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MLSCI250 HT LAB > LAB 2 Sectioning (Microtomy) > Flashcards

LAB 2 Sectioning (Microtomy) Flashcards

1
Q

Differentiate the course wheel and fine wheel

A
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2
Q

Function of the Micron Thickness Selector

A
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3
Q

Function of the Clearance Angle Adjustment

A

Adjusts the clearance angle between the tissue face and the lower facet of the knife wedge

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4
Q

Function of the Angle Adjuster Screws

A
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5
Q

What is the ideal clearance angle ?

A

3 to 8 degrees

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6
Q

What is the consequence to a clearance angle that is too shallow (knife is parallel to the tissue) ?

A

The pressure applied to the tissue is unstable, which results in alternating thick and thin sections
- this is dependent on factors like tissue and wax properties

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7
Q

What is the consequence to a clearance angle that is too steep (knife is perpendicular to the tissue) ?

A

The tissue will bend sharply before breaking off
- chattering will be seen microscopically (parallel lines like Venetian lines)

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8
Q

Waterbath temperature

A

5 to 10 degrees below melting temperature of the paraffin wax

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9
Q

T or F: the water bath has limited use in removing wrinkles from the tissue itself

A

TRUE; the water bath has limited use in removing wrinkles from the tissue itself

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10
Q

Since waterbaths are a source of carryover, what can be done to limit this ?

A

The surface should be cleaned between every sample using a tissue

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11
Q

Micron setting used for rough cutting (trimming)

A

20 to 25 microns

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12
Q

Which direction do you rotate the flywheel ?

A

Clockwise (away from you)

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13
Q

Consequence of the water bath being too hot

A

The ribbon will expand too quickly and tear sections apart

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14
Q

Consequence of the waterbath being too cold

A

Tissue section won’t flatten out

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15
Q

Why would the ribbon and consecutive sections be curved ?

A
  • face of block is not completely flat
  • knife blunt in one area
  • tissue has variable consistency
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16
Q

How to remedy ribbons that are curved ?

A
  • trim excess wax to make parallel
  • use a different part of a knife, or replace knife
  • vary speed of cutting or reorient block 90°
17
Q

Why would tissue sections alternate from being thick and thin ?

A
  • wax is too soft for tissue
  • block or knife is too loose
  • insufficient clearance angle (too shallow)
  • mechanism of microtome is faulty
18
Q

How to remedy tissue sections alternating from being thick or thin ?

A
  • cool block with ice
  • re-embed tissue with higher melting point wax
  • increase clearance angle gradually
  • check microtome for faults/ fix or send for maintenance/ replace microtome
19
Q

What causes chatters ?

A
  • Block or knife is loose
  • Knife angle is too steep
  • Tissue is unusually hard
20
Q

How to remedy chatters ?

A
  • tighten holder
  • reduce angle gradually
  • use a heavy duty microtome with a heavy duty knife
  • use softening agent on tissue
  • cut slowly at a thinner thickness (2 to 3 micrometers)
21
Q

Why would scoring or splitting occur at right angles to the knife edge ?

A
  • There’s a nick on the knife edge
  • Hard particles in tissue or wax
22
Q

How to remedy scoring or splitting at right angles to the knife edge ?

A
  • use different part of knife, or replace knife
  • remove hard particles
  • re-embed tissue
23
Q

Why would tissue sections not form a ribbon ?

A
  • wax is too soft for cutting conditions
  • debris on knife edge
  • incorrect knife angle
  • cutting above optimal speed
  • sections are too thick
24
Q

How to remedy sections not forming a ribbon ?

A
  • cool block
  • remove block and clean areas around knife
  • adjust knife angle
  • adjust cutting speed
  • reduce micron setting
25
Q

Why would tissue sections become attached to block on the return stroke ?

A
  • debris on knife edge
  • top edge of tissue is not smooth or has debris
  • clearance angle is insufficient (too shallow)
  • static electricity on ribbon
26
Q

How to remedy sections becoming attached to block on the return stroke ?

A
  • remove block and clean areas around knife
  • lock microtome flywheel and use a gloved thumb to run back and forth along edge of block
  • increase clearance angle slightly
  • humidify or ionize air to remove static electricity
27
Q

Why would areas of tissue be missing in the ribbon sections ?

A
  • incomplete impregnation of tissue (ie. tissue not flat against mold )
  • insufficient rough cutting
28
Q

How to remedy areas of tissue not being present in ribbon sections ?

A
  • return tissue to paraffin wax for at least one hour
  • set micron selector to 20 and turn wheel until full face of tissue is exposed, then re-cool block
29
Q

Why would tissue sections become excessively compressed ?

A
  • cutting too quickly
  • wax is too soft
  • knife is too dull
30
Q

How to remedy excessive compression of tissue sections ?

A
  • slower, more uniform cutting
  • cool block
  • use different part of knife, or replace knife
31
Q

Why would sections expand and disintegrate on the water surface ?

A
  • water temperature too high
  • poor impregnation of tissue
32
Q

How to remedy sections expanding or disintegrating on water surface ?

A
  • decrease water temperature
  • return tissue to wax for at least one hour
33
Q

Why would sections roll into a tight coil instead of remaining flat ?

A
  • section is too thick
  • clearance angle is insufficient (too shallow)
34
Q

How to remedy sections from rolling into a tight coil instead of remaining flat ?

A
  • ensure micron selector is set properly
  • reduce knife tilt
35
Q

Why would sections stick to the knife pressure plate ?

A
  • dirty pressure plate
36
Q

How to remedy sections sticking to the knife pressure plate ?

A
  • clean plate with xylene/ xylene substitute, or kimwipe with pressure
  • when picking up ribbon, use an upward sweeping motion
37
Q

Why would the knife gouge out parts of the block ?

A
  • Flywheel turned before ensuring block can clear the knife edge
  • Knife or tissue is loose
38
Q

How to remedy knife from gouging out part of the block ?

A
  • remove block and use scalpel to create a straight bottom edge of cassette
  • if there is no layer of wax left before the tissue, the block must be re-embedded
  • ensure tissue block and knife are tightened properly

MLSCI250 HT LAB (5 decks)

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