[1S] UNIT 1.2 Instrumentation in Histopathology Flashcards
(99 cards)
1
Q
Medical Technologists are also known as
A
Clinical Laboratory Scientists
2
Q
Overview of Different Laboratory Procedures
A
Numbering
Fixation
Decalcification*
Dehydration
Clearing
Impregnation
Embedding
Blocking
Trimming
Sectioning
Staining
Mounting
Labelling
3
Q
A procedure in which we assign a specific code/id for a particular specimen as the cassette containing the specimen is too small to include every specific detail
A
Numbering/Accessioning
4
Q
T/F: Numbering may be a combination of numbers and letters
A
T
5
Q
T/F: Site numbers (different areas of an organ) are not included if several sites are used
A
F; may also be included
6
Q
The process of preserving the tissues and its cellular constituents
A
Fixation
7
Q
● To prevent autolysis
● For the pathologists to read clearly and correctly to get an accurate diagnosis
A
Fixation
8
Q
Fixation reagent
A
Formalin
9
Q
The process of removing calcium ions to soften the tissue for easier cutting
A
Decalcification
10
Q
T/F: All specimens undergo decalcification
A
F; not all
11
Q
Decalcification reagent
A
Nitric Acid
12
Q
The process of removing water or excess water
A
Dehydration
13
Q
Why is water removed in a specimen?
A
Excess water is removed so that infiltration is performed properly
14
Q
Dehydration reagent
A
Alcohol
15
Q
● The process of removing alcohol
● Alcohol is removed to perform Impregnation or infiltration properly
A
Clearing
16
Q
Clearing agent
A
Xylene
17
Q
● The process of infiltrating the tissue with solution that will fill up the natural spaces to maintain the integrity of the specimen appearance
○ This provides rigid support for the tissues
○ To maintain the appearance and shape of the tissue microscopically
A
Impregnation / Infiltration
18
Q
Impregnation reagent
A
Paraffin Wax
19
Q
● The process of forming tissue block using a mold
● This is needed to perform cutting on a particular site
A
Embedding & Blocking
20
Q
● The process of removing excess wax
● _________ __________ is created so that we can prepare a tissue ribbon properly
A
Trimming
Truncated pyramid
21
Q
The process of cutting the tissue into micro thin slices of tissue
A
Sectioning / Cutting
22
Q
● The process of coloring the tissues to clearly identify cells clearly, correctly, and accurately
● To differentiate cell from one another
A
Staining
23
Q
● The process of securing the slide with a coverslip (permanent)
● To protect the stained slide
● Performed to protect the slide during handling and storage
A
Mounting
24
Q
● The process of providing a particular label
● Assigned number is also used for this process
A
Labelling
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How many steps are in tissue processing without decalcification?
12
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Characterized by the transfer of tissues, contained within a basket, through a series of stationary reagents arranged inline or in a circular carousel plan
AUTOMATED TISSUE PROCESSOR: Tissue Transfer Processors (Carousel-Type)
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TISSUE TRANSFER PROCESSORS (CAROUSEL-TYPE)
● ___ reagent positions & ___ wax positions
● Capacity = __________
● 9-10 and 2-3
● 30-110 cassettes
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are the instruments where sample tissue is placed
Cassettes
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Characterized by processing fluids pumped to and from a retort in which the tissues remain stationary
AUTOMATED TISSUE PROCESSOR: Fluid Transfer Processors
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FLUID-TRANSFER PROCESSORS
● _____ reagent stations with temperatures adjustable between ______
● ___ paraffin wax stations with variable temperature settings between ______
● Capacity: _________
● 10-12 reagent stations with temperatures adjustable between 30-45°C
● 3-4 paraffin wax stations with variable temperature settings between 48-68°C
● Capacity: 100-300 cassettes
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ADVANTAGES OF USING BOTH TISSUE-TRANSFER AND FLUID-TRANSFER
T/F: Flexibility of reagent selection
T
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ADVANTAGES OF USING BOTH TISSUE-TRANSFER AND FLUID-TRANSFER
T/F: The conditions and schedule are designed to provide optimum processing even for the small batches of tissues
T
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TISSUE PROCESSING
Used for making tissue blocks
Tissue Embedding Center
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TISSUE PROCESSING
Advantage:
○ It has a refrigeration system
■ The cold platform in the equipment
■ To solidify the tissue
■ Paraffin is solidified for the tissue to be stable
Tissue Embedding Center
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TISSUE PROCESSING: TISSUE EMBEDDING CENTER
Run the tissue embedding center for about _ hours to make sure paraffin wax is melted in the paraffin chamber
5
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TISSUE PROCESSING: TISSUE EMBEDDING CENTER
T/F: Not all blocking steps can be performed with the help of this machine
F; All blocking steps can be performed
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EMBEDDING PROCESS
What temperature should the paraffin be and wax melting point?
61°C
55-57°C (wax melting point)
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EMBEDDING PROCESS
T/F: Place cassettes in the paraffin reservoir in numerical order
T
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PARTS OF THE TISSUE EMBEDDING CENTER
The cool system of the Tissue Embedding Center
Freon Refrigeration System
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PARTS OF THE TISSUE EMBEDDING CENTER
Where embedded tissues, with their respected molds to hasten solidification
Cold Orientation Platform
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PARTS OF THE TISSUE EMBEDDING CENTER
○ A button that provides a non clogging flow of molten paraffin for casting the mold
○ Same button just like in water dispenser
Microswitch Dispenser
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PARTS OF THE TISSUE EMBEDDING CENTER
Hot orientation platform materials
Forceps
Tissue
Metal Mold
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PARTS OF THE TISSUE EMBEDDING CENTER
T/F: During embedding, the orientation is provided by the pathologist
T
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PARTS OF THE TISSUE EMBEDDING CENTER
○ Top left part of the machine
○ Above the cold orientation platform
○ Catches excess paraffin
Waste Drawer
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PARTS OF THE TISSUE EMBEDDING CENTER
Area where we can preheat the forceps
Hot Well
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Basic instrument used that is capable of cutting section at a predetermined thickness by sliding the block into a cutting tool which is fixed and attached to the machine.
Microtomy
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PRINCIPLE: A spring balance teeth or pawl is brought into contact with, and turns a ratchet feed wheel connected to a micrometer screw, which in turn rotated, moving the tissue block at a predetermined distance towards the knife for cutting sections at uniform thickness.
Microtomy
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KINDS OF MICROTOME
● Invented by Paldwell Trefall in 1881
● Simplest among the different types of microtome
Rocking Microtome
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KINDS OF MICROTOME
● Consist of a heavy base and two arms:
● Used to cut small and large blocks of paraffin tissues.
● Not recommended for serial sections because tissues are cut in slightly curved planes
Rocking Microtome
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KINDS OF MICROTOME
● Disadvantage:
○ Restrictions in size of tissue block that can be cut
○ Difficulty of reorienting the block
Rocking Microtome
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KINDS OF MICROTOME
● Minot microtome
● Invented by Minot in 1885-1886
Rotary Microtome
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KINDS OF MICROTOME
Most common type used for both routine and research laboratories
Rotary Microtome
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KINDS OF MICROTOME
● Operated by the rotation of the flywheel
● Causes reciprocal motion of the knife over the block
● Thickness of the section being automatically regulated by the ratchet feed wheel
Rotary Microtome
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ROTARY MICROTOME DIFFERENCE FROM THE ROCKING MICROTOME
T/F: The knife and the block holder are brought together by upward and vertical motions
T
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ROTARY MICROTOME DIFFERENCE FROM THE ROCKING MICROTOME
T/F: Cuts sections in a perfectly flat plane
T
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ROTARY MICROTOME DIFFERENCE FROM THE ROCKING MICROTOME
● Heavier and more stable
● More complex in design and construction
● More expensive
T
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ROTARY MICROTOME DIFFERENCE FROM THE ROCKING MICROTOME
T/F: The blade is placed in a blade-down position which is relatively dangerous
F; blade-up
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KINDS OF MICROTOME
● Developed by Adams in 1789
● Especially recommended for cutting extremely hard and rough tissue blocks
Sliding Microtome
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KINDS OF MICROTOME
● The knife is moved horizontally against a fixed block which progresses against it in an inclined plane
● Most dangerous type of microtome
Sliding Microtome
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KINDS OF MICROTOME
Consist of two movable pillars holding the adjustable knife clamps, allowing the knife to be set at an angle for cutting celloidin sections
Base-Sledge Microtome
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KINDS OF MICROTOME
Favored in laboratories where very hard tissue or large blocks are usually sectioned
Base-Sledge Microtome
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KINDS OF MICROTOME
• Suited for sectioning specimens embedded in all forms of media
• Comparatively more stable
Base-Sledge Microtome
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KINDS OF MICROTOME
The block remains stationary while the knife is moved backward and forward during the process of sectioning.
Standard Sliding Microtome
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KINDS OF MICROTOME
• Developed mainly for cutting celloidin-embedded tissue blocks
• Inherently more dangerous
Standard Sliding Microtome
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KINDS OF MICROTOME
• Invented by Queckett in 1848
• The stage for block holder is hollow and perforated around its perimeter, attached to a reinforced flexible lead pipe thru which carbon dioxide passes from a cylinder.
Freezing Microtome
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KINDS OF MICROTOME
• A simple lever operated valve allows the release of rapid intermittent burst of carbon dioxide
• A second cooling device for lowering temperature of the knife to facilitate sectioning.
Freezing Microtome
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KINDS OF MICROTOME
• Used to cut undehydrated tissues in a frozen state
• When histological demonstration of fat is needed
Freezing Microtome
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KINDS OF MICROTOME
• When certain neurological structures are to be
studied
• When sensitive tissue constituents to be studied are damaged or destroyed by heat.
Freezing Microtome
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KINDS OF MICROTOME
• Consists of microtome kept inside a cold chamber which has been maintained at a temperature between -5 to -30°C (-20°C) by an adjustable thermostat
• Capable of freezing fresh tissues within 2-3 minutes
• Can cut sections of 4µ with ease.
FREEZING MICROTOME: Cryostat
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KINDS OF MICROTOME
Provides a means of preparing thin sections of fresh frozen tissues for fluorescent antibody staining techniques or histochemical enzyme studies.
FREEZING MICROTOME: Cryostat
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KINDS OF MICROTOME
Most commonly used for rapid preparation of urgent tissue biopsies for intraoperative diagnosis.
FREEZING MICROTOME: Cryostat
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are used to mount tissues in the Cryotome
OCTs
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KINDS OF MICROTOME
Primarily used for cutting tissue sections at 0.5 micra for electron microscopy
Ultrathin Microtome
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KINDS OF MICROTOME
The knife used for cutting the ultrathin sections consists mainly of selected fragments of ______ ______ ______.
broken plate glass
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KINDS OF MICROTOME: ULTRATHIN MICROTOME
The specimen used is small, fixed in ______ ________, and embedded in plastic.
osmium tetroxide
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Other Kinds of Microtome
1. Vibrating Microtome
2. Hand Microtome
3. Saw Microtome
4. Laser Microtome
5. Computerized Microtome
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CARE OF THE MICROTOME
T/F: All the accumulated paraffin and small pieces of tissues must be brushed away with soft brush and not allowed to stay in the microtome
T
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CARE OF THE MICROTOME
T/F: After drying the machine and knife holder, the parts should be wiped with xylene
F; xylol not xylene
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CARE OF THE MICROTOME
T/F: Movable portions should be oiled thoroughly to prevent rusting
T
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CARE OF THE MICROTOME
T/F: Microtome must always be covered when not in use
T
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Trimming and section-cutting are done with a?
Microtome Knife
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TYPES OF MICROTOME KNIVES
• Usually 25 mm in length
• One side of the knife is flat while the other is concave.
Plane-Concave Knife
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TYPES OF MICROTOME KNIVES
• Usually 120 mm in length
• With both sides concave, recommended for cutting
paraffin embedded sections on a rotary microtome.
Biconcave Knife
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TYPES OF MICROTOME KNIVES
• Usually 100 mm in length
• Have both sides straight
• Recommended for frozen sections or for cutting extremely hard and tough specimens embedded in
paraffin blocks using a base-sledge type or sliding mircrotome.
Plane-Wedge Knife
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TYPES OF MICROTOME KNIVES
• Have a sharp cutting edge that can cut 2-4 µ thick sections with ease.
• Cheaper to use.
Disposable Blades
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TYPES OF MICROTOME KNIVES
• Used for trimming and semi-thin sectioning of tissue blocks for electron microscopy.
• Commercially prepared
• Should be prepared and stored in dust-free boxes with lids, just before use, to avoid contamination.
Glass Knives
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TYPES OF MICROTOME KNIVES
• Used to cut any type of resin block for electron microscopy
• Are brittle and expensive, but very durable
Diamond Knives
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TYPES OF MICROTOME KNIVES
They are already mounted in a metal block designed to fit directly into the knife holder of the ultrathin microtome when purchased.
Microtome Knives
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OTHER EQUIPMENT FOR TISSUE SECTIONING
• Temperature: About 10°C below the melting point of the paraffin wax.
• A small amount of detergent may be added to water to reduce surface tension and allow section to flatten out.
Floatation Water Bath
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OTHER EQUIPMENT FOR TISSUE SECTIONING
Temperature is set at the melting point of the wax, to 2-5 degrees Celsius above*
Drying Oven or Hot Plate
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OTHER EQUIPMENT FOR TISSUE SECTIONING
● To melt the paraffin
● Can be used to remove excess paraffin so that only the tissue is on the slide
Microwave
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OTHER EQUIPMENT FOR TISSUE SECTIONING
Both are needed for handling sections during cutting and removing folds and creases during “floating out” in waterbath.
Forceps & Squirrel/Camel Hairbrush
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Staining equipment should be?
Acid and alkali resistant, high temperature resistant
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OTHER EQUIPMENT
Programmed to perform H&E routine staining without any supervision. This has a continuous load and unload drawer access and robotic arm system that routinely transfers stained slides to another reagent or stain.
Recommended in a busy histopathology laboratory
Automated Stainer/Autostainer
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OTHER EQUIPMENT
These are covered glass vessels or containers, usually with grooves inside, and are used as containers of staining reagents during manual staining procedures.
Coplin Staining Jars
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OTHER EQUIPMENT
These are covered glass vessels or containers, usually with grooves inside, and are used as containers of staining reagents during manual staining procedures.
Coplin Staining Jars
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OTHER EQUIPMENT
Used to hasten the process of heating the slides after fishing the tissue section in the floatation water bath during sectioning in preparation for staining.
Up to 6 slide racks (30 slide capacities)
Slide Dryer
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PARTS OF TISSUE EMBEDDING CENTER
It filters particles or sediments
Microscreen
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PARTS OF TISSUE EMBEDDING CENTER
It forms a penthouse that contains molten paraffin. It has a microswitch dispenser usually on the lower left od the penthouse.
This chamber maintains an optimal paraffin temperature with adjustable thermostatic control
Paraffin Melting Chamber
