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Histo 2 > Exam 1 > Flashcards

Exam 1 Flashcards

1
Q

what are some basic safety procedures to remember?

A
  • blades go in sharps container

- xylene gets drained in a separate container

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2
Q

what are some examples of infectious waste?

A
  • microbiological or culture material
  • pathological material
  • blood
  • sharps
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3
Q

what are some examples of mechanical hazards?

A
  • blades
  • needles
  • scalpels
  • razors
  • glassware
  • electrical equipment
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4
Q

what are examples of health hazards?

A
  • infectious disease
  • carcinogens
  • cryogenic sprays
  • toxins (reproductive)
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5
Q

what are some examples of chemical hazards?

A
  • corrosives
  • irritants
  • sensitizers
  • carcinogens
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6
Q

what are some examples of fire hazards?

A
  • alcohol

- hydrocarbons

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7
Q

what should you see on a properly processed & stained slide?

A
  • nuclei should show a variety of chromatin patterns with crisp blue nuclear membrane
  • cytoplasm should be well preserved & should stain well with eosin
  • NO artifactual spaces between individual cells
  • NO cell shrinkage
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8
Q

what are some causes of delayed fixation?

A
  • specimens obtained long after blood supply has been compromised
  • specimen not open so that fixative can come into contact w/ all surfaces
  • specimen not thinly cut
  • inadequate volume of fixative
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9
Q

what does delayed fixation look like?

A
  • nuclei may show loss of chromatin
  • blue halo
  • fading nuclei
  • disappearance of nuclei
  • cell shrinkage
  • disruption of cytoplasm
  • some cells may completely disappear
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10
Q

what does incomplete fixation look like?

A
  • nuclei is muddy/smudgy

- tissue morphology not well maintained

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11
Q

how is incomplete fixation caused?

A
  • tissue sections not allowed enough time in fixative
  • inadequate amount of fixative relative to tissue volume
  • sections grossed too thick for good penetration
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12
Q

what does excessively dehydrated tissue look like?

A
  • microchatter most commonly seen at the edges of the tissue

- hairline cracks may also been seen

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13
Q

how is excessively dehydrated tissue caused?

A
  • too much time allowed in dehydrating solutions
  • molecularly bound water is removed as well as free water
  • processing biopsy tissues on same schedule as larger specimens
  • processing biopsy tissues on a schedule that is too long
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14
Q

what does poorly processed tissue look like?

A
  • section may appear cloudy with nuclei varying in staining properties
  • no chromatin definition in the nuclei
  • some nuclei may appear very washed out
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15
Q

what causes poorly processed tissue?

A
  • residual water remaining in tissue when they are placed in clearing agent
  • fixative left in tissues when placed in clearing agent
  • incomplete paraffin infiltration
  • too much clearing agent in paraffin
  • too much heat during processing
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16
Q

what does cell shrinkage look like?

A
  • cells are shrunken and there are a lot of artifactual spaces around the cells
  • nuclei appear pyknotic (a nucleus of a damaged cell that has decreased in volume & become darker due to some degree of condensation of nuclear chromatin)
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17
Q

how is cell shrinkage caused?

A
  • inadequate fixation followed by excessive dehydration

- drying before submersion in fixative

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18
Q

what does formalin or mercury pigment in tissue look like?

A
  • brown to black pigments in tissue
  • formalin pigment is most prevalent in bloody tissues
  • usually lie on top of cells (but rarely can occur within cells)
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19
Q

how is formalin or mercury pigment caused?

A
  • formalin solution has a pH below 5.5

- tissue has not been treated to remove mercury pigment

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20
Q

what does nuclear bubbling look like?

A
  • nuclei look as if they contain soapsuds

- chromatin pattern is disturbed and bubbly

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21
Q

how is nuclear bubbling caused?

A
  • incomplete fixation with formalin
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22
Q

what does poor section orientation look like?

A
  • all layers are not demonstrated on tissues that have walls or layers
  • lumen cannot be seen in tubular structures such as fallopian tube or arteries
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23
Q

what causes poor section orientation?

A
  • poor specimen embedding technique

- tissue incompletely fixed and hardened before grossing, so layers have rolled

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24
Q

what are the general steps of H & E staining?

A
  1. ) incubation at 60 degrees C
  2. ) xylene for deparaffination
  3. ) ethanol rinse for rehydration
  4. ) rinse w/ tap water
  5. ) hematoxylin stain & replace xylene vats
  6. ) rinse using hematoxylin rinsing vat
  7. ) ethanol rinse for differentiation
  8. ) eosin stain (counter-stain) & replace ethanol vats
  9. ) rinse with eosin rinsing vat
  10. ) repeat step 7 (dehydration)
  11. ) xylene rinse (clearing)
  12. ) mount with coverslip
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25
Q

what is the cause of nuclei not being crisp or appearing to be smudgy with no distinct chromatin patterns?

A
  • fixation poor or incomplete
  • water was not completely removed during dehydration & clearing
  • slides were exposed to excessive heat during processing or drying
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26
Q

what causes the 3 shades of eosin to not be seen?

A
  • inadequate fixation
  • improper processing occurred
  • poor differentiation of the eosin
  • eosin solution is not at the correct pH
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27
Q

what causes poor contrast?

A
  • nuclear stain is too dark for cytoplasmic stain
  • nuclear stain is too pale for cytoplasmic stain
  • cytoplasmic stain is too dark for nuclear stain
  • cytoplasmic stain is too pale for nuclear stain
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28
Q

what causes the cytoplasmic stain to be too dark?

A
  • exposure of sections to eosin solution is prolonged
  • inadequate eosin differentiation in the alcohols that follow the eosin stain; aqueous formulations stain tissue darker then alcohol-based stains
  • alcohol rinse is not performed properly after the eosin stain
  • eosin may be too concentrated, especially if phloxine is present
  • if using an alcoholic eosin product, water contamination may have interfered with complete differentiation
  • isopropyl alcohol was used as the dehydrating solution (isopropyl does not differentiate eosin within tissue sections in the same manner as ethyl alcohol)
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29
Q

what causes the cytoplasmic stain to be too light?

A
  • eosin solution is exhausted
  • the pH of eosin staining solution is greater than 4.5
  • bluing solution is not completely washed out before the slides were transferred to eosin solution
  • differentiation in diluted alcohols is prolonged
  • alcohol rinse after the eosin stain is incorrectly performed
  • staining time in eosin is too brief
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30
Q

what causes the nuclear stain to be too dark?

A
  • hematoxylin solution is too concentrated
  • too much time was allowed in the hematoxylin solution
  • pH of the hematoxylin solution is incorrect
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31
Q

what causes the nuclear stain to be too light?

A
  • incomplete deparaffinization
  • hematoxylin solution is exhausted or used beyond its shelf life
  • hematoxylin solution is diluted by carryover from a previous water rinse
  • sections are over-differentiated by using overly concentrated acid-alcohol solutions, or sections remained too long in acid-alcohol
  • staining time is too short in the hematoxylin stain
  • tap water is used in the water rinses before and after staining (can act as excess mordant & restrain hematoxylin)
  • poor fixation and/or processing, resulting in tissues that are unable to bind stain
32
Q

what causes uneven eosin or hematoxylin staining?

A
  • section may be thick and thin
  • some solutions are not high enough to cover the entire slide
  • water rinse was not adequate after hematoxylin staining
  • water rinse was not adequate after acid-alcohol (which is necessary to stop decolorization)
  • water rinse was not adequate after bluing
  • alcohol rinse (dehydration) was not adequate after eosin staining
33
Q

what causes a mounting artifact to appear?

A
  • mounting medium has retracted from the edges of the coverslip
  • air bubbles are trapped under the coverslip
  • mounting medium on top of the coverslip
34
Q

what causes red-brown nuclei?

A
  • sections have not been sufficiently blued

- hematoxylin is breaking down

35
Q

what causes blue staining of cellular elements (such as collagen or smooth muscle)?

A
  • pH of hematoxylin solution is greater than 3.0
  • inadequate differentiation
  • slides have spent too much time in dehydrating/differentiation alcohols
  • eosin is contaminated by carryover from the bluing solution
  • acid-alcohol is too weak
  • amount of time spent in acid-alcohol is too short
  • hematoxylin stain formulation is not selective enough
36
Q

what causes dark precipitate scattered throughout the section?

A
  • deteriorated hematoxylin (hematoxylin beyond its expiration date or damaged from improper storage conditions)
  • metallic film on surface solution due to the hematoxylin not being filtered before use
37
Q

what causes bubbles on the tissue section or in tissue spaces?

A

section is not completely dehydrated

38
Q

what causes sections with an overall hazy appearance, or eosin bleeding throughout the section?

A
  • solutions that are contaminated with water from the previous solution
  • sections that were not adequately dehydrated after eosin staining
39
Q

what causes brown “pigment” throughout section & glossy black nuclei?

A

tissue section dried out before coverslip was applied

40
Q

is acetic acid present in regressive H&E staining?

A

yes

41
Q

is acetic acid present in progressive H&E staining?

A

no

42
Q

what is the rate of stain uptake in regressive H&E staining?

A

rapid

43
Q

what is the rate of stain uptake in progressive H&E staining?

A

slow

44
Q

is regressive H&E staining easy to control?

A

no

45
Q

is progressive H&E staining easy to control?

A

yes

46
Q

is differentiation required in regressive H&E staining?

A

yes

47
Q

is differentiation required in progressive H&E staining?

A

no

48
Q

what are the components of hematoxylin?

A
  • dye
  • oxidizer
  • mordant
  • acidifier
  • stabilizer
  • glycerin
49
Q

what is the active dye source in hematoxylin?

A

hematein

50
Q

what is the dye precursor?

A

hematoxylin

51
Q

what does hematoxylin stain?

A

basophilic (negatively charged) components such as nuclear material (nuclei, etc.)

52
Q

what is the oxidizer in hematoxylin?

A

sodium iodate/mercuric oxide

53
Q

what is the purpose of oxidizer in hematoxylin?

A

used to speed up the oxidation process to hematein

54
Q

what is the mordant in hematoxylin?

A

ammonium alum/potassium alum

55
Q

what is a mordant in general?

A

a metal with a valence of at least 2+ charge

56
Q

what is the purpose of a mordant?

A

links the dye to the tissue by means of a covalent and coordinate bond

57
Q

what is the acidifier in hematoxylin?

A

citric acid/acetic acid

58
Q

what is the purpose of an acidifier?

A

adjust pH to extend shelf life

59
Q

what is the stabilizer in hematoxylin?

A

chloral hydrate/absolute alcohol

60
Q

what is the purpose of a stabilizer?

A

help control the rate of oxidation

61
Q

what is the purpose of glycerin in hematoxylin?

A

delays over-oxidation & discourages fungal growth

62
Q

what is eosin stain for?

A

used to demonstrate general structure of tissue & provides contrast to the now stained nuclei

63
Q

what does eosin stain & what kind of dye is it?

A
  • stains positively charged structures in the cytoplasm

- is an acidophilic dye

64
Q

is a mordant required for eosin stain?

A

no

65
Q

what causes wrinkles?

A

not properly stretching tissue on waterbath

66
Q

what causes folds?

A

picking up tissue incorrectly from waterbath onto slide or tissue not adhering properly to the slide & folding during staining

67
Q

what causes Venetian blinding/chatter/wash-boarding?

A

loose or worn microtome parts

68
Q

what causes parched earth?

A

incomplete fixation

69
Q

what causes microchatter?

A

overdehydration or cutting too rapidly

70
Q

what does OCT stand for & what is it for?

A
  • Optimal Cutting Temperature compound

- for cryostat

71
Q

what acidifier is in progressive (Mayer) stain?

A

citric acid

72
Q

what acidifier is in regressive (Harris) stain?

A

acetic acid

73
Q

what causes knife line?

A

nicked/damaged blade

74
Q

what causes muddy/faded nuclei?

A

poor fixation

75
Q

what causes moth-eaten holes?

A

dull knife or poorly processed tissue

Histo 2 (2 decks)

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