Dehydration, embedding, and sectioning with a microtome are crucial steps in preparing tissue samples for microscopic examination. Dehydration removes water from the tissue, followed by embedding in a medium like paraffin wax to provide support for sectioning. The microtome is then used to cut thin slices (sections) of the embedded tissue, which are subsequently mounted on slides and stained for viewing under a microscope.

1. Dehydration and Embedding
Section with Microtome

2.  Dehydration
• Dehydration is a crucial step in the preparation of
permanent microscope slides. It’s essential for removing
water from biological specimens, which allows for
proper clearing and mounting.
• Purpose of Dehydration:
• Prevent Decay: Water promotes bacterial
growth and decomposition, which would
degrade the specimen.

3. This stepwise approach prevents excessive distortion or shrinkage of
the tissue that could occur if it were abruptly placed in pure alcohol.
• Typical steps might include: 70% ethanol, 95% ethanol, and multiple
changes of 100% ethanol.
• Time:
• The duration of immersion in each alcohol
solution varies depending on the size and type of
specimen.
• Generally, several minutes are spent in each
solution to ensure complete dehydration.

4. Important Considerations:
It is very important to use fresh, high-quality ethanol,
especially for the final dehydration steps.
The time spent in each solution must be controlled.
The solutions must be kept clean.
Following Dehydration:
After dehydration, the specimen undergoes:
• Clearing: The alcohol is replaced with a clearing agent (e.g., xylene),
which makes the tissue transparent.

5. Embedding and sectioning with a microtome are essential techniques in
histology and pathology, allowing for the preparation of thin tissue
slices for microscopic examination. Here’s a breakdown of the process:
1. Embedding:
Purpose:
• Embedding provides support to the delicate tissue,
allowing for the creation of thin, uniform sections.
• Paraffin wax is the most commonly used embedding medium.
 Embedding and
sectioning with microtome

6. Process:
After dehydration and clearing, the
tissue is infiltrated with molten paraffin
wax.
The tissue is then placed in a mold, and
more molten paraffin is added to create
a solid block.
• The paraffin block is allowed to
solidify, encasing the tissue.

7. 2. Sectioning with a Microtome:
Purpose:
• The microtome is a precision
instrument that cuts extremely thin
sections of the embedded tissue.
• Process:
• The paraffin block is mounted in the
microtome.
• A sharp microtome blade is used to
slice thin sections, typically ranging
from 3 to 10 micrometers in thickness.
• The sections form a ribbon of paraffin,
which is carefully collected.

8. Microtome Blade Sharpness: A sharp blade is crucial for
obtaining high-quality sections.
• Section Thickness:
• The desired section thickness depends on the specific
application.
• Water Bath Temperature: The water bath temperature must
be carefully controlled to prevent damage to the tissue.
• Safety:
• Microtome blades are extremely sharp, so proper safety
precautions must be taken.

9. • Sass.J.E.(1958) Botanical micro technique,The Towa State
College Press.
• https://nios.ac.in
 Reference