last one to go” (reproduced in Article
56 in Inoue´ 2008a). At Misaki, rather than use
a commercial polarizing microscope, I started
from scratch by assembling parts on a cast-off
machine gun base. (The Station had been taken
Parts on wood blocks
over by the Japanese Navy for the last year of
secured by twine on discarded
the war as a miniature submarine base, so some machine gun base
destroyed weapon parts were scattered.) On the
cast-iron base, I tied by string a Zeiss micro-
scope that Katie let me modify, a calcite polar-
izing prism loaned by Professor Koana of the Shinya-Scope 1
Physics Department of Tokyo University, and (Misaki, March 1948)
an AH-4 mercury arc lamp that I found at a
Floor
surplus store and that I placed in a tea can.
Figure 3
Using this home-made instrument
The author’s handmade polarizing microscope, built from salvaged
(Figure 3), we could finally repeat Schmidt’s
components at the Misaki Marine Biological Station in 1948. Reproduced from
observations. For several minutes before the
Inoue´ (2008a) with kind permission of World Scientific Publishing.
egg underwent cleavage, we saw the birefrin-
gent spindle and asters emerge and grow, then a thin sheet and placed it on the microscope’s
the spindle splitting into two parts as the asters rotatable substage filter holder so that its ori-
grew larger. But the initial success was dashed entation could be adjusted, namely, so that it
when I tried to improve the image by rotating would act as a Brace-Koehler compensator.
the objective lens to minimize the stray light Now, even though the microscope field was
introduced by strain birefringence in the not completely dark, we could see the brighter
lenses. The field between crossed polarizers or darker football-shaped spindle against a gray
did, in fact, become darker, but where were background (Figure 4). In fact, we could see
the birefringent spindles? They had simply quite a bit more than was reported in Schmidt’s
vanished! publication and even guess at the orientation of
Katie’s admonition to me was, “I told you to the component molecules.
leave well enough alone.” But I was really curi- By way of explanation, a compensator in-
ous and wanted to make the system work better. troduces uniform birefringence over the whole
It did take a whole month, but I finally realized field of view so that, between crossed polarizers,
that the birefringence of the strained lens was, the specimen appears brighter or darker, de-
in fact, helping by acting as a compensator and pending on whether its birefringence is adding
raising the image contrast of the weakly bire- or subtracting from the birefringence of the
fringent spindle. So I split a piece of mica into compensator. As shown in Figures 4 and 5,
www.annualreviews.org • Microtubule Dynamics in Mitosis 5
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
Figure 4
(a,b) Two-cell-stage jellyfish (Spirocodon sp.) egg with metaphase spindles observed with the microscope
shown in Figure 3. In these live, optically clear cells, the positively birefringent spindle appears bright or
dark, depending on its orientation relative to the compensator (see Figure 5 for an explanation). The cell
surface, which is negatively birefringent, appears in opposite contrast where it lies parallel to the spindle.
Unlike in many other genera, eggs of jellyfish do not produce a fertilization envelope. (c,d,e) Fertilized,
developing eggs of a sand dollar, Clypeaster japonica. The Clypeaster eggs (which are exceptionally transparent)
are surrounded by a fertilization envelope, which shows a strong, tangentially positive birefringence. The
spindle and asters also show a positive birefringence along their long axes. Double-headed arrows indicate
the slow-axis direction of the compensator. From Inoue´ & Dan (1951).
where the slow axis of the specimen (e.g., the see, e.g., Bennett 1950, Appendix III in Inoue´
length of the spindle filaments) lies parallel to 1986, and Inoue´ 2002.]
the slow axis of the compensator, the specimen In the paper reporting these observations
appears brighter. Where the axes are crossed (Inoue´ & Dan 1951), I also calculated the op-
(lie in opposite quadrants), they appear darker, timum amount of compensation required to
i.e., are compensated. [Every birefringent (i.e., maximize the image contrast of weakly birefrin-
doubly refractive) material has two refractive gent objects in the presence of stray background
indexes that reflect the arrangement of their light (as also published nearly concurrently by
molecular lattice or fine structure. The direc- Swann & Mitchison in 1950).
tion for which (the electrical vector of ) the light Kayo Okazaki and Katie extensively used the
wave suffers the greatest refraction is called the microscope I built in Misaki (and sketched from
slow axis, and the one with the lowest refraction memory in approximately 1989; see Figure 3)
is called the fast axis. For further explanations, to follow the development of biocrystalline
6 Inoue´
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
skeletal spicules in sea urchin embryos (Okazaki
& Inoue´ 1976). After my departure to Princeton
in 1948, Kayo and Katie called it the “Shinya-
Scope.”
In 1948, Jean Dan returned to Misaki from
her first post–World War II trip back home to
the United States. She was full of news about
their friends in the States, especially those at
the MBL in Woods Hole. And she brought
home, as a present for her husband Katie, a
Bausch & Lomb phase contrast microscope (the
first one available in the United States and ac- 20 μm
quired courtesy of the American Philosophical
Society). Jean, who soon discovered the acroso- F
mal reaction, used this microscope extensively P A'
to study sperm-egg interactions at fertilization.
S S'
For me, she arranged a financial loan from her
A P'
sister Peggy Chittick of Milford, Connecticut,
F'
so that I could travel and study in the States.
Figure 5
Micromere formation during the fourth division in a developing egg of a sand
TO PRINCETON (1948–51) dollar, Echinarachnius parma (the images were taken with a rectified polarizing
microscope in the 1970s). (a) The spindles in these four cells have converged to
In the fall of 1948, with a postwar Japanese pass-
the egg’s vegetal pole (the four animal pole cells are out of focus). Where the
port (which I recall was number 50), I arrived at spindle long axis (orientation of microtubules) lies parallel to the compensator
′
Princeton’s Biology Department. There, while slow axis (SS ), the positively birefringent spindle appears bright. Where the
axes are crossed, the spindle appears dark. (b) Cleavage planes bisect the spindle
building what I hoped was a better polarizing
remnants and give rise to four micromeres (predecessor of spicules and gonads)
microscope, I was introduced to classical cytol-
′ ′
and four macromeres. In the diagram below the photographs, PP and AA
ogy by my mentor, Kenneth W. Cooper. Ken
show the transmission axes of the polarizer and the analyzer (which are
had studied with Franz Schrader, who in turn crossed), respectively, and SS′ and FF′ are the orientations of the slow and fast
had followed E.B. Wilson’s steps; all three were axes of the compensator, respectively. Reproduced from Inoue´ (1981) with kind
permission of The Rockefeller University Press.
at Columbia University in New York. It was,
therefore, natural for me to wonder how chro-
mosomes moved in mitosis and about the enig- may not really pre-exist approximately as such
matic properties of the mitotic spindle. in the living cell” and cautions us not to “pre-
At Princeton I was exposed to Wilson’s clas- maturely condemn a theory which may yet be
sical volume on the hereditary role of chro- reconcilable with the so-called dynamical the-
mosomes and the structure and function of ories” (Chapter II, Section IV, The Mechanism
the mitotic spindle in cell division (Wilson of Mitosis, in Wilson 1928).
1928). Summarizing four-decades-long studies The more physicochemically oriented pro-
on fixed and stained cells made by many cytol- ponents of the dynamical theory experimented
ogists, he describes the fibrillar structure of the with living cells and tended to be skeptical of the
achromatic spindle and astral rays. Still, he is existence of the fibrous elements of the spindle
puzzled about the ephemeral nature and invis- and asters. These investigators considered such
ibility of the spindle fibrils in living cells and elements to be artifacts of fixation.
questions the validity of the contractile fibril- In a 1929 article, Karl Be˘larˇ compared
lar hypothesis for chromosome movement fa- the behavior of live grasshopper spermatocytes
vored by many. At the same time, he is reluc- with that of carefully fixed and stained cells
tant to accept “that the fibrillae seen in sections (Figure 6). Although unable to see any spindle
www.annualreviews.org • Microtubule Dynamics in Mitosis 7
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
up not of coherent filaments but of short thin
rods, as in a liquid crystal (Wada 1950). Simi-
larly, observing the migration of chromosomes
that appeared to cut right through kinetochore
¨
fibers, Gunnar Ostergren (1949) also favored
the liquid crystalline nature of spindle fibers.
These and other views in the early 1950s
on the physical nature of the mitotic spin-
dle, as well as various proposals on how chro-
mosomes move in mitosis, are summarized in
Franz Schrader’s monograph Mitosis: The Move-
ment of Chromosomes in Cell Division (Schrader
1953). In this volume, he points out two cases in
which spindle fibers were actually observed in
intact dividing cells. These observations were
made by L. R. Cleveland (1938) in Barbu-
lanympha, a symbiotic protozoan in the wood-
eating cockroach Cryptocercus, and by Kenneth
Cooper (1941) in the eggs of a grass mite,
Pediculopsis graminum. Still, Schrader points out
that these were exceptional cases and could not
be taken to represent cells undergoing mitosis
generally. Thus, the reality of spindle fibers and
their nature remained major unresolved issues.
While I was at Princeton, we also saw
fascinating movies of dividing grasshopper
spermatocytes, filmed by Kurt Michelle of
Karl Zeiss, using its phase contrast micro-
scope. Subsequently, Kyojiro Shimakura cap-
tured higher-resolution images of similar live
cells (Figure 7), and Andrew and Wishia Bajer
(1951, 1956) made many films of dividing
Figure 6
endosperm cells of the African blood lily
Spermatocytes of a grasshopper, Chorthippus lineatus, at different stages of
Haemanthus katherinae (Figure 8; see Sup-
meiosis I. The drawings were by Beˇlarˇ (1929) of 6-μm sections of cells fixed in
Flemming-Meves solution, stained with iron-hematoxilin. mi, mitochondria; plemental Movie 1; for all supplemental
x, sex chromosome; H, heteromorph tetrad.
material, follow the Supplemental Material
link from the Annual Reviews home page at
structure, he observed in healthy, live cells http://www.annualreviews.org). In these di-
Brownian motion preferentially along the di- viding cells, the phase contrast microscope dis-
rection of fibrils that would appear after fix- played the movement and shape change of chro-
Supplemental Material ation. Also, from the distortion of live cells mosomes most strikingly.
treated with hyperosmotic media, he concluded The phase contrast microscope accentuates
that spindle fibrils or some longitudinal lamel- the image contrast of those bodies whose refrac-
lar material must exist in the living cell despite tive indexes are somewhat greater or less than
their invisibility. the refractive indexes of their surroundings.
By observing chromosome movement in di- In contrast to the chromosomes themselves,
viding stamen hair cells of Tradescantia, Bungo the spindle fibers that were supposed to move
Wada proposed that spindle fibers were made the chromosomes, and the fibrils laying down
8 Inoue´
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
ANRV356-CB24-01 ARI 27 September 2008 2:16
the cell plate in plant cells, were not visible in
phase contrast.
In these early post–World War II years, the
electron microscope also started to reveal many
important cellular fine structures (see Sabatini
2005). But little could be seen of the fine struc-
ture in the spindle until glutaraldehyde fixa-
tion was introduced nearly two decades later
(Sabatini et al. 1963).
Thus, the challenge for me in the late 1940s
was to develop a polarizing microscope that
had enough sensitivity and image resolution to
show what, in fact, was going on inside dividing,
living cells.
THE PRINCETON MICROSCOPE
At Princeton, I decided to start from scratch
again so that I could improve on the microscope
that I had built at Misaki. By then I was more
aware of the standard use of polarizing (or pet-
rographic) microscopes to study crystals and to Figure 7
identify minerals and ores (e.g., Hartshorne & Phase contrast image of a live spermatocyte in a grasshopper, Chloealtis
Stuart 1960, Rinne & Bereck 1953, Wahlstrom genicularibus, at full metaphase. Photograph courtesy of Dr. Kyojiro Shimakura
of Hokkaido University. Reproduced from Inoue´ (1964) with kind permission
1960, Wright 1911). Biologists also used these
of Elsevier.
microscopes to study mineralized tissue, skele-
tal muscle, plant cellulose walls, etc., which
were all highly birefringent (see, e.g., Ambronn gence, and polarizers and analyzers providing
& Frey 1926, Bennett 1950, Frey-Wyssling very high extinction, was not enough. The crit-
1953, Schmidt 1924). Schmidt (1937) also ex- ical factor turned out to be the very fact that
plored an extensive array of cellular com- microscope lenses had to refract light to form
ponents and cell products, many with much an image. And the greater the angle of refrac-
weaker birefringence, as summarized in this tion (and high-NA lenses are characterized by
second monograph. The commercially avail- high angles of refraction), the greater is the loss
able polarizing microscopes were, however, not of extinction and, therefore, loss of sensitivity
optimally designed for observing or measuring to detect weak birefringence.
the intricately organized, and very weakly bire- I examined this paradox in detail at Prince-
fringent, minute organelles in living cells. ton (Inoue´ 1952b) but had no basic solution, so
It turned out that there was an inherent I went ahead and built my second polarizing mi-
incompatibility between achieving high sensi- croscope, using the best arrangement and com-
tivity for detecting weak birefringence and for ponents available to me at that time (includ-
gaining image resolution high enough to study ing strain-free objectives selected from several
structural details inside a living cell. At low con- hundred by Bausch & Lomb, a much brighter
denser and objective lens numerical apertures AH-6 water-cooled high-pressure mercury arc
(NAs), one could achieve high extinction and lamp, a Leitz photo stand, etc.). The resulting
gain great sensitivity, but then the resolution microscope is illustrated in Figure 9.
was limited. Selecting objective and condenser With this microscope, I found that I could
lenses with exceptionally low strain birefrin- indeed gain moderately high resolution (if not
www.annualreviews.org • Microtubule Dynamics in Mitosis 9
Annu. Rev. Cell Dev. Biol. 2008.24:1-28. Downloaded from arjournals.annualreviews.org
by University of California - Davis on 01/20/09. For personal use only.
