Sectioning Flashcards
Microtome model that we use in the lab
Leica RM2125 RTS Microtome
Measurement of sections produce by the Leica RM2125 RTS Microtome
0 - 60um
2 dot meaning
Initial Sectioning
Initial sectioning measurement
25um
1 dot meaning
Final Sectioning
Final sectioning measurement
10um
Floatation bath temp
5-10C below melting point of paraffin wax
Most commonly used adhesive
Meyer’s Egg Albumin
Ratio concentration for Meyer’s Egg Albumin
1:1
Meyer’s Egg Albumin composition
1mL Egg white & 1mL Glycerin
Added to Meyer’s Egg Albumin to prevent molds
Phenol
Adding Phenol to Meyer’s Egg Albumin will allow the slide to last for how long?
3 - 4 months
Other names for Sectioning
Microtomy
Cutting
The process where tissues are cut into uniformly thin slices to facilitate microscopic studies
Sectioning
Most paraffin embedded tissues are sectioned to what measurement
4 - 8um
Histotechnologists are the _______ of the laboratory
artists
Common artifacts introduced in sectioning
Tearing
Ripping
Holes
Folding
4 types of microtome knives
Standard Thick Metal
Thin Disposable Blades
Glass Knives
Diamond Knives
This knife allows for sharpening to your liking and is considered to be expensive
Standard thick metal
Does the same job as the standard thick metal knife, and probably better, for a much lesser cost
Thin disposable blades
Glass knives are for tissue sections embedded in what types of plastic materials
Methacrylate
Araldite
Epon
A glass knife can section down to what measurement
About 1um
Knife used for electron microscopy
Diamond knives
Diamond knife can section to what measurement
About 0.25um
Angle that prevents the contact between the knife facet and the face of the block
Clearance angle
Recommended clearance angle
0 - 15 degrees
Bevel angle is aka?
Facet angle
The angle between the two facets that form the cutting edge
Bevel Angle
Facet angle
approx. 35 degrees
Consequences of incorrect knife angle will vary on the differences in what factors?
Specimen types
Fixation
Processing Qualities
Knife Flexibility
Causes if the ribbon is crooked rather than straight
- edges of the block face are not parallel to each other
- uneven knife edge
Causes if the ribbon will not form and each section comes off separately
- Dull blade edge
- Knife has wrong tilt
- Edges of paraffin block face are crumbled or rounded
Causes if sections are compressed or folded
- dull knife
- knife angle is set too close to the vertical
- knife is clogged with paraffin
- sections are too thin
- paraffin is too soft due to warm room
- paraffin is contaminated with xylene
Causes if the specimen crumbles or falls out of the paraffin section
- tissue is inadequately dehydrated or cleared
- tissue is too hard and compact
- tissue was in the paraffin oven too long and at too high temp
- tissue is too hard for paraffin matrix
Causes if the ribbon continuously splits or scratches vertically
- crystals or dirt particles are caught on the knife edge or on paraffin face
- nicked knife edge
Causes if the ribbon wraps itself around your finger, the knife blade, or anything else
- static electricity
In prepping the tissue block for sectioning, trimming the lateral surface should result in resembling what shape
Trapezoid
After resulting in a trapezoid resembling shape, the tissue block must still be surrounded by a frame of how much wax
2-3mm
Wipe the microtome blade using what solution?
Chloroform or Xylene
In the blade preparation, the angle of the center line of the blade with the face of the block should be about how many degrees
20 degrees
During sectioning, students should start at how many microns
10um
In sectioning, how much paraffin should be allowed to move up and down until the ribbon begins to bow up
first inch or two
How long must the ribbon be before detaching it form the edge of blade
8 inches
How must we detach the ribbon from the edge of the blade
In an upward stroke
How should we brush the blade
away from the cutting of the blade
What should the orientation of the ribbon be when laying it in the ribbon box
Dull surface up
When mounting sections, slide should be rinsed in what
95% ethanol
What should be used to write information on the slide
Diamond-tipped pencil
Lead pencil
Paraffin expands to how many percent
20%
What is the measurement of the cover slip used after staining
22mm x 22mm
At what temperature should the slide warmer be
40 - 45C
