Pathologists study patient slides carefully
to make their assessments with the aide of sophisticated
microscopes. Histology is the science dealing with the
microscopic identification of cells and tissue. Before
the pathologist can begin this process, a slide has
to be made. The following guide will take you through
the steps required to produce a slide.
Tissues must be immersed in fixative immediately
after removal from the body to prevent decomposition,
autolysis and artifact formation. 10% Neutral
Buffered Formalin is the routine fixative.
||1. Specimen Accessioning
Upon verification that the specimen was received
in the proper fixative, the specimen is then assigned
an accession number. This accession number consists
of a letter-number code used to identify the specimen
throughout the complete process.
2. Gross Specimen Examination
A pathologist, pathologist assistant or resident
visually examines the specimen. A description
of the specimen is recorded and the specimen is
then cut into the proper size and placed into
a color-coded cassette labeled with the assigned
||Inking a gross specimen for margins
(not shown) Different color tattoo inks
are used to identify margins or other structures
After the specimens are macroscopically examined
and placed into the color-coded cassettes, the
cassettes are then loaded into baskets that are
placed into a tissue processor. Tissue processing
consists of four steps: Fixation, Dehydration,
Clearing and Infiltration lasting between 8 to
A tissue processor is a machine that automatically
transfers one chemical to the next inside the cassette
Properly oriented and arranged tissues are cast
into liquified paraffin that hardens upon cooling.
Enclosing the tissue in the infiltration medium
(paraffin) used for processing and then allowing
the medium to solidify.
with a microtome
The microtome advances the tissue block, toward
a sharp knife. The tissue is moved in an up and
down motion to cut the tissue into a preset number
of micrometers. The paraffin and tissue shavings
are shaved off like a ribbon.
||The ribbon of tissue
is then carefully placed into a water bath. The
water bath is a dish of heated, distilled water.
The paraffin ribbon floats on top of the water.
on glass slide
Placing a positively charged slide under the section
and lifting up picks up the tissue section. The
tissue adheres to the charged slide
||5. Drying Oven
The unstained slides are then placed into a metal
tray that gets placed into a drying oven. The oven
is warm and helps the section of tissue adhere to
||6. Tissue Staining
When the slides are removed from the oven, they
are placed into an automated staining machine. The
slides with the tissue on them are immersed in chemicals
and dyes that stain the cells. By applying the colored
dyes to the cells, the pathologists are able to
view the structure of the tissue better.
||H & E Staining
Hematoxylin and Eosin staining is the routine stain
for all tissues. Hematoxylin stains the nucleus
and the Eosin stains the cytoplasm of tissue cells.
A protective glass coverslip is
attached to the slide with mounting medium. This
protects the tissue from being scratched. Better
microscopic examination at various magnifications
is also obtained by the use of coverslips.
When rapid diagnosis is required, the pathologist
may need to perform a frozen section. A frozen section
is cut using a cryostat. A cryostat is a refrigerated
unit that has a microtome. The tissue is frozen
solid enough to be sectioned by the microtome and
put directly onto a slide for H & E staining.
Frozen sections are stained by hand.