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Histopathology > Practice Questions > Flashcards

Practice Questions Flashcards

1
Q

The organisation responsible for accrediting diagnostic histopathology laboratories in Australia is the:

a) National Association of Testing Authorities
b) Royal College of Pathologists of Australasia
c) National Association of Testing in Australia
d) National Pathology Accreditation Advisory Council
e) Australian Institute of Medical Scientists

A

A - National Association of Testing Authorities

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2
Q

The optimal tissue section thickness for specimens prepared for electron microscopy is:

a) 2-5 μm
b) 30-50 μm
c) 60-90 μm
d) 2-5 nm
e) 60-90 nm

A

E - 60-90nm

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3
Q

Name the 8 types of staining.

A
  1. Non-vital
  2. Vital/phagocytic
  3. Histochemical
  4. Lysochrome
  5. Impregnation
  6. Injection
  7. Fluorochrome
  8. Immunostaining
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4
Q

Describe non-vital staining.

A

Staining of dead tissue that’s been fixed, processed and sectioned. (Bulk of routine staining - HandE)

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5
Q

Describe vital/phagocytic staining.

A

Living tissues/cells, less toxic because of dilute dyes or phagocytic action of macrophages ingesting the dye particles. (Eg. Janus Green)

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6
Q

Describe histochemical staining.

A

Used to visualise biological structures. Tissue must be fixed. (Eg. PAS)

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7
Q

Describe lysochrome staining.

A

Stains neutral lipids and fats. Stains dissolve into fats. (Eg. Sudan Black and Oil O Red)

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8
Q

Describe impregnation staining.

A

Example is reticular staining with the use of silver salts. The silver salts are reduced to form black coating/deposit around the individual fibres. They require a reducing agent. (Eg. Reticular stains)

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9
Q

Describe injection staining.

A

Not technically a stain but classed as one. Injection of coloured compound (not always a dye) to highlight structures. (Eg. Latex dye)

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10
Q

Describe fluorochrome staining.

A

Fluorescent molecules combined with a tissue entity is visualised under fluorescent light. (Eg. Fluorescent antigen technique)

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11
Q

Describe immunostaining.

A

Antibody-antigen labelling. A labelled antibody is used. (Eg. Direct and indirect IHC).

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12
Q

What is a chromophore?

A

A colour bearer. Atomic groupings which are required for a stain to work. The more chromophores within a compound, the deeper the colour.
Eg. Quinonoid, Azo and Nitro groups

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13
Q

What is an auxochrome?

A

Ionizing group which gives the dye a net positive charge.
It assists the dye to bind to the tissue itself. It can also intensify the colour.

  • If auxochrome gives net +ve charge = basic/cationic dye
  • If auxochrome gives net -ve charge = acidic/anionic dye

Eg. Hydroxyl, methyl and amino groups.

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14
Q

What are 7 stains for liver biopsies and what do they test for?

A
  • H and E: general structure
  • Orcein: Hep B antigens
  • PAS: Glycogen storage
  • PAS-D: Neutral mucins, (adenocarcinomas secrete neutral mucins.)
  • Reticulin: Reticular fiber general tissue architecture and changes therein = cirrhosis.
  • Perls: Iron deposits and haemachromatosis
  • Van Gieson: Collagen increased in cirrhosis.
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15
Q

What is a modifier?

A

They are additions of various side groups to the dye molecule which can enhance or change the dye colour. Eg. Eosin B or Eosin Y

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16
Q

What is a leuco compound?

A

A colourless solution which destructs the chromophore (reduces) to make the compound more soluble. They are oxidised in a subsequent reaction to add colour.
They produce a colour on contact with a certain tissue element that it’s trying to bind to.
Eg. Schiff’s reagent

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17
Q

What is the difference between regressive and progressive staining?

A

Regressive is when you OVER stain and then differentiate by removing excess stain. Progressive staining is when stain applied until the desired intensity of tissue coloration is attained.

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18
Q

What is a trapping agent?

A

A chemical which inhibits removal of dyes from tissues. Eg. Iodine in gram staining

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19
Q

What is differentiation?

A

Removal of excess stain (as in regressive staining) or using differentiation agents to distinguish between multiple tissue components.
Differentiation agents include mordants, acids and oxidising agents.

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20
Q

What can depth of colouration be affected by?

A
  • Chemical affinity
  • Density
  • Permeability and diffusion of dye.
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21
Q

What is trichrome staining?

A

A staining method that uses two or more acid dyes in conjunction with a polyacid. Three different dyes of 3 different particle sizes will penetrate the tissue differently depending on density, chemical affinity and permeability.

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22
Q

Why use a weak positive control?

A

A weak positive is going to show up a result, but only just and this means our tests will be very sensitive.

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23
Q

Which stain has a copper atom at its centre?

A

Toluidine blue

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24
Q

What is the difference between antigen and antibody?

A

Antigens are molecules capable of stimulating an immune response.
Antibodies are Y shaped proteins produced by B cells of the immune system in response to exposure to antigens.

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25
Q

How does formalin work?

A

It works by cross linking the amino acid side chains within the tissue.

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26
Q

What are 4 benefits of IHC autostaining?

A
  • Standardisation
  • Onboard antigen retrieval
  • Integrated IT system
  • Digital image capture
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27
Q

What are the benefits of using IHC stains?

A
  • Sensitive and specific
  • Results are more consistent/highly reproducible
  • Doesn’t require tissue to be fresh
  • Can study almost any Ab
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28
Q

What is a sentinel node?

A

The closest lymph node to a tumour is usually the 1st site of metastases.

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29
Q

Ideal fixative type?

A

Generally 10% neutral buffered formalin

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30
Q

What is an advantage of using microwave HIER (Heat induced epitope retrieval)?

A

It’s very cheap

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31
Q

What are two disadvantages of using microwave HIER (Heat induced epitope retrieval)?

A
  • The time depends on the microwave wattage

- Hot and cold spots

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32
Q

What are some advantages of using pressure cooker HIER (Heat induced epitope retrieval)?

A
  • Very few hot and cold spots
  • No evaporation
  • Can accommodate large number of slides
  • Short incubation period at higher boiling point (120C)
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33
Q

What are some disadvantages of using pressure cooker HIER (Heat induced epitope retrieval)?

A
  • It’s slow to heat up and cool down

- Potential hazards

34
Q

What is an advantage of using water bath HIER (Heat induced epitope retrieval)?

A
  • Low investment needed
35
Q

What are two disadvantages of using water bath HIER (Heat induced epitope retrieval)?

A
  • Difficult to control temperature (hot and cold spots like with microwave
  • Not suitable for large numbers of slides
36
Q

What is the primary role of IHC?

A

To demonstrate morphological appearance of cell and tissue architecture.

37
Q

What is ischemia time?

A

The time the sample is taken out of surgery and prior to placement into fixative. Ideally no longer than an hour.

38
Q

True or False?
With proteolytic digestion antigen retrieval method, length of time in proteolytic solution is proportional to length of time in fixative?

A

False. It is NOT proportional.

39
Q

What factors affect metachromasia?

A
  • Increasing concentration of dye
  • Lowering temperature
  • Lowering pH
  • Increasing bonding (water can be used for this as it’s polar)
40
Q

Which IHC method is more sensitive? Direct or indirect?

A

Indirect

41
Q

Define “malignancy”.

A

A change in normal cell biology of cells. They grow rapidly and will often spread (metastasize).

42
Q

What are the 5 tumour groups?

A
  • Carcinoma (Variation: Adenocarcinoma)
  • Lymphoma (Variation: Leukemia)
  • Melanoma
  • Mesothelioma
  • Sarcoma (Variation: Liposarcoma from adipose tissue)
43
Q

What are two advantages of using “antibody cocktails”?

A
  • Fewer sections required

- Have relevance in specifically defined areas.

44
Q

What does “in situ” mean?

A

In it’s original place.

45
Q

Name some common antibodies used in IHC.

A
  • Cytokeratins (CK7, CK5, CK6)
  • CD antibodies (cluster of differentiation)
  • Smooth muscle actin
  • LCA
  • HER2
  • p16, p53, p63
  • Broad spectrum cytokeratin
  • S100
  • Vimentin
46
Q

Why create a control using the test section but omitting the primary antibody in IHC?

A

To rule out endogenous enzyme activity or non-specific absorption of other reagents.

47
Q

Thickness of sections for light microscope?

A

4um

48
Q

Thickness of sections for electron microscope?

A

60-90nm

49
Q

What is Koehler illumination?

A

Efficient, bright and even illumination.
It is the first step and most crucial step in setting up a microscope to minimize any internal stray of light from occurring.

50
Q

Give an example of a leuco dye?

A

Schiff’s reagent

51
Q

Polarisation microscopy

A
  • Need a polariser, detector and an analyser
  • When you have a crystal, it will scatter the light and the detector/analyser will pick up where the light scatters to.
    USED FOR: uric acid crystals (in gout) and amyloid (glows green).
52
Q

Bright field microscopy

A
  • Most common in labs
  • Uses Koehler illumination for a uniform view across the slide.
    USED FOR: routine special stains
53
Q

Phase contrast microscopy

A
  • Need a condenser and phase plate
  • Changes amplitude and thus affects the brightness.
    USED FOR: RBCs in urine, sperm motility, living/unstained tissues.
54
Q

Fluorescence microscopy

A
  • Need proper filters to excite the particles and make them fluoresce.
  • Need fluorophores
    USED FOR: IHC, autofluorescence and FISH (fluorescence in situ hybridisation)
55
Q

Confocal microscopy

A
  • Similar to fluorescence but has higher resolution.
  • You can go through the entire sample.
  • VERY expensive so only used in research labs.
    USED FOR: Research
56
Q

Darkfield microscopy

A
  • Need a darkfield condenser
  • The light scatters at acute angles and reaches specimen to allow you to see it.
    USED FOR: Syphilis
57
Q

TRUE OR FALSE?

Light emitted from a single source can be regarded as a series of waves travelling in straight lines in all directions.

A

TRUE

58
Q

What is the purpose of the condenser?

A

To focus light produced by the lamp onto the specimen giving maximum and even illumination

59
Q

Inverted microscope

A

Unstained samples and live cultures.

Can accommodate tissue culture flasks or petri dishes

60
Q

Dissection/stereo microscope

A

Commonly used for renal core biopsies to identify presence of glomeruli.
Allow for fine manipulation of samples.

61
Q

What are the advantages and disadvantages of multidiscussion (multiheader) microscopes?

A

Advantages: Allow multiple people to view a specimen simultaneously so good for teaching.

Disadvantages: Old school. Take up a lot of room and can get the same outcome from sharing the image on a high definition TV screen.

62
Q

What are the advantages and disadvantages of digital image capture?

A

Advantages: Allows you to capture images straight from the microscope and you can feed it straight to a screen.

Disadvantages: You’re restricted by the quality of the camera.

63
Q

What are the advantages and disadvantages of telemicroscopes?

A

Advantages: The microscopes have a camera you can relay the image to someone over the internet. Great when you have urgent samples that you need to show a specialist.

Disadvantages: The person locally is controlling the microscope, the person over the phone is viewing and is saying “move, up, down, stop” etc. Time consuming.

64
Q

What are the advantages and disadvantages of slide scanning?

A

Advantages: Allows you to place slides into a tray and view them on a computer. You can share the images and someone remotely can control the machine. Helps with slide storage (as opposed to physical storage).

Disadvantages: Limiting factors are only the screens definition that you can look at them with. Also issues with patient confidentiality and paying for digital storage.

65
Q

All of the following are used to stain carbohydrates except:

a) PAS
b) Alcian Blue
c) Orcein
d) Toluidine Blue
e) IHC

A

C - Orcein

66
Q

What is an alternative fixative to glutaraldehyde and osmium tetroxide for TEM sections if you are also wanting to do IHC?

A

Paraformaldehyde

67
Q

Polarizing microscopy is useful for:

a) assessing HandE stained sections
b) Assessing sperm motiility
c) Identification of Troponema pallidum
d) Identification of autofluorescent molecules
e) Identification of uric acid crystals

A

E - Identification of uric acid crystals.

68
Q

Metachromasia results from:

a) Uptake of dye into living cells
b) Polymerisation of some basic dyes
c) Impurities within some dyes
d) The action of a trapping agent
e) The use of a mordant

A

B - Polymerisation of some basic dyes

69
Q

Antigen retrieval is required to:

a) Unmask antigenic sites
b) Unmask antibody sites
c) Form new methylene bridges
d) Form new protein cross links
e) Preserve immunoreactivity in alcohol fixed tissue

A

A - Unmask antigenic sites

70
Q

The ionising group that enables binding of a dye to tissue is termed a/an:

a) Amino group
b) Auxochrome
c) Chromophore
d) Modifier
e) Leuco compound

A

B - Auxochrome

71
Q

Could a glass lens be used to focus the electron beam in a TEM?

A

No, because glass would stop the block of electrons.

72
Q

TRUE or FALSE?

Chromatic aberration, spherical aberration and astigmatism all occur in electron microscopy too.

A

TRUE

73
Q

Is electron microscopy good for tumour diagnosis?

A

It was first thought to be but it isn’t. IHC is better

74
Q

What are the 3 applications for TEM?

A
  • Tumour diagnosis
  • Renal disease (glomerulus is principle structure to be examined)
  • Muscle biopsy
75
Q

A broad-spectrum cytokeratin antibody could be used to identify:

a) Lymphoma
b) Melanoma
c) Carcinoma
d) Sarcoma
e) Nerve-derived tumour

A

C - Carcinoma

76
Q

Which of the following haematoxylin stains is naturally ripened?

a) Harris’
b) Mayers’
c) Delafields
d) Weigert’s
e) Cole’s

A

C - Delafields

77
Q

What are the names of the two organisations responsible for accreditation and advisory for histopathology lab standards?

A

NPAAC - National Pathology Accreditation Advisory Council

NATA - National Association of Testing Authorities

78
Q

In electron microscopy are cells commonly fixed in paraformaldehyde?

a) True
b) False

A

FALSE

79
Q 
Schiffs reagent is known as a/an : 

a) Basic Dye 
b) Leuco Dye
c) Acid Dye 
d) Modifier 
e) Leuco compound
A

E - Leuco compound

80
Q 
Which of the following stains will detect Melanin? 

a) Masson Fontana
b) Perls
c) Sudan black
d) Rhodamine
e) Grimelius
A

A - Masson Fontana

Histopathology (3 decks)

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