Infiltration And Embedding Flashcards
Removal of the clearing agent
Impregnation/ Infiltration
It fill all the open spaces by the infiltrating or impregnating medium
Impregnation/ infiltration
It gives the tissue a firmer consistency for better sectioning
Impregnation/ infiltration
Types of impregnating medium
Paraffin
Celloidin
Gelatin
Plastic
It is a solid hydrocarbons with several melting points
Paraffin wax
Simplest most common and best embedding medium used for routine tissue processing
Paraffin wax
Serial sections are cut easily without distortion, very rapid and many staining procedures are permitted
Paraffin wax
Methods of infiltration
Manual method
Automated method
Vacuum embedding
It needs four changes of paraffin wax at 15 minutes intervals.
Manual method
Temperature should be 2-5C higher than the melting point of wax
Manual method
It uses autotechnicon (elliot bench type)
Automated Method
2-3 changes of wax is needed with constant agitation, with 12 stations
Automated Method
Wax impregnation under negative atmospheric pressure inside an embedding oven
Vacuum embedding
It is the most rapid type
Vacuum embedding
Recommended for urgent biopsies such as lungs, bones, eyes, spleen and CNS tissues
Vacuum embedding
Average melting point of paraffin wax
56-58C
If lab temperature is at 20-24C
54-58C
If lab temperature is 15-18C
50-54C
Advantages of paraffin wax
Rapid
Compatible with most staining methods
Allows cutting of serial sections
Disadvantages of paraffin wax
Tissue brittleness
Prolonged fixation may cause shrinkage and hardening
Not for fatty tissues
Inadequate process may promote retention of clearing agent
Tissues that are difficult to infiltrate may require longer_________
Immersion time
Important components of the autotechnicon
Glass beakers
Transfer arms
Electrical clock
Cover plate
Spring loaded plunger
Example of automatic tissue processing machine
Elliot bench type processor
Controlled by electrical current moves the tissues from one processing reagent to another
Transfer arm
Serves to control the time needed for each processing step
Electrical clock
Factors affecting paraffin wax impregnation
Temperature of the oven
Nature and size of the tissue
Type of clearing agents to be removed
Substitutes for paraffin wax
Paraplast
Embeddol
Bioloid
Tissue mat
Ester wax
Carbowax and PEG
Highly purified paraffin and synthetic plastic polymer
Paraplast
Has a 56-57C melting point
Paraplast
More uniform blocks, better ribbons
Paraplast
Synthetic wax and has a 56-58C melting point
Embeddol
Semi synthetic form of paraffin and recommended for eye specimen
Bioloid
Rubberized paraffin and has a similar property with paraplast
Tissue mat
Does not require clearing
Harder than paraffin wax
Ester wax
46-48C melting point
Used in sliding or sledge type of microtome
Ester wax
Water soluble waxes
Does not require dehydration and clearing
Carbowax and PEG
Ideal for fats and lipids, enzyme and histochemical studies
Hygroscopic
Tissues are difficult to float out
Carbowax and PEG
Type of impregnating medium that is highly volatile and a nitrocellulose dissolved in ether and alcohol
Celloidin/ collodion
Type of impregnating medium that does not require heat and allows cutting of hard tissues without distortion
Celloidin/ collodion
Type of impregnating medium that is slow, difficult to cut serial sections and pictomicrograph are difficult to obtain
Celloidin/ collodion
Methods of celloidin impregnation of tissue
Wet celloidin
Dry celloidin
Nitrocellulose method
Methods of celloidin that is recommended for bones, teeth, brains and whole organs
Dissolved in 70-80% alcohol
Wet celloidin
Methods of celloidin that is recommended for whole eye section, dissolved in Gibson’s mixture
Dry celloidin
Methods of celloidin that can be used in high concentrations and forms harder tissue block
Nitrocellulose method
Method of celloidin that is explosive
Nitrocellulose method
A type of impregnating medium that is used for histochemical and enzyme studies
Gelatin
It is for delicate specimens and frozen section
Gelatin
It does not require dehydration and clearing, volume should be 25x of the tissue volume
Gelatin
It contains phenol to prevent molds
Gelatin
It is also known as casting or blockin
Embedding
It is a process where an impregnated tissue is placed into a precisely arranged position in a mold, containing an embedding medium and allowed solidify
Embedding
Orientation for embedding
Placing the tissue in the center of embedding mold
If you use paraffin for embedding:
Cool the paraffin in the mold at -5C
Types of molds
Leuckhart’s Embedding mold
Compound embedding unit
Plastic embedding rings and base molds
Tissue tek
Disposable molds
A 2 L shaped strips of heavy metal
Leuckhart’s embedding mold
Examples of these are peel away, ice tray, and paper boat
Disposable molds
Other embedding medium that is used for hard tissues and large sections of whole organ
Celloidin or nitrocellulose method
Other embedding medium which infiltration with celloidin is needed and subsequently embedded with paraffin
Double embedding method
Other embedding medium that is used for electron microscopy sections
Plastic emebedding
Examples of plastic (resin) embedding
Epoxy- reduces antigenicity, toxic and damage tissue
Polyester
Acrylic
Removal of excess wax to form a truncated pyramid
Trimming
What length of wax should surround the tissue bock?
2mm
A type of mold that has the advantage of embedding more specimens at a time, thereby reducing the time needed for blocking
Compound embedding unit
A type of mold that serves as the block holder during cutting
Plastic embedding rings and base mold
A type of mold that consists of a white plastic cassette mold with detachable hinge, it is used to hold the tissue specimen throughout fixation, dehydration, clearing and wax impregnation
Tissue tek
