IMPREGNATION AND EMBEDDING Flashcards
___has a wide range of melting points which is important for use in the fifferent climactic regions of the world
Inexpensive
Provides quality sections
Can be used for most routine and special stains
Paraffin wax
LOW MELTING POINT PARAFFIN – usually softer
HIGH MELTING POINT PARAFFIN – usually harder
Melting points range from ___ (commonly 56C)
40-70C
Paraffin wax that contains plasticizers or other resin additives are commercially available
These mixtures create paraffin with the desired hardness for the tissues to be embedded
Additives have a high melting point than paraffin which makes the tissues brittle.
Examples:
Beeswax Rubber Ceresin Plastic polymers Diethylene glycol disteareate
Sub for paraffin wax.
Is a mixture of highly purified paraffin and synthetic plastic polymers
Melting point: 56-57C
More elastic and resilient than paraffin wax thereby permitting large dense tissue blocks such as bones and brain to be easilt cut
Better ribboning of tissues and serial sectioning is allowed
PARAPLAST
similar to paraplast with a melting point of 56-58C which is less pressible than paraplast
EMBEDDOL
recommended for embedding eyes
BIOLOID
paraffin plus rubber with the same property as paraplast
TISSUE MAT –
Lower melting point (46-48C) but is harder than paraffin
Non water soluble but is soluble in 95% ethyl alcohol and other clearing agents
Three to four changes are needed
Cutting should be done with a sliding or sledge type microtome
ESTER WAX
Mostly are polyethylene glycols with melting points of 38-42C or 45-56C
WATER SOLUBLE WAXES
appears solid at room temperature
Does not need dehydration and clearing
Does not remove neutral wax
Good for enzymatic studies
Used in four changes : 70% (30mins)-90% (45 mins) and 100% (X2) (1H) at a temperature of 56C
Tissues are difficult to float out due to water miscibility (remedied by _ or _ )
CARBOWAX
soap or 10% polyethylene glycol 900
Also known as COLLODION
Is a purified form of nitrocellulose soluble in many solvents suitable for specimens with large hollow cavities which tend to collapse, for hard and dense tissues such as bones and teeth and large tissue sections of the embryo
Supplied in:
CELLOIDIN
Thin (2%)
Medium (4%)
Thick (8%)
Disadv.
Impregnation is very slow (days to week) Very thin sections (less than 10u) are difficult to cut Serial sections are different to prepare Vapor of ether solvent is very flammable Photomicrographs are difficult to obtain Very volatile
CELLOIDIN
Cellodin Methods
Recommended for bones, teeth, large brain sections, and whole organs
Recommended for the eyes
WET METHOD
DRY METHOD
involves the enclosing of properly processed, correctly oriented specimens in a support medium that provides external support during microtomy
EMBEDDING
consists of two L shaped strips of heavy brass or metal arranged on a flat metal plate
LEUCKHART’S EMBEDDING MOLD
is made up of a series of interlocking plates resting on a flat metal base forming several compartments
COMPOUND EMBEDDING UNIT
DISPOSABLE EMBEDDING MOLDS
3
Peel away
Plastic ice trays
Paper boats
BLOCKING OUT MOLDS 4
DISPOSABLE EMBEDDING MOLDS
PLASTIC EMBEDDING RINGS AND BASE MOLD
COMPOUND EMBEDDING UNIT
LEUCKHART’S EMBEDDING MOLD
Advantages:
Ease of use
Speed
Tissue and holder are firmly attached creating a single unit
Blocks filled immediately after sectioning
Permanent identification
EMBEDDING CENTERS
EMBEDDING CENTERS Consists of three modules:
–allows the paraffin to solidify and ensures a small crystalline structure producing fewer artifacts when sectioning the tissue
COLD PLATE
PARAFFIN DISPENSER
HEATED STORAGE AREA
COLD PLATE
Arteries, veins, fallopian tubes and vas deferens
Should be cut in cross sections of the lumen
TUBULAR STRUCTURES
Cut in the plane at right angles to the surface
Oriented so that the epithelial surface is cut last minimizing compression and distortion of the epithelial layer
SKIN, INTESTINE, GALLBLADDER, and OTHER EPITHELIAL BIOPSIES
Sections containing both transverse and longitudinal planes
MUSCLE BIOPSIES
Oriented side by side with the epithelial surface facing the same directions
MULTIPLE PIECES OF TISSUES
The process where tissues are first infiltrated with Celloidin and subsequently embedded in parrafin
Used to facilitate cutting of large blocks of dense firm tissues like the brain
DOUBLE EMBEDDING METHOD
AUTOMATED TISSUE PROCESSING
The basic principle for tissue processing requires the exchange of fluids using a series of solutions for a predetermined length of time in a controlled environment
No change for years
Recent advances now includes specialty MICROWAVE OVENS and the emergence of constant THROUGH-PUT processors
_ and _ were the first automated tissue processors used in histology laboratory.
The CAROUSEL TYPE PROCESSOR (tissue transfer) and the SELF CONTAINED FLUID EXCHANGE SYSTEMS
AUTOMATED TISSUE PROCESSORS
Makes use of an automatic tissue processing machine called
AUTOTECHNICON
The machine fixes, dehydrates, clears and infiltrates tissues, thereby decreasing the time and labor needed during the processing of tissues
Impregnation is hastened by constant tissue agitation
AUTOTECHNICON
Permits the turning of platforms and easy access to beakers and wax baths.
It makes use of 12 individual processing steps with 1 liter capacity glass beakers and two thermostatically controlled wax baths.
AUTOMATED TISSUE PROCESSORS
THE ___ controlled by electrical current moves the tissues from one reagent to another (by clock schedules)
TRANSFER ARM
Involves the wax impregnation under negative atmospheric pressure inside the embedding oven to hasten removal of air bubbles and clearing agent from the tissue block thereby promoting a more rapid wax penetration of tissues
VACUUM EMBEDDING
Time is reduced from 25-75%
Tissues are not over-exposed to heat - brittleness, shrinkage, and hardening of tissues is therefore prevented
VACUUM EMBEDDING
Hourly throughput of up to 120 specimens.
Tissue-Tek Xpress® Continuous Rapid Tissue Processor
This instrument uses
an innovative, low-wattage microwave technology
molecular-friendly reagents
traditional vacuum infiltration techniques
Specimen slides now make their way to the pathologist faster than ever, enabling same-day diagnosis and thus drastically reducing patient stay and anxiety.
Tissue-Tek Xpress® Continuous Rapid Tissue Processor
The Xpress® reagents preserve _ _ and _ in the paraffin block, eliminating the need to use fresh tissue for molecular studies.
DNA, RNA and proteins
The reagents provide excellent morphology, sharp nuclear detail and crisp cellular images equal to or better than conventional tissue processing.
Compatible with special staining and immunohistochemistry methods, these reagents can often eliminate the need for antigen retrieval.
Tissue-Tek® Xpress® Continuous RapidTissue Processor
is the fifth generation of instruments for the enclosed automatic tissue processing of human and animal tissue specimens.
The Tissue-Tek® VIP® 5 Vacuum Infiltration Processor
20 user-defined processing programs for fixation, dehydrating, clearing and paraffin infiltration of tissue specimens.
The Tissue-Tek® VIP® 5 offers
provides more efficient processing using a redesigned control panel for easy monitoring of the system status and several preventive functions to protect against unexpected situations.
Capacity of __ cassettes
The Tissue-Tek® VIP® Jr.
150
The system to fully automate your embedding
No more opening of the cassette from grossing through sectioning
Continuous loading of your blocks after processing
A throughput of 120 blocks per hour, matching the capacity of the Tissue-Tek® Xpress® Continuous Rapid TissueProcessor
Tissue-Tek® AutoTEC® Automated Embedder
MAINTENANCE TIPS
Any spillage or overflow should be wiped away immediately
Accumulation of wax in any surface should be removed
Temperature of the paraffin bath should be set to 3C above the melting point of the paraffin
Timing should be checked when placing tissue cassettes in the processor, specially when delayed schedules are selected
MAINTENANCE TIPS
Any spillage or overflow should be wiped away immediately
Accumulation of wax in any surface should be removed
Temperature of the paraffin bath should be set to 3C above the melting point of the paraffin
Timing should be checked when placing tissue cassettes in the processor, specially when delayed schedules are selected
Rarely used today
There can be circumstances the tissue sample requiring the tissue sample to be manually processed, including
Power failure and equipment malfunction
Large tissue samples requiring more time that allocated by the automated processor
Small biopsies, such as transplant specimens needing a rapid diagnosis
MANUAL TISSUE PROCESSING
