Page 54 - Atlas of Histology with Functional Correlations

Frozen Section Procedures

Besides paraffin sections, there are also procedures in which the tissues are first
frozen for rapid microscopic examination of a specimen that may be taken
during surgery. This type of procedure is called cryosection; however, frozen

sections are of lesser quality than those fixed in formalin and embedded in
paraffin. The instrument for cryosectioning is the cryostat, which is a microtome
inside a freezer. The tissue is removed from an organism, placed onto a metal
stub, embedded in a gellike medium, and frozen to about −20°C to −30°C; such
low temperatures are required for fat or lipid-rich tissue. The specimen is

secured in a chuck and is cut frozen on the microtome 5 to 10 μm thick.
Individual or single frozen sections are cut at low temperature, picked up and put
on a glass slide, and then stained. The freezing of tissue eliminates the need for

chemical fixation, is faster, and maintains most enzyme and immunological
functions. It also can be used to examine temperature-sensitive or lipid-soluble
molecules or where rapid analysis of the tissue is needed. Sectioning with the
cryostat is similar to that of paraffin sectioning; both methods use a rotary type
of microtome.

In addition to rotary microtomes, there are nonrotary or sliding or sledge
microtomes. In these units, the tissue sample is put into a holder, which then is
moved back and forth across the knife. The sledge or sliding microtome is
primarily used for cutting large samples of tissues or organs embedded in

paraffin, such as sections of the brain, kidneys, and other biological structures
for histological examinations. Typical section thickness produced by a sledge
microtome is between 1 and 60 μm. After sectioning, the samples undergo
routine preparation for staining with different type of stains.

Transmission and Scanning Electron Microscopy

Examining the tissue sections with a transmission electron microscope (TEM)
allows for much higher magnification and greater resolution. The fixatives and

procedures are different from those of tissue preparation for histological slide
examination. The specimen that is to be collected is either previously perfused
with the fixative in the body or removed from the organism, cut into small
pieces, and directly immersed in the fixative for rapid fixation. In addition, the

primary fixative for TEM specimens is cold-buffered glutaraldehyde in which

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