170
5.
SECTIONING
5.18, Mounting the Knife
It might seem that mounting the knife in the microtome hardly deserves
a section in this book, yet the proper knife tilt is a most important thing
for successful sectioning and care must be taken in determining it. Also,
insofar as possible, it is important to recognize the symptoms of improper
tilt so that suitable corrections can be made while sectioning is in
progress.
A slight tilt is necessary; about 3 to 5° seems best (Fig. 18).
Too much tilt encourages "chattering" (Chapter 5.22) which is the
technical bugaboo of ultrathin microtomy, for the chatter is often on
such a microscopic scale that it is not visible while sections are being
cut. It is disappointing to discover when looking with the electron microscope that seemingly well cut sections are quite useless for this reason.
Too much tilt probably promotes chatter because the knife edge tends
to dig into the block, and then to bounce back when elastic forces
become great enough. One should recall how flexible thin glass is as we
experience it in a fine capillary or in glass wool. It is a wonder indeed
that the very fine edges of glass knives can have enough support to cut at
all. They gain maximum support when the front face is as nearly vertical
as possible, and the main mass of the glass can absorb the vertical thrust.
Although one can be deluded into thinking that he is sectioning well
when the knife is tilted too much, no such illusion is possible when the
knife is leaning slightly backwards. Then the block coming down misses
the edge and rubs against the back of the knife. This compresses the
block a little and the same thing happens again on the next cycle. Thus,
no sections at all are cut for a number of cycles of the microtome, but
then suddenly a thick section is cut when the block recoil catches up with
the additive effects of compression, and the knife digs in. Thus, when this
is happening, there is excessive skipping and occasional thick sections.
Also, one can sometimes see the block rubbing on the glass by looking
through the acetone and through the glass to the front face of the knife.
Where the block makes contact with the glass the reflectivity of the glass
surface is altered and a black spot may be visible.
In almost all microtomes it is also necessary to adjust the height of the
knife fairly exactly so that the block passes the knife during a particular
part of its cycle. How then is one to determine the correct angle and
height? Some of the new microtomes have rather elaborate knife-holding
5.
SECTIONING
5.18, Mounting the Knife
It might seem that mounting the knife in the microtome hardly deserves
a section in this book, yet the proper knife tilt is a most important thing
for successful sectioning and care must be taken in determining it. Also,
insofar as possible, it is important to recognize the symptoms of improper
tilt so that suitable corrections can be made while sectioning is in
progress.
A slight tilt is necessary; about 3 to 5° seems best (Fig. 18).
Too much tilt encourages "chattering" (Chapter 5.22) which is the
technical bugaboo of ultrathin microtomy, for the chatter is often on
such a microscopic scale that it is not visible while sections are being
cut. It is disappointing to discover when looking with the electron microscope that seemingly well cut sections are quite useless for this reason.
Too much tilt probably promotes chatter because the knife edge tends
to dig into the block, and then to bounce back when elastic forces
become great enough. One should recall how flexible thin glass is as we
experience it in a fine capillary or in glass wool. It is a wonder indeed
that the very fine edges of glass knives can have enough support to cut at
all. They gain maximum support when the front face is as nearly vertical
as possible, and the main mass of the glass can absorb the vertical thrust.
Although one can be deluded into thinking that he is sectioning well
when the knife is tilted too much, no such illusion is possible when the
knife is leaning slightly backwards. Then the block coming down misses
the edge and rubs against the back of the knife. This compresses the
block a little and the same thing happens again on the next cycle. Thus,
no sections at all are cut for a number of cycles of the microtome, but
then suddenly a thick section is cut when the block recoil catches up with
the additive effects of compression, and the knife digs in. Thus, when this
is happening, there is excessive skipping and occasional thick sections.
Also, one can sometimes see the block rubbing on the glass by looking
through the acetone and through the glass to the front face of the knife.
Where the block makes contact with the glass the reflectivity of the glass
surface is altered and a black spot may be visible.
In almost all microtomes it is also necessary to adjust the height of the
knife fairly exactly so that the block passes the knife during a particular
part of its cycle. How then is one to determine the correct angle and
height? Some of the new microtomes have rather elaborate knife-holding