Basics from Practice

Question 1

Describe the sections of the NFPA diamond

Answer 1

Question 2

What fixative is used for the preservation of enzymes and cell surface antigens?

Answer 2

Acetone

Question 3

What is miscible with hydrocarbons?

Answer 3

Paraffin is the only reagent that is miscible with hydrocarbons (xylene)

Question 4

What may cause nuclear bubbling?

Answer 4

Incomplete fixation before processing
Microwave drying of slides
Insufficient draining of slides before drying

Question 5

What pigments reduce silver?

Answer 5

Formalin (reducing substance) –> may give a + reaction with Fontana-Masson & Schmorl

Question 6

Urate crystals are demonstrated with?

Answer 6

Gomori methenamine silver method turns urate crystals black

Question 7

ATPase stains were performed at pH 9.4 and 4.3. Each of these stains shows large type grouping of both dark and light staining fibers. The patterns of dark and light stained fibers are reversed at the different pHs. This result is most likely due to:

Answer 7

Large type grouping is indicative of a neuropathic disease process.

Question 8

What pH should a solution containing hydrochloric acid be before it is discarded in the sink?

Answer 8

between pH 3 and pH 11

Question 9

What controls are preferred for reticulin procedures?

Answer 9

Normal liver tissue

Question 10

What is demonstrated following the Steiner procedure?

Answer 10

Spirochetes (stained black)

Question 11

What stains are best for the demonstration of the myelin sheath?

Answer 11

Luxol fast blue and Weil stains and sections should be cut at 10-15um

Question 12

What best demonstrates Pneumocystis jirovecii?

Answer 12

Grocott methenamine silver stain
- is the “gold standard” procedure

Question 13

What is the result of using Osmium tetroxide as a fixative?

Answer 13

Fixes fats, interferes with H&E staining

Question 14

What are the characteristics of Bouin solution?

Answer 14

Excellently preserves structure with soft and delicate structures
Lyses RBC
Good penetration
Mordants connective tissues

Question 15

Define the primary function of auxochromes in artificial dyes.

Answer 15

Give the dye affinity for the tissue

Question 16

What is this tissue stained with?

Answer 16

Periodic acid-Schiff stain
- depends on the formation of aldehydes

Question 17

What pigments are birefringent?

Answer 17

Formalin, melanin, hemosiderin, lipofuscin

Question 18

What is the toughest connective tissue fiber?

Answer 18

Collagen

Question 19

How do you calculate the gravimetric factor?

Answer 19

[present dye] / [new dye]

Question 20

Explain reduction and oxidation

Answer 20

A substance that gains hydrogen (e-) is reduced.
A substance that loses hydrogen (e-) is oxidized.

A reducing agent (e- donor) is oxidized in the reaction.
An oxidizing agent (e- acceptor) is reduced in the reaction.

Question 21

In humans, what kind of epithelium lines the esophagus?

Answer 21

Nonkeratinizing stratified squamous epithelium

Question 22

In humans, what kind of epithelium covers the skin?

Answer 22

Keratinizing stratified squamous epithelium

Question 23

In humans, what kind of epithelium lines the bladder?

Answer 23

Transitional, or urothelium

Question 24

In humans, what kind of epithelium lines the trachea?

Answer 24

Ciliated pseudostratified columnar epithelium

Question 25

What do the objectives of psychomotor relate to?

Answer 25

Teaching and learning technical skills

Question 26

A pigment caused by chromate-containing fixatives can be prevented by treating the tissue prior to processing with:

Answer 26

Running water

Question 27

What is the key feature of argentaffin cells?

Answer 27

Their ability to take up silver stains (Fontana-Masson)

Question 28

What is the substance demonstrated?

Answer 28

Glycogen

Question 29

If hematoxylin is unavailable, which of the following dyes is recommended as a substitute in a routine hematoxylin and eosin procedure?

Answer 29

Celestine blue

Question 30

What stain is this?

Answer 30

The methyl green-pyronin technique for differentially staining nucleic acids (DNA: blue to blue-green, RNA: red)

Question 31

What is the light chain found on in some antibodies?

Answer 31

lambda

Question 32

To help maintain morphology in formalin-fixed tissue prior to freezing, what may the tissue be placed in?

Answer 32

The tissue should be infiltrated with 30% sucrose before freezing

Question 33

What is this tissue?

Answer 33

Skeletal muscle

Question 34

How many changes of paraffin wax are recommended for adequate infiltration?

Answer 34

3

Question 35

What are bluing agents?

Answer 35

Convert the hematoxylin stain from a reddish hue to a blue color - Ammonia water - Lithium carbonate solution - Scott tap water substitute

Question 36

What could prevent this problem?

Answer 36

Immediate and prolonged fixation

Question 37

What is Carnoy solution recommended for?

Answer 37

Preservation of nucleic acids

Question 38

What is Clarke solution used for?

Answer 38

Lyse RBC Should be used to treat very bloody cytology smears

Question 39

At what pH does the best eosin staining occur?

Answer 39

4.5 - 5.0

Question 40

Why is Bouin contraindicated for Feulgen stains?

Answer 40

The picric acid in Bouin solution is strong enough to hydrolyze nucleic acid and should not be used if stains for DNA or RNA are anticipated.

Question 41

Can acetone bleach melanin?

Answer 41

No

Question 42

What can correct this cryostat problem?

Answer 42

The holes seen in the cryostat section of brain tissue are ice crystal artifacts caused by slow freezing of the tissue; more rapid freezing will prevent this artifact.

Question 43

What is the differential staining of nuclei and cytoplasm with Giemsa solution an example of?

Answer 43

Polychromasia

Question 44

Improper washing after which fixative will form an insoluble pigment in tissue?

Answer 44

Zenker fluid (due to the chromium trioxide)

Question 45

Why do you not want to freeze tissue slowly?

Answer 45

It is likely to show disruption in morphology caused by large ice crystals. This is especially evident in skeletal muscle biopsies.

Question 46

What technique is used?

Answer 46

alkaline phosphatase, identified by the red color of the chromogen

Question 47

What may coolant sprays cause during sectioning?

Answer 47

Coolant sprays used on a block during microtome may cause cracking of the block, which will result in a parched earth appearance in the section.

Question 48

How do you differentiate granulocytes?

Answer 48

Romanowsky type stain

Question 49

What kind of stain will reliably type skeletal muscle fibers?

Answer 49

ATPase is the only reliable stain to show the difference between type I and type II fibers

Question 50

What is the best freezing method to demonstrate muscle enzymes?

Answer 50

Isopentane chilled to –150°C

Question 51

Why is beeswax added to some paraffins?

Answer 51

To decrease crystal size

Question 52

What DNA base pairs with adenine?

Answer 52

Thymine

Question 53

What is the oxidizer used in this technique?

Answer 53

The oxidizer in the technique shown (PAS) is periodic acid and it demonstrates neutral mucopolysaccharides

Question 54

What are hematoxylin mordants classified as?

Answer 54

The mordants linking hematoxylin to tissue are generally classified as metallic.

Question 55

What technique is shown here?

Answer 55

Movat pentachrome stain

Question 56

What features can be seen from this image?

Question 57

What is a common use of acetic acid in a fixative?

Answer 57

Counteract the shrinkage caused by another component and coagulate nucleoproteins

Question 58

Which staining technique immerses sections in silver nitrate solutions and exposed to bright light?

Answer 58

von Kossa technique for calcium

Question 59

What does Gill hematoxylin stain?

Answer 59

Mucin stained blue

Question 60

What kind of basic enzyme-reaction is Acid phosphatase?

Answer 60

Hydrolases

Question 61

What are nissl substances?

Answer 61

Granular structures found in the cell bodies of neurons and are composed of large aggregates of granular endoplasmic reticulum with the RNA content - demonstrated after extraction with ribonuclease

Question 62

What is PAS used to stain?

Answer 62

glycogen, neutral polysaccharides, and some glycoproteins

Question 63

What requires sections be cut at 8-10um?

Answer 63

- Amyloid (Lieb crystal violet, Congo red) - Neural stains (Bielschowsky)

Question 64

What is the most common microtomy artifact?

Answer 64

Wrinkles and folds (30% of slides)

Question 65

How can you resolve peripheral chatter?

Answer 65

Soaking block in ice water to counteract overdehydration

Question 66

What thickness should basement membranes be cut at?

Answer 66

2um

Question 68

What is this tissue and how thick should it be sectioned?

Answer 68

Kidney, 2um

Question 69

What effect does positively charged slides have on sectioned tissue?

Answer 69

A chemical reaction occurs leaving the amino groups linked by covalent bonds to the silicon atoms of the glass

Question 70

What is most likely the cause of incomplete cryostat sections?

Answer 70

Incorrect temperature for the tissue

Question 71

What are variable costs?

Answer 71

Indirect or direct costs that vary in direct proportion to test volume.

Question 72

How can overdecalcification impact staining?

Answer 72

Can result in a lack of nuclear staining.

Question 73

What step should occur after completion of decalcification?

Answer 73

Washing the tissue in water to stop the activity of the acid.

Question 74

What will result from bone not being left in decalcification solution long enough?

Answer 74

The H&E will show areas of very dark purple staining due to residual calcium.

Question 75

What can add an overall pink-orange coloration to embedded tissue?

Answer 75

Adding eosin or phloxine to the processing alcohols

Question 76

What is the major disadvantage of aliphatic hydrocarbons in processing?

Answer 76

Intolerance for water and incompatibility with some mounting media BUT are low toxicity & sensitization

Question 77

What are the key features of microwave processing?

Answer 77

- Xylene can't be used - Speeds up processing time - Tissues should be fixed prior

Question 78

What gas is released during decalcification?

Answer 78

Carbon dioxide

Question 79

What can cause precipitated phosphates in tissues?

Answer 79

Buffered formalin that is not rinsed out before the 95% alcohol - can cause difficulties in microtomy

Question 80

What can excessive heat during processing cause?

Answer 80

Hazy blue nuclear staining is the result of the use of heat in the tissue processor alcohol and xylene steps.

Question 81

What is the common name for ethylenediaminetetraacetic acid?

Answer 81

EDTA - chelating agent in decal when pH = 7-7.4

Question 82

How do you remove essential oils (cedarwood oil) from tissue?

Answer 82

Xylene

Question 83

What should not be used for checking the endpoint of decalcification?

Answer 83

Electrolytic procedures

Question 84

Which decalcification method may cause heat damage to the specimen?

Answer 84

Electrolytic methods of decal may cause damage because of the heat generated

Question 85

How can pigment from mercury containing fixatives be removed?

Answer 85

Iodine-sodium thiosulfate

Question 86

What is Zamboni fixative used for?

Answer 86

The buffered formaldehyde-picric acid fixative that is a good dual purpose fixative; it is good for both light and electron microscopy.

Question 87

What is the best fixative for blood smears?

Answer 87

Methanol

Question 88

What fixative must be post treated for mercuric chloride pigment?

Answer 88

B-5 fixative contains mercuric chloride, sodium acetate, and formaldehyde

Question 89

What does Carnoy contain?

Answer 89

Chloroform, glacial acetic acid, 100% alcohol

Question 90

What is Millonig formaldehyde?

Answer 90

A dual-purpose fixative that allows both light and electron microscopy.

Question 91

Paraffin blocks containing tissue fixed in Bouin solution are retrieved from storage after several years. New sections are cut and stained with H&E, and no nuclear staining is present, although the nuclei of the original slides were well stained. To prevent this from happening in the future on tissue to be stored, one must:

Answer 91

Neutralize the picric acid before processing; if allowed to remain in the tissue when embedded, the staining characteristics will change over time

Question 92

What is calcium-formalin recommended for?

Answer 92

The fixation and preservation of phospholipids

Question 93

What about microwave fixation can cause distorted architecture and pyknotic nuclei?

Answer 93

Overheating - carefully controlling the temperature is a must

Question 94

What is commonly added to formalin to preserve immunoreactivity?

Answer 94

Zinc salts (also a protein coagulant and used for ultrastructural studies)

Question 95

What is the osmolality of an isotonic solution?

Answer 95

340 mOsm

Question 96

What fixative irreversibly blocks tissue antigens?

Answer 96

Glutaraldehyde