Microtechniques 3 - Sectioning

Question 1

What are the 2 types of microtomes?

Answer 1

1. Rotary: fixed knife, moveable block (most common)

2. Sledge: moveable knife, fixed block

Question 2

What are the 3 types of knives?

Answer 2

1. Reusable: steel, regular sharpening, used on paraffin blocks
2. Disposable: steel, 3 grades (soft, medium, hard), preferred, used on paraffin blocks
3. Non-metal: glass, sapphire, diamond, can’t be used on paraffin, instead used on plastic blocks in EM

Question 3

5 steps in the sectioning procedure?

Answer 3

1. Trim block
2. Attach to chuck (block holder)
3. Attach to microtome
4. Cut sections
5. Collect ribbons and flatten them on a waterbath

Question 4

What is the CLEARANCE ANGLE?

Answer 4

Angle between block face and lower facet of blade (3-8°)

Question 5

What are 3 ways a specimen can be damaged during sectioning?

Answer 5

1. Knife marks - right angles to knife edge
2. Chatter - paralel to knife edge, when CA is too wide
3. Compression - flattened, squashed, round things appear oval in shape e.g. from cutting from a warm block

Question 6

What are 4 ways a specimen can be damaged during sectioning?

Answer 6

1. Bubbles
2. Cracking - poor fixation
3. Folds
4. Floating artefact

Question 7

2 ways to adhere sections to slides more easily?

Answer 7

1. Subbing using egg white

2. Using charged slides

Question 8

What is dewaxing? Why do we do it? What happens if it isn’t done properly?

Answer 8

Removing wax e.g. using Histoclear, uneven staining if not done properly