A. Autotechnicon / Tissue Processing: Discuss The Principles and Concepts of The Following

Discuss the principles and concepts of the following:
a. Autotechnicon / Tissue Processing
Autotechnicon is making the sample process painless and quickly,

where tissue processing is a step that require taking the tissue of an
animal or human from the fixation into the state where it completely
infiltrated and can be ready for the section cutting on microtome.
The various solutions used for processing tissue are placed into ten
separate nylon beakers. These beakers are placed on the circular deck
of the instrument. There are also two paraffin baths that are mounted on
the deck. Each paraffin bath contains an internal heater that is
thermostatically controlled.
Tissue processing is concerned with the diffusion of various substances
into and out of porous tissues. Diffusion results from the tendency of
processing reagents to equalize concentrations both inside and outside
blocks of tissue. The reagent molecules diffuse down a concentration
gradient and move from where they are at a high concentration to where
they are at a lower concentration. This movement requires no energy
since it always progresses down the concentration gradient.
PRINCIPLES OF TISSUE PROCESSING
Vacuum, it removes the reagents from the tissue but only if they are more volatile
than the reagents that being replaced. It is also used on the automated processor
to prevent deterioration and damage to the tissue
Fixation, it stabilized the proteins rendering the cells and its component resistant
to the further autolysis by the inactivating the lysosome enzymes.
Post-fixation treatment, this include the special fixation techniques that may require
additional steps before the process are initiated.
Heat, this increases the rate of exchange fluid and penetration and must be used
to reduce the possibility of shrinkage, hardening or embrittlement of the tissue
sample.
Viscosity, it is the property of resistance of the flow of fluid wherein, the smaller
size of the molecules in the solution has the faster rate of the fluid penetration (low
viscosity) and the larger the size in the solution has the slower rate of the fluid
penetration (high viscosity).
Factors influencing the rate of processing, it occurs when the tissue is immersed
in the fluid.
Agitation, this increases the flow of the fresh solutions around in the tissue.
b. Immunofluorescence
Immunofluorescence is an assay which is used primarily on biological samples and

is classically defined as a procedure to detect antigens in cellular contexts using
antibodies. The specificity of antibodies to their antigen is the base
for immunofluorescence. It also uses fluorescently labeled secondary antibodies
to visualize proteins in cells and tissues and can provide information about the
tissue distribution of a given protein as well as its subcellular distribution.
THE THREE CONCEPT
1. Immunohistochemistry is an assay for confirming the expression and

expression location of proteins in nature tissue section.
2. Immunocytochemistry is performed on sample of intact cells.
3. Immunofluorescence is an assay which is used primarily on biological

samples and is classically defined as a procedure to detect antigens in
cellular contexts using antibodies. Immunofluorescence has been widely
used in biological research and medical research yield and becomes one
most important and effective method. There are two different
immunofluorescence assay which include indirect immunofluorescence
assay and direct immunofluorescence assay.
c. Frozen Section / Cryostat
The frozen section is the rapid tissue section by cooling the tissue
with the help of cryostat to provide immediate report of the tissue
sample.
The cryostat is the instrument to freeze the tissue and also to cut
the frozen tissue for microscopic section. The rapid freezing of the
tissue sample converts the water into ice. The firm ice within the
tissue acts as embedding media to cut the tissue. A cryostat has a
cryo-chamber that can hold up to 10 specimen disks. It also has a
stainless-steel rotary microtome that can cut specimens in a
specified thickness level.
The frozen section is mainly used for rapid diagnosis of the lesion
for intraoperative management, to know the extent of the lesion, to
do enzyme immunocytochemistry and immunofluorescence study
and also to stain lipid and certain carbohydrate in the tissue.
d. Electron Microscope
An electron microscope is a microscope that uses a beam of

accelerated electrons as a source of illumination.
It is a special type of microscope having a high resolution of images, able to
magnify objects in nanometers, which are formed by controlled use of
electrons in vacuum captured on a phosphorescent screen.
TWO TYPES OF ELECTRON MICROSCOPE
1. The transmission electron microscope (TEM)

The transmission electron microscope is used to view thin specimens through
which electrons can pass generating a projection image.
The TEM is analogous in many ways to the conventional (compound) light
microscope.
TEM is used, among other things, to image the interior of cells (in thin sections),
the structure of protein molecules (contrasted by metal shadowing), the
organization of molecules in viruses and cytoskeletal filaments (prepared by
the negative staining technique), and the arrangement of protein molecules in
cell membranes (by freeze-fracture).
2. The scanning electron microscope (SEM)

Conventional scanning electron microscopy depends on the emission of
secondary electrons from the surface of a specimen.
Because of its great depth of focus, a scanning electron microscope is the EM
analog of a stereo light microscope.
It provides detailed images of the surfaces of cells and whole organisms that
are not possible by TEM. It can also be used for particle counting and size
determination, and for process control.

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Discuss the principles and concepts of the following:

a. Autotechnicon / Tissue Processing

Autotechnicon is making the sample process painless and quickly,

PRINCIPLES OF TISSUE PROCESSING

Immunofluorescence is an assay which is used primarily on biological samples and

THE THREE CONCEPT

1. Immunohistochemistry is an assay for confirming the expression and

2. Immunocytochemistry is performed on sample of intact cells.

3. Immunofluorescence is an assay which is used primarily on biological

c. Frozen Section / Cryostat

An electron microscope is a microscope that uses a beam of

TWO TYPES OF ELECTRON MICROSCOPE

1. The transmission electron microscope (TEM)

2. The scanning electron microscope (SEM)

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