5.22.
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177
other, leaving the acute region for quality sectioning later. If a good
portion of the knife edge is used for moderate facing of the block, and
this is done with delicacy and without taking too large a number of
sections, it may be possible to continue with ultrathin sectioning without
moving the knife.
Coarse and fine adjustments allow the knife to be brought up to the
block face. The perspective with which one views the block and knife is
such that it is not always easy to see just how close the knife is to the
block face. Often, however, the light source can be adjusted so that the
knife edge will cast a thin dark shadow on the block face if the latter is
dropped a little below the edge of the knife. The narrowing shadow
observed as one manipulates the controls indicates the closing gap
between the knife and block. Under no circumstances should the knife
actually be allowed to touch the block, for most of the time this will so
contaminate that portion of the edge that it must be abandoned.
When the block and knife are as close together as one dares place them
further advance is made by cycling the microtome while still advancing the
knife by small adjustments. Eventually the microtome will start cutting
"thick" sections, although the advance should be so cautious that these
"facing" sections should be only about a micron thick. If thicker than this
the knife may dig and gouge, or chip out portions of the block.
The most convenient time to fill the trough is just after the knife has
made contact with the block. The acetone solution is added with a pipette
until the edge is wet. So long as thick facing sections are to be cut, it is
best to keep the meniscus low to minimize the danger of wetting the block
face.
When final facing is completed and one is ready to begin thin sectioning it is usually desirable to get rid of the thick sections floating around
in the trough, and to clean the surface and adjust the level of the acetone
in preparation for the ultrathin sections. This is done with a glass spatula
previously wet with the acetone solution, in a manner described in detail
in Chapter 5.26.
If we assume that there has been an interruption to clean the acetone
surface, thin sectioning is then begun by first taking one or two slightly
thick sections by delicate manipulation of the fine adjustment of the knife
holder. Then one continues on, simply cycling the microtome. Success
will depend in part upon factors already considered: (1) a clean knife,
(2) a proper tilt, (3) a meniscus at the proper level, and (4) a sufficiently
small block, one half the size that the novice hopes to cut. Success also
SECTIONING
177
other, leaving the acute region for quality sectioning later. If a good
portion of the knife edge is used for moderate facing of the block, and
this is done with delicacy and without taking too large a number of
sections, it may be possible to continue with ultrathin sectioning without
moving the knife.
Coarse and fine adjustments allow the knife to be brought up to the
block face. The perspective with which one views the block and knife is
such that it is not always easy to see just how close the knife is to the
block face. Often, however, the light source can be adjusted so that the
knife edge will cast a thin dark shadow on the block face if the latter is
dropped a little below the edge of the knife. The narrowing shadow
observed as one manipulates the controls indicates the closing gap
between the knife and block. Under no circumstances should the knife
actually be allowed to touch the block, for most of the time this will so
contaminate that portion of the edge that it must be abandoned.
When the block and knife are as close together as one dares place them
further advance is made by cycling the microtome while still advancing the
knife by small adjustments. Eventually the microtome will start cutting
"thick" sections, although the advance should be so cautious that these
"facing" sections should be only about a micron thick. If thicker than this
the knife may dig and gouge, or chip out portions of the block.
The most convenient time to fill the trough is just after the knife has
made contact with the block. The acetone solution is added with a pipette
until the edge is wet. So long as thick facing sections are to be cut, it is
best to keep the meniscus low to minimize the danger of wetting the block
face.
When final facing is completed and one is ready to begin thin sectioning it is usually desirable to get rid of the thick sections floating around
in the trough, and to clean the surface and adjust the level of the acetone
in preparation for the ultrathin sections. This is done with a glass spatula
previously wet with the acetone solution, in a manner described in detail
in Chapter 5.26.
If we assume that there has been an interruption to clean the acetone
surface, thin sectioning is then begun by first taking one or two slightly
thick sections by delicate manipulation of the fine adjustment of the knife
holder. Then one continues on, simply cycling the microtome. Success
will depend in part upon factors already considered: (1) a clean knife,
(2) a proper tilt, (3) a meniscus at the proper level, and (4) a sufficiently
small block, one half the size that the novice hopes to cut. Success also