Infiltration and Embedding

Question 1

the 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

Answer 1

Infiltration/ Impregnation

Question 2

the 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.

Answer 2

Embedding

Question 3

What are the 4 types of infiltration and embedding medium?

Answer 3

1. Paraffin wax
2. Celloidin (collodion)
3. Gelatin
4. Plastic

Question 4

• simplest, most common, and best embedding medium used for routine tissue processing
• a polycrystalline mixture of solid hydrocarbons produced during the refining of coal and mineral oils
• solid at room temperature but melts at temperatures up to about 65°C or 70°C

Answer 4

Paraffin wax

Question 5

the most common melting point at different temperatures for histological use of paraffin wax

Answer 5

56°C - 58°C

Question 6

The traditional advice with paraffin wax

Answer 6

use about 2°C above its melting point

Question 7

To decrease viscosity and improve infiltration of the tissue, technologists often increase the temperature to

Answer 7

60°C or 65°C

Question 8

Paraffin wax is traditionally marketed by its melting points which range from

Answer 8

39°C to 68°C

Question 9

What happens when the paraffin was overheated?

Answer 9

brittle

Question 10

What happens if prolonged impregnation of paraffin wax?

Answer 10

tissue shrinkage and hardening

Question 11

Tissues become soft and shrunken, and tissue blocks crumble when sectioned and break up when floated out in a water bath.

Answer 11

Inadequate impregnation that promote retention of the clearing
agent

Question 12

In a laboratory with temperature ranging from 20-24°C, paraffin wax with a melting point of______ is indicated. If the laboratory temperature is between 15-18°C, the melting point of wax to be used should be between _______

Answer 12

54-58°C
50 and 54°C

Question 13

3 ways by which paraffin wax infiltration and embedding of tissues may be performed

Answer 13

1. manual processing
2. automatic processing
3. vacuum embedding

Question 14

at least 4 changes of wax are required at 15-minute interval and then the specimen is immersed in another fresh solution of melted paraffin for approximately 3 hours

Answer 14

Manual Processing

Question 15

One example of automatic tissue processing machine

Answer 15

Elliot Bench-Type Processor

Question 16

Presence of any odor in the clearing agent during final paraffin wax bath in automatic processing indicates

Answer 16

paraffin wax needs to be changed

Question 17

the most critical stage of tissue processing

Answer 17

dehydration

Question 18

Wax bath thermostats should be set at least ______ the melting point of the wax

Answer 18

3 degrees above

Question 19

• involves wax impregnation under negative atmospheric pressure inside an embedding oven
• this technique is particularly recommended for urgent biopsies, for delicate tissues such as lung, brain, connective tissues, decalcified bones, eyes, spleen, and central nervous system.

Answer 19

Vacuum Embedding

Question 20

With vacuum embedding, the time required for complete impregnation is reduced by _________ of the normal time required for tissue processing

Answer 20

25% -75%

Question 21

In vacuum embedding oven, the vacuum chamber is enclosed in a thermostatically controlled water-jacket, usually maintained at a temperature of _____ the melting point of the wax

Answer 21

2-4°C above

Question 22

In vacuum embedding oven, the degree of the vacuum should not exceed

Answer 22

500 mm. Hg

Question 23

This is provided to prevent water from being sucked back into the trap bottle and vacuum chamber when the water or suction pump is closed

Answer 23

stopcock

Question 24

Out of the 3 methods of paraffin wax impregnation, which one gives the fastest result?

Answer 24

Vacuum Impregnation

Question 25

infiltration in overheated paraffin (above 60°C) will

Answer 25

produce shrinkage and hardening of tissues and destroy lymphoid tissues completely

Question 26

To avoid shrinkage, hardening of tissues, and destroying lymphoid tissues completely due to infiltration in overheated paraffin _______

Answer 26

the paraffin oven must be maintained at a temperature 2 to 5°C above the melting point of paraffin to be used for impregnation

Question 27

Example of a coarse filter paper

Answer 27

Green's No. 904

Question 28

When wax has been reused, some amount of water inevitably is mixed with it, how can you remove the water?

Answer 28

heating the wax to 100-105°C, thereby raising its melting point

Question 29

Substitute for Paraffin wax

Answer 29

1. Paraplast 2. Embeddol 3. Ester wax 4. Water soluble waxes 5. Dimethyl sulphoxide

Question 30

- a mixture of highly purified paraffin and synthetic plastic polymers, with a melting point of 56-57°C. - more elastic and resilient than paraffin wax thereby permitting large dense tissue blocks such as bones and brain to be cut easily

Answer 30

Paraplast

Question 31

- synthetic wax substitute similar to Paraplast with a melting point of 56-58°C. - less brittle and less compressible than Paraplast

Answer 31

Embeddol

Question 32

a semisynthetic wax recommended for embedding eyes

Answer 32

Bio/aid

Question 33

a product of paraffin, containing rubber, with the same property as Paraplast.

Answer 33

Tissue Mat

Question 34

- has a lower melting point (46-48°C), but it is harder than paraffin - not soluble in water, soluble in 95% ethyl alcohol, hence can be used for impregnation without prior clearing of tissue

Answer 34

Ester wax

Question 35

microtome used for ester wax-impregnated tissue

Answer 35

sliding or sledge type microtome

Question 36

- plastic polymers, mostly polyethylene glycols with melting points of 38-42°C or 45-56°C

Answer 36

Water soluble waxes

Question 37

- a polyethylene glycol containing 18 or more carbon atoms, which appears solid at room temperature - It is soluble in and miscible with water; hence does not require dehydration and clearing of the tissue - Cytologic details are excellently preserved

Answer 37

carbowax

Question 38

Routine processing of carbowax

Answer 38

70% for 30 minutes 90% for 45 minutes 100% for 1 hour 100% for 1 hour temperature at 56°C

Question 39

What will reduce tissue distortion and promote flattening and "floating out" of sections?

Answer 39

Adding soap to water or using 10% Polyethylene Glycol 900

Question 40

- added to proprietary blends of plastic polymer paraffin waxes reduces infiltration times and facilitates thin sectioning - scavenges residual transition solvent and probably alters tissue permeability by substituting for or removing bound water thus improving infiltration.

Answer 40

Dimethyl sulphoxide (DMSO)

Question 41

Some individuals who handle DMSO-paraffin wax may experience

Answer 41

unpleasant and annoying oyster or garlic taste caused by DMSO metabolites

Question 42

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 for large tissue sections of the whole embryo.

Answer 42

Celloidin (Collodion)

Question 43

2 methods for celloidin impregnation

Answer 43

1. Wet Celloidin Method 2. Dry Celloidin Method

Question 44

celloidin method recommended for bones, teeth, large brain sections and whole organs

Answer 44

Wet Celloidin Method

Question 45

Celloidin method preferred for processing of whole eye sections

Answer 45

Dry Celloidin Method

Question 46

made up of equal parts of chloroform and cedarwood oil

Answer 46

Gilson's mixture

Question 47

another form of celloidin soluble in equal concentration of ether and alcohol, with a lower viscosity, allowing it to be used in higher concentrations and still penetrate tissues rapidly

Answer 47

Low Viscosity Nitrocellulose (L.V.N.)

Question 48

examples of plasticizers

Answer 48

oleum ricini castor oil

Question 49

- rarely used except when dehydration is to be avoided and when tissues are to be subjected to histochemical and enzyme studies - used as an embedding medium for delicate specimens and frozen tissue sections because it prevents fragmentation of tough and friable tissues when frozen sections are cut. - water-soluble, and does not require dehydration and clearing - has a low melting point and does not cause over-hardening of tissues by heating.

Answer 49

GELATIN

Question 50

serves to prevent the growth of molds in gelatin impregnation

Answer 50

1% phenol

Question 51

The process by which a tissue is arranged in precise positions in the mold during embedding, on the microtome before cutting, and on the slide before staining

Answer 51

Orientation

Question 52

4 Types of Blocking-out Molds

Answer 52

Leuckhart’s Embedding Mold Compound Embedding Unit Plastic Embedding Rings and Base Mold Disposable Embedding Molds

Question 53

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

Answer 53

Leuckhart’s Embedding Mold

Question 54

an embedding mold that is made up of a series of interlocking plates resting on a flat metal base, forming several compartments. It has the advantage of embedding more specimens at a time, thereby reducing the time needed for blocking.

Answer 54

Compound Embedding Unit

Question 55

an embedding bold that is consist of a special stainless steel base mold fitted with a plastic embedding ring, which later serves as the block holder during cutting.

Answer 55

Plastic Embedding Rings and Base Mold

Question 56

3 types of disposable embedding molds

Answer 56

Peel-away Plastic Ice Trays Paper Boats

Question 57

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. It may be placed directly in the chuck or block holder of the microtome.

Answer 57

Peel-Away

Question 58

such as those used in ordinary refrigerators may be recommended for busy routine laboratories. Each compartment may be utilized for embedding one tissue block, which may then be removed by bending once the wax has solidified or by smearing the inner mold with glycerin or liquid paraffin before embedding

Answer 58

Plastic Ice Trays

Question 59

- are normally utilized for embedding celloidin blocks but are equally useful for paraffin wax blocks - They have the advantage of being cheap and easy to make. - They provide easy and accurate identification of specimen, thereby avoiding confusion and interchange of tissue blocks.

Answer 59

Paper Boats

Question 60

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

Answer 60

Celloidin or Nitrocellulose Embedding Method

Question 61

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 (obsolete)

Answer 61

Double-Embedding

Question 62

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 and bone marrow biopsies

Answer 62

PLASTIC (RESIN) EMBEDDING

Question 63

3 types of Epoxy

Answer 63

1. Bisphenol A (Araldite) - slow 2. Glycerol (Epon) - low viscosity 3. Cyclohexene dioxide (Spurr) - very low viscosity

Question 64

- for electron microscopy - most widely applied - carcinogenic due to vinyl cyclohexane dioxide (VCD)

Answer 64

Epoxy

Question 65

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

Answer 65

Polyester plastics

Question 66

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

Answer 66

Acrylic plastics

Question 67

popular embedding medium for light microscopy because it is extremely hydrophilic, allowing many staining methods to be applied, yet tough enough when dehydrated to section well on most microtomes.

Answer 67

Polyglycol methacrylate (GMA)

Question 68

- has found an increasing number of applications for the embedding of biological tissue for transmission electron microscopy (TEM), for the preservation and observation of fine structure not previously subjected to solvent dehydration. - forms only non-crosslinked straight chains on polymerization and therefore requires no hardener

Answer 68

glycol methacrylate (GMA)

Question 69

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

Answer 69

Benzoyl peroxide

Question 70

widely used because of its hardness as the ideal embedding medium for undecalcified bone and is widely used for bone histomorphometry and bone marrow hematopathology.

Answer 70

methyl methacrylate (MMA)