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HISTOPATHOLOGY > CHAPTER 13 CUTTING SECTIONS > Flashcards

CHAPTER 13 CUTTING SECTIONS Flashcards

1
Q

is a process whereby tissues are cut into uniformly thin slices or “sections” with the aid of a microtome, to facilitate the studies under the microscope.

A

Sectioning

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2
Q

Three general types of tissue sections may be made:

A
  1. PARAFFIN SECTIONS
  2. CELLOIDIN SECTIONS
  3. FROZEN SECTIONS
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3
Q
  • for paraffin embedded tissue blocks which may be cut by rocking and rotary microtome.
A
  1. PARAFFIN SECTIONS
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4
Q
  • for celloidin embedded tissues which are usually cut by means of the sliding microtome.
A
  1. CELLOIDIN SECTIONS
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5
Q
  • which may be cut from tissues that have been fixed and frozen with CO2 or for fresh or fixed tissues frozen with the cryostat.
A
  1. FROZEN SECTIONS
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6
Q

Once the tissues have been embedded and the wax has solidified, the
wax block is removed from the mold, the identification number is noted and the excess wax is cut off from the block to expose the tissue surface in preparation for actual cutting.

A

TRIMMING

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7
Q

Only thin slices are taken out at a time to prevent the block from cracking.

A

TRIMMING

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8
Q

Depending upon the size and orientation of the tissue sample, shave conservatively into the block surface taking appropriate cuts that may measure between

A

4-60 micrometers.

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9
Q

Coarse facing is done on the microtome at approximately [?] at a time until the entire tissue surface is exposed.

A

30 microns

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10
Q

After [?], a heated spatula is held between the tissue block and the block holder until the wax begins to melt.

A

coarse trimming

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11
Q

The block is allowed to harden for cutting proper by facing them down in ice cold water or refrigerator for

A

5-10 minutes.

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12
Q

Fine trimming may be done by either setting the thickness adjuster at [?] or by advancing the block using the coarse feed mechanism.

A

15 mm

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13
Q

The knife is usually tilted at [?] angulation on a microtome to allow a clearance angle between the cutting facet and the tissue block.

A

0-15°

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14
Q

require smaller clearance angles than wedge-shaped knives.

A

Biconcave knives

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15
Q

Sections are cut between [?] in thickness for routine histologic procedures, after the block has been fixed and secured to the block holder.

A

4-6μ

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16
Q

provides better support for the harder elements in a specimen allowing thinner sections to be obtained.

A

Cold wax

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17
Q

wax provides better support for the harder elements in a specimen allowing thinner sections to be obtained.

A

Cold

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18
Q

The sections are then floated out on a water bath set at [?], approximately [?] lower than the melting point of the wax used for embedding the tissue.

A

45-50°C

6-10°C

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19
Q

Sections should not be left on the water bath for a long time ([?] will be enough) to avoid undue expansion and distortion of tissue.

A

30 seconds

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20
Q

Sections may also be flattened out by placing them on a slide which has been flooded with [?], producing convection currents which will serve to remove the creases in the tissue within a few seconds.

A

20% alcohol

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21
Q

should expand the section to its original dimensions and ensure that it is completely flat.

A

Flotation

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22
Q

Flotation
The temperature will need to be [?] below the melting point of the wax.

A

5 - 9 ̊C

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23
Q

The mounted section is then placed in a [?] paraffin oven for [?] or until water droplets are no longer visible on the slides.

A

70oC

20 minutes

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24
Q

Besides the paraffin oven which is maintained at a temperature of [?] above the melting point of the paraffin used, small thermostatically controlled incubators may be used, regulated at [?], and at [?], for enzyme digestion, chemical extraction, metallic impregnation and enzyme localization techniques.

A

2-5°C

37°C

45-55°C

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25
Q

Dry sections for between [?]. Some delicate specimens will produce best results when dried at [?] for a longer time (several hours to overnight).

A

5 and 30 minutes

37oC

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26
Q

Metal racks with 25-slide divisions are used to store the mounted sections during the drying process which usually takes about [?] in the heated oven.

A

5 minutes

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27
Q

Extended storage (?) of unstained formalin-fixed paraffin embedded slides should be avoided as this may result in the loss of antigens.

A

usually more than 3 days

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28
Q

Surfaces and edges of the block are not parallel

A

Re-trim the block

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29
Q

Horizontal surface of the block is not parallel to the knife

A

Re-adjust and re- orient the block

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30
Q

Paraffin wax is too hard

A

Coat horizontal edges of the block with wax of lower melting point

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31
Q

Knife is tilted too much

A

Reduce the tilt

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32
Q

Sections are too thick

A

Readjust the thickness of the sections

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33
Q

Knife is dull

A

Hone and strop

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34
Q

Knife is blunt

A

Sharpen the knife

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35
Q

Tilt of knife is too great

A

Reduce the tilt

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36
Q

Knife edge is dirty

A

Clean the knife edge

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37
Q

Blunt or dull spot on the knife, producing an irregular knife edge

A

Adjust the knife so that knife edge will present a uniformly sharp edge to the block, or sharpen

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38
Q

Edges of the block are not parallel but round or wedge shaped

A

Re-trim the block

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39
Q

Knife is not parallel to the block

A

Readjust the knife and block

40
Q

Knife is blunt or dull

A

Re-sharpen the knife

41
Q

Paraffin block is warm and soft

A

Cool the block on ice water until firm

42
Q

Knife edge is coated with paraffin

A

Clean the knife edge

43
Q

Sections are too thin

A

Readjust thickness of the section

44
Q

Microtome set screw is loose

A

Tighten the screw

45
Q

Tilt of knife is too vertical

A

Reduce the tilt

46
Q

Bevel of knife is lost due to incorrect sharpening

A

Re-sharpen, using a knife back or automatic knife sharpener

47
Q

Bubble or dirt formed in the embedding medium

A
48
Q

Tissue is not processed properly and will not form a section (especially if center is raw)

A

Re-process tissue

49
Q

Under-processed portion of tissue bursts on contact with warm water

A

Re-process tissue

50
Q

Hard spot in tissue due to calcium

A

Once embedded in paraffin wax, decalcification is impractical; use a base-sledge microtome with a wedge knife

51
Q

Tilt of knife is too great or bevel is not cleared, hence object is compressed against the knife edge

A

Reduce the tilt

52
Q

Clamp set screw on knife or block holder is loose

A

Tighten the screw

53
Q

Blocks are too large

A

Cut blocks into smaller fragments

54
Q

Blocks are too hard

A

Soften the blocks in detergent or phenol

55
Q

Static electricity due to low atmospheric humidity

A

Breathe out or blow gently on the bock and knife to break up static electricity, or boil water in the room to increase humidity

56
Q

Knife edge is dirty

A

Clean the knife edge

57
Q

Knife edge is dull

A

Sharpen the knife

58
Q

Knife tilt is too great

A

Reduce the tilt

59
Q

Nicks or damage on the knife edge

A

Sharpen the knife

60
Q

Dirty embedding

A

Re-embed in freshly filtered wax

61
Q

Knife edge is dirty

A

Clean knife edge with xylene

62
Q

Tilt of knife is too great

A

Reduce the tilt

63
Q

Knife tilt is too great

A

Reduce the tilt

64
Q

Knife is dull

A

Sharpen the knife

65
Q

Paraffin is too soft or room temperature is warm

A

Cool paraffin wax in ice water

66
Q

Tilt of knife is too small, paraffin block is therefore compressed against the base of the knife towards the end of stroke

A

Increase the tilt

67
Q

Knife edge vibrates due to hardness of tissue

A

Treat with phenol during processing or collodionize

68
Q

Tilt of knife is too great

A

Reduce the tilt

69
Q

Knife is blunt

A

Sharpen the knife

70
Q

Knife is not clamped properly

A

Adjust the knife so that knife edge will present a uniformly sharp edge to the block, or sharpen

71
Q

Knife or block holder is loose

A

Tighten adjusting and locking screws

72
Q

Knife tilt is too small that block is compressed by bevel and section is not cut

A

Increase the tilt

73
Q

Tilt of knife is too slanted or too big

A

Readjust the tilt

74
Q

Tissue is too hard

A

Take fresh block treated with phenol during processing

75
Q

Knife blade is too thin

A

Change the knife

76
Q

Freezing is not adequate

A

Refreeze the tissue block

77
Q

Tissue is frozen too much

A

Warm the tissue with the fingers

78
Q

Top and bottom edges of block are not parallel to edge of blade/sides of block are not perpendicular to the blade

A

Adjust the block holder to make the block edges parallel to the knife

79
Q

Wrong micrometer setting

A

Microtome needs recalibration

80
Q

Clearing agent not completely removed due to insufficient impregnation

A

Block is trimmed down nearest to the tissue. Remaining wax is melted on embedding oven and paraffin impregnation is repeated, changing the paraffin at least once before embedding

81
Q

Insufficient clearing

A

Block is trimmed down nearest to the tissue.

82
Q
A

Repeat clearing; if object has already been embedded, prolong clearing up to 12 hours, then re- embed

83
Q

Insufficient dehydration, therefore incomplete clearing and impregnation

A

Repeat the whole procedure

84
Q

Contaminated wax

A

Re-embed in freshly filtered wax

85
Q

Block not cooled rapidly enough

A

Re-embed in freshly filtered wax

86
Q

Insufficient paraffin

A

X
Repeat paraffin impregnation, then re-embed

87
Q

is a slow process, usually taking weeks, and does not
produce sections as thin as those produced by paraffin embedding.

A

Celloidin embedding

88
Q

The advantage is that it completely avoids the use of heat at any stage.

A

Celloidin embedding

89
Q

As a consequence, heat produced artifacts are avoided.
In particular, shrinkage is absolutely minimal, if there is any, and structural relationships of the various types of tissue components can be seen clearly.

A

Celloidin embedding

90
Q

The disadvantages are the longer time to cut, the thickness of the sections, the necessity for staining to be done on free floating section, the inconvenience of having to store the blocks in sealed jars with tight lids to prevent complete evaporation of 70% ethanol, the resulting restrictions on the type of staining methods that may be used.

A

Celloidin embedding

91
Q

Celloidin is used in form of solution, usually in a 1:1 mixture of ethanol-ether at concentrations of [?].

A

2%, 4% and 8%

92
Q

The fastest way to dissolve celloidin is to soak it first in half the final volume of anhydrous ethanol to soften it (?) with intermittent mixing in a tightly stoppered container.

A

50 mL for each 8 grams celloidin

93
Q

The next day, an equal volume of [?] is added and intermittently mixed until an evenly consistent solution is obtained.

A

diethyl ether

94
Q

The [?] may then be made by simple dilution of the [?] with an equal parts mixture of ethanol and diethyl ether.

A

2% and 4% solutions

8% solution

95
Q

The block is then trimmed, leaving about [?] of celloidin all around the tissue, then a distinctive cut is made on one corner for orientation.

A

3 -5 mm

96
Q

The knife is held at a significant slant so that most of the blade edge is used during the cutting stroke, and is quite long, often in excess of [?].

A

25 cm

HISTOPATHOLOGY (14 decks)

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