MICROTOMY

Question 1

After tissues have been through processing and has been embedded in wax, they are now ready to undergo the process called_____.

Answer 1

microtomy

Question 2

Microtomy, also known as____ or _____, is the process where tissues are cut into uniformly thin slices to facilitate microscopic studies.

Answer 2

sectioning or cutting

Question 3

for most paraffin embedded tissues these are sectioned from ___ to ____

Answer 3

4 to 8 microns

Question 4

The microtome mechanism slowly moves a block into the path of an extremely sharp_____

Answer 4

steel knife

Question 5

Microtome knives may be generally grouped into four types:

Answer 5

(1)standard thick metal
(2) thin disposable blades
(3) glass knives
(4) diamond knives

Question 6

These knives allows sharpening to your liking.

Answer 6

Standard thick metal knives

Question 7

These knives are considered to be expensive.

Answer 7

STMK

Question 8

In the contrary the______ blades do the same job and probably better for a much lesser cost.

Answer 8

thin disposable blades

Question 9

Tissue sections that are embedded in plastic material such as methacrylate, araldite, or epon are sectioned with a….

Answer 9

glass or a diamond knife

Question 10

Tissue sections that are embedded in plastic material such as (3) are sectioned with a glass or a diamond knife.

Answer 10

methacrylate, araldite, or epon

Question 11

A glass knife can section down to about____

Answer 11

1 micron

Question 12

For electron microscopy thin sections are preferred and they are sectioned to about _______utilizing a diamond knife.

Answer 12

1/4 of a micron

Question 13

Sectioning tissues is a real art and takes much skill and practice. Histotechnologists are the artists of the laboratory.

It is important to have a properly____ and ____ block or much artifact can be introduced in the sectioning.

Answer 13

fixed and embedded block

Question 14

Common artifacts include…

Answer 14

tearing, ripping, holes, folding, etc.

Question 15

Once sections are cut, they are floated on a_____ that helps remove wrinkles.

Then they are picked up on a____

Answer 15

warm water bath

glass microscopic slide

Question 16

As a general principle,_____ of the section at the cut line is never advantageous in histology or material sectioning.

Answer 16

excessive bending

Question 17

Errors of too shallow and too steep an angle are damaging to the specimen. This leads inevitably to the conclusion that the _____should always be set

Answer 17

correct knife angle

Question 18

Position the lower bevel face____ to the specimen block and the plane of motion.

Then raise the angle slightly above the bevel angle to avoid having the lower bevel face slide over the specimen block and possibly produce friction damage.

The correct knife angle positioning should be consistent regardless of the type of microtome, or any specimen property.

Answer 18

parallel

Question 19

The ______prevents contact between the knife facet and the face of the block.

Answer 19

clearance angle

Question 20

The________ is the angle between the two facets that form the cutting edge.

Answer 20

facet or bevel angle

Question 21

For routine use knives and disposable blades are made with a facet angle of approximately____, but this angle can vary with the blade type and from manufacturer to manufacturer.

Therefore for each blade type the clearance angle must be optimally set.

Answer 21

35°

Question 22

For rotary microtome knives and blade holders a clearance angle between_______ is recommended.

Answer 22

0° and 15°

kay ma’am - 5°

Question 23

Problems in sectioning

Answer 23

A. If the ribbon is crooked rather than straight

B. If a ribbon will not form and each section comes off separately

C. If the sections are compressed or folded

D. If the specimen crumbles or falls out of the paraffin section

E. If the ribbon continuously splits or scratches vertically

F. If the ribbon wraps itself around your finger, the knife blade or anything else

Question 24

Remedies

If the ribbon is crooked rather than straight:

Answer 24

• The upper and lower edges of the block face are not parallel to each other or to the knife edge.

• The knife edge may be uneven. Try another section of the knife edge.

Question 25

Remedies

If a ribbon will not form and each section comes off separately.

• The blade edge is dull.

• The knife has the wrong tilt.

• The upper and lower edges of the paraffin block face are crumbled or rounded.

Answer 25

Sharpen it.

Adjust it.

Retrim with a sharp razor blade.

Question 26

Remedies

If the sections are compressed or folded:

• The knife is dull.

• The knife angle is set too close to the vertical.

• The knife is clogged up with paraffin.

• The sections are too thin.

• The paraffin is too soft because the room is warm.

• The paraffin is still contaminated with xylene.

Answer 26

Sharpen it.

Increase the angle.

Clean the edge with chloroform.

Increase the thickness setting.

Pack the blade with ice cubes. Sometimes holding an ice cube against the face of the block helps.

Re-embed.

Question 27

Remedies

If the specimen crumbles or falls out of the paraffin section :

• The tissue is inadequately dehydrated or cleared.

• The tissue is too hard and compact (e.g., liver).

• The tissue was in the paraffin oven too long at too high a heat.

• The tissue is too hard for a paraffin matrix (e.g., bone, cuticle, heavy plant fibers, or xylem).

Answer 27

Dissolve off the paraffin, rehydrate, and clear.

Soak it in water or 70 % ethanol to soften it.

Throw it away.

Try celloidin embedding.

Question 28

Remedies

If the ribbon continuously splits or scratches vertically:

• Crystals or dirt particles are caught on the knife edge or on the paraffin face.

• The knife edge is nicked.

Answer 28

Wipe the knife edge carefully and clean it with chloroform.

Use another section of the knife or sharpen it.

Question 29

Remedies

If the ribbon wraps itself around your finger, the knife blade or anything else:

• The problem is_____, which is often present in very dry room.

• Let _____stand nearby, section in the morning hours or make short ribbons.

Answer 29

static electricity

boiling water

Question 30

Sectioning
 Mechanism

Answer 30

• slow moving of the block into the path of an extremely sharp knife

Question 31

• allows sharpening; expensive

Answer 31

standard thick metal

Question 32

• same with (or probably better than) std thick metal; cheaper

Answer 32

thin disposable blades

Question 33

• plastic-embedded (methacrylate, araldite or epon)

Answer 33

glass knives or diamond knives

Question 34

diamond knives
• for EM; ultrathin sxns (____)

Answer 34

~0.25 um

Question 35

Sectioning
Knife Angle

General principle: _____of sxn at the cut line is disadvantageous.

Answer 35

excessive bending

Question 36

Sectioning
Knife Angle

Errors happen if knife angle is_____
this damages the specimen

Answer 36

too shallow or too steep

Question 37

• prevents contact between knife facet and face of block

• for rotary microtome -_____

Answer 37

Clearance angle

0° to 15°

Question 38

• angle between the two facets that form the cutting edge

• knives and disposable blades

Answer 38

Bevel angle (facet)

~35° (variable)

Question 39

Knife Angle
• Considerations
- incorrect knife angle have various consequences, depending on:

Answer 39

• a specimen type
• fixation and processing qualities
• knife flexibility