PATHOLOGY
SHIFT 1
TISSUE PROCESSING, MICROTOMY,
AND STAINING
24 AUG 2022
AY 2022-2023
Dra. Celestine Marie G. Trinidad, MD, DPSP
03
TABLE OF CONTENTS
Tissue Processing Methods…………….………………1
1.1. Producing Small Tissue Sections
1.2. Methods of Tissue Processing
2. Steps in Tissue Processing.……………………….……2
2.1. Fixation
2.2. Dehydration
2.3. Clearing
2.4. Infiltration
3. Factors Affecting Tissue Processing……..……..……4
3.1. Tissue Density and Thickness
3.2. Agitation
3.3. Temperature
3.4. Pressure
3.5. Vacuum
3.6. Other Technical Considerations
4. Embedding/Casting/Blocking….………………….…….1
4.1. Embedding Molds
4.2. Embedding Machine
4.3. Steps in Embedding
5. Sectioning/Microtomy………..……..…………........……4
5.1. Parts of a Microtome
5.2. Steps in Sectioning
5.3. Artifacts
6. Staining……………………………..………….……….......6
6.1. Types of Stains
7. Mounting of Tissue Sections........................................7
7.1. Steps in Mounting
8. Labeling..........................................................................8
9. Stains for Cellular Components and Contents...........8
9.1. Periodic Acid Schiff (PAS) Stain
9.2. Masson’s Trichrome
9.3. Alcian Blue Stain
9.4. Reticulin Stain
9.5. Perl’s Prussian Blue
9.6. Lipid Stains
9.7. Amyloid Sstains
10. Stains for Organisms....................................................9
10.1. PAS Stain
10.2. Grocott-Gomori Methenamine Silver
10.3. Giemsa Stain
10.4. Warthin Starry
10.5. Gram Stain
10.6. Mycobacterial Stain (Acid Fast Stain)
1.
Fig 1. Tissue cassette
(Source: Instructor’s slide)
•
Fig 2. Paraffin blocks
(Source: Instructor’s slide)
Fig 3. Smaller sections on a glass slide
(Source: Instructor’s slide)
•
LEGEND
Take note
1.
Textbook
From Prof
Previous Trans
TISSUE PROCESSING METHODS MABB; ADG
1.1 PRODUCING SMALL TISSUE SECTIONS MABB
• After placing samples of tissue in tissue cassettes, we
turn them into even smaller sections that we can place
on a glass slide and view under the microscope.
.
We embed these tissues in paraffin blocks.
o These tissues have to be embedded in a solid
medium firm enough to support the tissue,
give it enough rigidity to be cut, and yet soft
enough not to damage the knife or the tissue
itself.
•
Tissue Processing
o Various steps to produce tissue sections
o Tissue must be adequately prepared and set
in a paraffin block ready for cutting
o Ensures that the tissue is firm enough and
completely embedded for sectioning
o Good quality tissue sections ensure adequate
interpretation of microscopic cellular changes
Steps in Tissue Processing
o Fixation
o Dehydration
o Clearing
o Infiltration
o Embedding
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PATHOLOGY
SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
1.2. METHODS OF TISSUE PROCESSING MABB; ADG
1.2.1 Manual Method
• For small batches of tissue
• You do everything by hand
• Convenient backup in the event of a power interruption
or breakdown of the tissue processor
4
5
FLUID TRANSFER
PROCESSOR
Fig 5. Tissue Transfer
Processor
Fig 6. Fluid Transfer Processor
“Dip-and dunk” machine
Also known as Enclosed
tissue processor
Cassettes stay in a single
chamber and fluids are
pumped in and out as
required
Specimens do not dry out
within the chamber
Reagent vapors are
vented through filters or
retained in a closed-loop
system
Preferably used
2. STEPS IN TISSUE PROCESSING MABB
Table 2. Sample Processing Schedule
Most tissue processors follow this one.
STATION
1
2
3
REAGENT
10% neutral
buffered formalin
70% alcohol
80% alcohol
1 hour
1 hour
1 hour
1 hour
Clearing
Infiltration
1 hour
1 hour
1 hour
1 hour
1 h
o
u
r
12 STATIONS lasting 1 hour each
Each station has a corresponding reagent that
corresponds to a particular step in tissue processing
“What I want you to remember from this slide [Table
1] is that the entire process takes 12 HOURS. So you
DON’T instantly get a paraffin block right after you
submit a specimen. Processing takes time. It would
be some time before you get the block and slides as
well.” (Trinidad, 2022)
Table 1. Types of Automated Tissue Processors
Specimen-containing
cassettes are moved
from one container to
another
Paraffin wax
7
•
•
TISSUE TRANSFER
PROCESSOR
12
6
1.2.2 Automated Method
• Using a tissue processor
• Method used by most laboratories
Fig 4. Tissue processor
(Source: Instructor’s slide)
8
9
10
11
95% alcohol
95% alcohol
100% (absolute)
alcohol
100% (absolute)
alcohol
Xylene
Xylene
Xylene
Paraffin wax
PURPOSE
TIME
Fixation
1 hour
Dehydration
1 hour
1 hour
2.1. FIXATION MABB; ADG
• Done even before submitting it to the histopathology
laboratory
• Also part of tissue processing
o 1st step
o 1 hour in 10% neutral buffered formalin
• Alteration of tissues by stabilizing proteins so that
the tissues become resistant to change
• Essentially kill the tissues
o Stop all metabolic processes
2.1.1. Fixative of Choice
• 10% Neutral Buffered Formalin
• Change the soluble contents of the cells into insoluble
structures
• Prevent autolysis (decay of tissue due to the action of
cellular enzymes) and putrefaction (bacterial attack)
• Stabilize structures to maintain the proper relationship
of cells and their stroma
2.1.2. Role of Fixation in Tissue Processing
• Allows thin sectioning of tissues by hardening tissue
• Protects tissue from the subsequent processing steps
• Improves cell avidity for special stains
2.1.3. Must Dos in Fixation
• Suboptimal/ delayed fixation = Irreversible tissue
damage and damage of sections
• Tissue should be fixed in a sufficient volume of
solution (10:1 to 20:1)
• Fixatives diluted and/or contaminated by bodily fluids
must be replaced to ensure effectiveness
o Quality control of tissue processing
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PATHOLOGY
SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
•
Fixation time
o Time tissue is exposed to formalin
o Acceptable range: 6 to 48 hours
o Failure to meet this range adversely affects
staining qualities
o Overfixation hardens the tissue and makes it
difficult to cut
2.2. DEHYDRATION MABB; ADG
• Takes about 6 hours (Stations 2 to 7)
2.2.1. Definition
Process of removing intracellular and extracellular
water from the tissue following fixation and prior to wax
impregnation
• Most of the water in a specimen must be removed
before it can be infiltrated with wax
o Because wax does not mix with water
o Wax cannot infiltrate inside a specimen if it still
has water
o To help remember the concept:
o Liquids: “Like dissolves like”
o Miscible: polar liquids dissolve in
polar
liquids;
non-polar
liquids
dissolve in non-polar liquids
o Wax is a non-polar substance and
water is a polar substance
•
2.2.2. Reagent For Dehydration Step
• Alcohol (Ethyl alcohol/ Ethanol)
• Recommended for routine dehydration of tissues
• Fast-acting, mixes with water and many organic
solvents
• Penetrates tissues easily
• Not poisonous and not very expensive
2.2.3. Dehydration Process
• Involve slow substitution of the water in the tissue
with the organic solvent
• Entails immersing the specimen in a series of ethanol
solutions in graded concentration
o 70% → 90% → 100% (absolute alcohol)
o Graded in order to minimize some shrinkage
and extraction of cell components
• Some components are drawn out alongside water:
o Water soluble proteins: removed at lower
alcohol concentrations
o Certain lipids may be dissolved: 100%
alcohol concentration
o Fatty tissues esp. those with benign
neoplasms of fat (lipoma): acetone or
isopropanol should be added first before the
final absolute ethanol
▪ 100% (absolute alcohol) ensures complete
dehydration
• Volume of ethanol: should not be less than 10x the
volume of tissue
o To ensure complete penetration of the tissue
2.3. CLEARING CMB; ADG
• A.k.a. de-alcoholization
• Takes 3 hours to complete; 3 stations (Stations 8, 9,
10)
o All use the same reagent (Xylene) in 3
different changes or batches
o Is new in each station to completely clear the
specimen
• After dehydration, tissues are water-free, but still
cannot be infiltrated with wax
o
wax and ethanol are immiscible
 Ethanol is polar
 Wax is non-polar
• Intermediate solvent that is fully miscible with both
the ethanol and wax is needed to remove alcohol and
other dehydrating solutions from tissues prior to
embedding
o Is of intermediate polarity that can mix with
both ethanol and wax
2.3.1. Definition
• Process whereby alcohol or dehydrating agent is
removed from the tissue and replaced with a
substance that will dissolve in the wax
• Removes a substantial amount of fat from the tissue to
ensure a complete wax infiltration (aside from some
fatty tissue already removed during dehydration)
o Clearing removes most of the fat from the
tissue
2.3.2. Reagent For Clearing Step
• Xylene
o Colorless clearing agent that is most
commonly used in histopathology
o Cost-effective
• Tissue is immersed in 1-3 different changes of xylene
solutions to gradually replace the ethanol with xylene
o Xylene dissolves alcohol and replaces it in the
tissue
2.4. INFILTRATION CMB; ADG
• A.k.a. impregnation
• Takes 2 hours in 2 stations
o changes (paraffin wax)
2.4.1. Definition
• Process whereby the clearing agent is completely
removed from the tissue and replaced by a medium
that completely fills the tissue cavities and gives a firm
consistency to the specimen
• Allows easier handling and cutting of suitably thin
sections without any damage or distortion to the tissue
and cellular components
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
2.4.2. Infiltration Process
• Tissue is infiltrated with a suitable histological wax
(paraffin)
• Paraffin is liquid at high temp (60°C) → solidify when
cooled (20°C)
o Allows sections to be cut
• Wax
o Mixture of purified paraffin wax and additives
(resins)
o Allows thin sectioning on microtomes
Fig 8. Perpendicular fluid flow through tissue
cassettes
(Source: instructor’s slides)
3.3. TEMPERATURE CMB; NADG
• Must be kept at 37 – 45 °C
•
High temperature
can cause tissue
to
shrink
and
become hard and
brittle
Fig 7. Dehydration-Clearing-Infiltration Process
(Source: Instructor’s slides)
3. FACTORS AFFECTING TISSUE PROCESSING CMB; NADG
•
Factors that impact the duration of tissue processing
and extent of infiltration:
o Tissue density and thickness
o Agitation
o Temperature
o Vacuum and pressure
3.1. TISSUE DENSITY AND THICKNESS CMB; NADG
• Spongy tissues: more rapidly infiltrated by wax than
hard and dense tissues
• Hard and dense tissues: take longer time to finish
their processing
• Tissue thickness: influences the rate of reagent
diffusion and processing time
• The thicker the block, the longer time it takes for
tissue processing to be over
• Recommended is typically not more than 4 mm
thick
• When samples are put in cassettes, it shouldn’t
be too thick as it would take longer to process
3.2. AGITATION (VERTICAL OR ROTARY OSCILLATION
MECHANISMS) CMB; NADG
In the chambers, in the tissue processor, there
are vertical or rotary oscillation mechanisms
(Trinidad, 2022).
•
•
•
Increases the flow of fresh fluids in and around tissues
(speeds up fluid exchange)
Increases the surface area available for fluid exchange
Cassette perforations should be perpendicular to the
fluid flow
•
Low temperature
increases
the
viscosity of reagents
used
in
tissue
processing
• ↓ rate of diffusion
(rate
the
substances move
through
the
tissue)
• ↑ processing time
3.4. PRESSURE CMB; NADG
• The baskets or chambers have corresponding
pressures which must be controlled
Reduced pressure
High pressure
• increased
(↑) • facilitates infiltration of
dense specimens with
infiltration rate
the more viscous
• decreased
(↓)
embedding media
tissue processing
time
3.5. VACUUM CMB; NADG
Chambers also have a vacuum mechanism in
them (Trinidad, 2022).
•
•
Improves processing quality and aid in removal of
trapped air from porous tissue
Can reduce the infiltration time when dealing with
dense and fatty tissues
3.6. OTHER TECHNICAL CONSIDERATIONS CMB;NADG
• Baskets and metal cassettes should be cleaned and
wax-free
• Tissues (inside baskets or chambers) should not be
packed too tightly in baskets
Ensure that there is some space within and that
the fluid can completely infiltrate the tissues
(Trinidad, 2022).
•
Processors must be free of spilled fluids and
accumulated wax
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
•
•
Fluid levels must be higher than the specimen
containers
o Check the level of fluid within the baskets.
Timing and delay mechanism must be correctly set
and checked against the appropriate processing
schedule
Processing usually takes overnight (Trinidad,
2022).
•
Quality Assurance Program: keep a processor log
(records)
o Number of specimens processed
o Processing reagent changes
o Temperature checks on the wax baths (kept at
a certain temperature for infiltration)
o
•
Completion of the routine maintenance
schedule
After 12 hours, tissue processing is done.
Fig 9. Correct Orientation in Embedding (yellow arrow)
(Source: Instructor’s slide)
4.1. EMBEDDING MOLDS PDEF; NADG
• Plastic Molds
o “Peel-away” type of plastic mold
o Disposable
GENERAL STEPS
Tissue Cassette → Paraffin Blocks → Small Tissue Sections
Fig 10. Plastic molds / “peel-aways”
(Source: Instructor’s slide)
•
4. EMBEDDING/CASTING/BLOCKING PDEF; NADG
•
Embedding
o Also known as casting or blocking
o The last step in tissue processing
o Process by which the impregnated tissue is
placed into a precisely arranged position in
a mold containing a medium which is then
allowed to solidify
Metal Molds
o Used with a tissue cassette
o Reusable
o NOT disposable
“This tissue has been impregnated by paraffin,
and we now need to put it in a mold containing the
same medium as well.” (Trinidad, 2022)
•
•
The tissue is oriented and placed in a mold that is filled
with molten wax to form a solid tissue block.
The CORRECT orientation of the tissue in a mold is
the most important step in embedding.
o Embedded with the surface to be cut facing
down in the mold
“It is important that you embed the tissue properly
so that what you [are] looking at is what you will
be cutting first.” (Trinidad, 2022)
Fig 11. Metal molds and tissue cassettes
(Source: Instructor’s slide)
4.2. EMBEDDING MACHINE PDEF; NADG
Figure 12. Embedding machine used in UST-FMS
(Source: Instructor’s slide)
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PATHOLOGY
SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
•
Paraffin Wax – used for embedding
o Simplest, most common, & BEST embedding
medium used for routine tissue processing
o Also used in infiltration step
Figure 13. Parts of an embedding machine
(Source: Instructor’s slide)
Embedding machine
• This is the standard method to
produce blocks of tissue for section
cutting.
• Usually, this procedure is performed
using
an
embedding
machine,
surrounding the tissues by a medium
such as paraffin wax, which when cooled
and solidified will provide sufficient
support for section cutting.
Fig 14. Paraffin Wax
(Source: Instructor’s slide)
4.3. STEPS IN EMBEDDING PDEF; NADG
•
STEP 1: Open the tissue cassette
o To expose the tissue
Table 3. Parts of an embedding machine
PART
DESCRIPTION
Wax
Dispenser
Molten wax is
placed here and
is dispensed by
this mechanism.
Forceps
Warmer
Where forceps
are warmed.
Tissue
Warmer
Hot Plate
Cold
Plate
Where tissues
are placed first
before they are
placed in the
embedding
machine.
To maintain the
heat or to heat
objects/samples
with controlled
heating usually up
to 65˚C.
To cool the
processed block
(see step 7 of
embedding)
usually up to a
temperature of 20˚C
PICTURE
Fig 15. Open the tissue cassette
(Source: Instructor’s slide)
•
STEP 2: Select the mold that fits the tissue
o Plastic or metal mold depends on size &
shape
o Allowance: at least 2 mm margin of wax
“You need to place the tissue in the middle and
you need it to have at least 2 mm of margin of wax
all around, so choose the right mold for the
tissues.” (Trinidad, 2022)
Fig 16. Select the mold that fits the tissue
(Source: Instructor’s slide)
•
STEP 3: Dispense wax in the mold
o about 1/2 or 2/3 full
Fig 17. Dispense wax in mold
(Source: Instructor’s slide)
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
•
STEP 4: Using clean and warm forceps, select the
tissue and orient on the mold, gently pressing it flat
on the wax
o This is the significance of the forceps warmer
part of the embedding machine
Fig 22. Remove the block from the mold when the wax is
completely cooled and hardened
(Source: Instructor’s slide)
Fig 18. Using a clean and warm forceps, select the tissue and
orient on the mold, gently pressing it flat on the wax
(Source: Instructor’s slide)
•
Fig 23. Paraffin block
(Source: Instructor’s slide)
STEP 5: Insert the identifying label on the mold, and
place tissue cassette on top
5. SECTIONING/MICROTOMY PDEF, RPT; NADG
●
Sectioning / Microtomy – a paraffin-embedded tissue
is trimmed and cut into uniformly thin slices using a
microtome to facilitate studies under the microscope
o The embedded tissues must be cut into thin
sections to be placed on a slide
Fig 19. Insert the identifying label on the mold, and place
cassette on top
(Source: Instructor’s slide)
•
STEP 6: Dispense wax, completely filling the mold
Fig 20. Dispense wax, completely filling the mold
(Source: Instructor’s slide)
•
STEP 7: Cool the block on the cold plate
Fig 24. A Microtome
(Source: Instructor’s slide)
5.1. PARTS OF A MICROTOME PDEF; NADG
● Microtome
o Knife and its Holder – knife is inserted here
o Block holder (Chuck)
o Base (Microtome Body)
o Rotary Wheel – turned to make the block
holder go nearer to the knife as you cut along
the sections
Fig 21. Cool the block on the cold plate
(Source: Instructor’s slide)
•
STEP 8: Remove the block from the mold when the
wax is completely cooled and hardened
o This will take about 30 minutes
o END PRODUCT → PARAFFIN BLOCK
ROTARY
WHEEL
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
•
STEP 4: Continue cutting until a ribbon of sections
is created
Fig 25. Microtome and its Parts
(Source: Instructor’s slide)
Fig 29. Continue cutting until a ribbon of sections is created
(Source: Instructor’s slide)
5.2. STEPS IN SECTIONING PDEF, RPT; KDSC
•
STEP 1: Insert the knife blade into the holder and
tighten
o Make sure that it is secure
o A loose and unsecured knife may result in
imperfect sections
Fig 30. Tissue ribbons
(Source: Instructor’s slide)
Fig 26. Insert the knife blade into the holder and tighten
(Source: Instructor’s slide)
•
STEP 2: Orient the block in the microtome chuck
o The block holder is also called the chuck
o The long axis of the block should be parallel to
the knife
•
STEP 5: Float the ribbon on the water bath
o Remove wrinkles or bubbles
o Separate sections from each other by teasing
them very carefully apart using forceps in the
water bath
o Paraffinized ribbons of serial tissue sections
can be removed from the microtome knife as
they are cut
▪ Slight traction is exerted on the end of
the ribbon stretching it gradually over
the blade while floating carefully in a
warm water bath
Fig 27. Orient the block in the microtome chuck
(Source: Instructor’s slide)
•
STEP 3: Turn the wheel on the side of the
microtome and start cutting
o Block will go nearer to the knife as you turn
the wheel
o In the initial rough trim of blocks, thickness of
the blocks will be 15-30 mm
▪ But it can be adjusted in the
microtome
o The actual section slices, or the thin slices
would be 3-5 microns
Fig 28. Turn the wheel on the side of the microtome and start
cutting
(Source: Instructor’s slide)
Fig 31. Float the ribbon on the water bath
(Source: Instructor’s slide)
o
Water bath temperature should be kept 5-10˚C
below the melting point of the embedding wax
▪ Too HOT: desiccated-looking section
▪ Too COLD: excessive tissue wrinkling
Fig 32. Water bath machine
(Source: Instructor’s slide)
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
Table 4. Problems and appearances of ribbons
APPEARANCE
PROBLEM
5.3. ARTIFACTS RPT
• Good microtomy techniques will minimize artifacts
that can lead to difficult diagnostic interpretation of
special stains
Thin
Desiccated (dried), almost transparent
Thick
Too opaque
Figure 29. A badly sectioned tissue
(Source: Instructor’s slide)
Normal
•
•
STEP 6: Mount sections on glass slide
o Fish the ribbons out of the water bath carefully
into the glass slide
Artifacts include:
o Tearing
o Ripping
o Creases
o Holes
o Folding
o Floaters
▪ Extraneous tissue pieces that are not
part of the sample
▪ During the fishing out step of the water
bath, you can get floating tissue that is
not part of your section
▪ May be part of the other samples that
have been floating in the water bath
“Thus, it is important to regularly change the
water in the water bath.” (Trinidad, 2021)
Figure 27. Mount sections on glass slide
(Source: Instructor’s slide)
•
Tissue slide is drained
o May be gently heated in a drying oven to
evaporate the water between the sections and
the glass and to melt the wax and improve
adhesion to the slide
Figure 30. Contaminant floaters from the fishing process
(Source: Instructor’s slide)
●
●
Figure 28. Drying oven
(Source: Instructor’s slide)
6. STAINING RPT; KDSC
Staining – process whereby tissue components are
made visible in microscopic sections by direct
interactions with a dye or staining solution
o Produce a contrast between different tissues and
cellular components based on their varying
affinities for most dyes or stains
Before staining the slides, you need to prepare them
first by:
o Dewaxing – heat slides in 60C° for at least 30
minutes to soften the wax
▪ Can also be done in the drying oven (the
one used to drain tissue slides)
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
o
After dewaxing, wash with xylene → absolute
alcohol → distilled water (this procedure is the
one commonly done in laboratories)
▪ Some laboratory protocols: xylene →
absolute alcohol → 70% alcohol → 50%
alcohol → distilled water
▪ Thus, protocols vary from lab to lab
Figure 31. Staining area in the UST-FMS Pathology lab
(Source: Instructor’s slide)
●
Basic Principle in Staining: “Opposites Attract”
o ACIDIC cell components (nucleus) has greater
affinity for Basic Dyes
o BASIC cell components (cytoplasm) has greater
affinity for Acidic Dyes
6.1. HEMATOXYLIN AND EOSIN STAIN (H&E) RPT; KDSC
• aka “the routine stain”; Used for routine
histopathologic examination
Figure 33. Examples of Other Stains
(Source: Instructor’s slide)
“Keep in mind that there are many other stains
that you can use with different uses depending on
what you are looking for in your specimens.”
(Trinidad, 2021)
●
●
Figure 32. Hematoxylin and Eosin Stain
(Source: Instructor’s slide)
Table 3. Difference between Hematoxylin and Eosin
●
●
●
●
●
●
●
●
●
●
●
HEMATOXYLIN
EOSIN
Basic dye
● Acidic dye
Stains cell nuclei
● Stains cytoplasm
purple or blue
(basic), connective
tissue and other
Nuclei is the acidic
extracellular
component of the cell
because it contains
substances pink or
red
nucleic acids (DNA,
RNA)
OTHER STAINS
Periodic Acid Schiff ● Grocott-Gomori
(PAS)
Methenamine Silver
Masson’s Trichrome
● Warthin Starry
Alcian Blue
● Giemsa
Reticulin
● Gram
Oil Red O
● Perl’s Prussian Blue
Sudan Black
Congo Red
Acid Fast
●
7. MOUNTING IN TISSUE SECTIONS RPT; KDSC
Last step in tissue processing
Results in a permanent histological preparation
suitable for microscopy, after adhesion of the sections
on to the slide and appropriate staining of the tissue
Mounting medium
o Applied between the section and coverslip after
staining and prevents movement of the coverslip
o Protects the stained section from getting scratched
o Facilitates easy handling and storage of slides
o Prevent bleaching or deterioration of slides
▪ In time, slides usually fade
▪ Putting mounting medium and cover slip
slows down the rate of the fading of slides.
7.1 STEPS IN MOUNTING RPT
1. Apply 1 or 2 drops of mounting medium
2. Put the cover slip over the tissue very carefully,
avoiding bubbles
o Bubbles prevent one from seeing the tissues
clearly in the microscope
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SHIFT 1 | LESSON 3 | TISSUE PROCESSING, MICROTOMY, AND STAINING
Figure 34. Steps in Mounting of Tissue Sections Stains
(Source: Instructor’s slide)
•
8. LABELING RPT; KDSC
Identification and correctly labeling the tissue blocks
and its corresponding slides
o Stickers may be used for samples in tissue slides.
o Barcodes are the most ideal label because they
can directly be inputted in the computer, helping
streamline the process.
Figure 35. Labeled tissue slides on the upper portion of the
glass slide
(Instructor’s slide)
REFERENCES
Main References
● Lecture Powerpoint and Video Recording by Dra.
Celestine Marie G. Trinidad, MD, DPSP
Supplementary References
● Pathology Trans from LEAPMed Batch 2021 and 2022
TWG EIC
TEG EIC
M.A.B.B.
N.A.D.G.
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