HISTO IMPREGNATION AND EMBEDDING CH 11

Process whereby the clearing agent is completely removed from the tissue and replaced by a medium that will completely fill all the tissue cavities and give a firm consistency to the specimen.

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.

The most popular infiltration and embedding medium in the histology laboratory

Vacuum reduces the time required for complete impregnation.

At what °C wax is normally used for routine work

Rarely used today

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

Makes use of an automatic tissue processing machine called

Example of an automatic tissue processing machine

Presence of any odor in the clearing agent during final paraffin wax bath indicates that the paraffin wax needs to be changed.

Dehydrating fluids should be changed once a year since dehydration is the most critical stage of tissue processing and inadequate dehydration is difficult to correct once the tissue is in paraffin.

Clearing agent and the dilute ethanols should be changed at least twice a week.

The CAROUSEL TYPE PROCESSOR (tissue transfer) and the SELF CONTAINED FLUID EXCHANGE SYSTEMS were the first automated tissue processors used in histology laboratory.

Shortens the processing time from hours to minutes

Involves 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

In vacuum embedding, degree of the vacuum should not exceed

Vacuum impregnation gives the fastest result.

Fresh wax should be filtered before use in a wax oven at what temperature higher than its melting point.

When wax has been reused, some amount of water inevitably is mixed with it.

When using an automatic tissue processing machine, wax usually becomes admixed with the clearing agent, especially in the first beaker; hence, water must be discarded.

Usually softer

Usually harder

Heating the paraffin to low temperatures alters the properties of the wax.

Is a mixture of highly purified paraffin and synthetic plastic polymers

What is the melting point of paraplast?

During the winter, 54 to 56°C Paraplast may be used if the tissue is cut in a cool room

During the summer it may be necessary to use 60 to 63°C, although this is to be avoided if possible in order to not to "cook" the tissue.

Similar to paraplast with a melting point of 56-58°C which is less pressible than paraplast

Recommended for embedding eyes

Paraffin plus rubber with the same property as paraplast

Lower melting point (46-48°C) but is harder than paraffin

Mostly are polyethylene glycols with melting points of 38- 42°C or 45-56°C

Most commonly used in water soluble waxes and a polyethylene glycol containing 18 or more carbon atoms, appears solid at room temperature

Water soluble waxes is good for enzymatic studies

Added to proprietary blends of plastic polymer paraffin waxes reduces infiltration times and facilitates thin sectioning

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.

Celloidin impregnation is very slow

Recommended for bones, teeth, large brain sections, and whole organs

This is done to avoid dehydration and shrinkage of tissues.

Recommended for whole eye sections

The dry method does not make use of alcohol due to the presence of cedarwood oil in the block

More explosive than celloidin and should therefore be handled with care.

Another form of celloidin soluble in equal concentration of ether and alcohol, with a lower viscosity,

Rarely used except when dehydration is to be avoided and when tissues are to be subjected to histochemical and enzyme studies.

Involves the enclosing of properly processed, correctly oriented specimens in a support medium that provides external support during microtomy

After impregnation, the tissue is placed into a mold containing the embedding medium and this medium is allowed to solidify.

Once tissues have been embedded, they may be stored in a room temperature place indefinitely until they are cut.

Consists of two L-shaped strips of heavy brass or metal arranged on a flat metal plate which can be moved to adjust the size of the mold to the size of the specimen

Made up of a series of interlocking plates resting on a flat metal base, forming several compartments

Consist of a special stainless steel base mold fitted with a plastic embedding ring, which later serves as the block holder during cutting.

This instrument uses – an innovative, lowwattage microwave technology – molecular-friendly reagents – traditional vacuum infiltration techniques

Hourly throughput of up to 120 specimens.

The Xpress reagents preserve DNA, RNA and proteins in the paraffin block, eliminating the need to use fresh tissue for molecular studies

The fifth generation of instruments for the enclosed automatic tissue processing of human and animal tissue specimens

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 150 cassettes

The system to fully automate your embedding

Disposable thin plastic embedding molds, available in 3 different sizes, are simply peeled off one at a time, as soon as the wax has solidified, giving perfect even block without trimming.

Such as those used in ordinary refrigerators may be recommended for busy routine laboratories

Normally utilized for embedding celloidin blocks but are equally useful for paraffin wax blocks

Allows the paraffin to solidify and ensures a small crystalline structure producing fewer artifacts when sectioning the tissue

Arteries, veins, fallopian tubes and vas deferens. Should be cut in cross sections of the lumen

Sections containing both transverse and longitudinal planes.

Oriented side by side with the epithelial surface facing the same directions

Used to be recommended for embedding hard tissues such as bones and teeth, and for large sections of whole organs like the eye, since the delicate layers of the eyeball are difficult to keep intact when other media are used.

The process by which tissues are first embedded or fully infiltrated with a supporting medium such as agar or nitrocellulose, then infiltrated a second time with paraffin wax in which they are subsequently embedded.

Has provided superior results for light microscopic studies, particularly in hard tissues such as undecalcified bone and for high resolution light microscopy of tissue sections thinner than the usual 4-6 µm, such as renal biopsies a n d bone marrow biopsies.

Made up of a carefully balanced mixture of epoxy plastic, catalysts and accelerators.

Slow, partly because the epoxy plastic itself is a large molecule

Have a lower viscosity but are often sold as mixtures of isomers.

Can be obtained pure, have very low viscosity, and infiltrate fastest.

Known to be carcinogenic.

Originally introduced for electron microscopy in the mid- 1950s, but have been superseded by more superior epoxides, and are now seldom used.

Made up of esters of acrylic or methacrylic acid, and are used extensively for light microscopy.

Added to the plastic as a catalyst that decomposes to form phenyl radicals acting as an active site for the polymerization of acrylics.