MICROTOMY part 2 Flashcards
TYPES OF TISSUE SECTIONS
Paraffin
Celloidin
Frozen sections
Paraffin section size
4-6 um
Successive sections will usually stick edge-to-edge.
Paraffin section
Sections are removed in ribbons of ten to allow easy location of serial sections.
Most common
Paraffin sections
Celloidin section size
10-15um
The blocks are trimmed in the same manner as in paraffin blocks.
To avoid dehydration and shrinkage, sections are usually cut by the wet method, with both the sections and the block being kept moist with _____during cutting.
Celloidin
70% alcohol
_____sections do not come off in ribbons and have to be collected into 70% alcohol immediately.
Celloidin
Frozen Sections
Methods of preparing frozen section:
- Cold knife procedure
- Cryostat procedure (cold microtome)
three basic types or shapes of microtome knives:
Plane-concave knife
Bioconcave knife
Plane-wedge knife
Plane-Concave Knife (usually_____. in length)
25 mm
Biconcave Knife (usually____. in length)
120 mm
Plane-Wedge Knife (usually____. in length)
100 mm
One side of the knife is flat while the other is concave.
Plane concave knife
Less concave sides are recommended for cutting_______-embedded tissue blocks on a______ microtome.
celloidin
sliding
More concave sides are used to cut_____ sections on base-sledge, rotary or rocking microtome
paraffin
with both sides concave
Biconcave
Biconcave knife
recommended for cutting_____ - embedded sections on a_____ microtome.
paraffin
rotary
have both sides straight
Plane-wedge knife
Plane-wedge knife
recommended for______ sections or for cutting extremely hard and tough specimens embedded in_____ blocks, using a_____ or _____
frozen
paraffin
base sledge type or sliding microtome.
recommended for frozen sections or for cutting extremely hard and tough specimens embedded in paraffin blocks, using a base sledge type or sliding microtome.
Plane-wedge knife
_____ and _____ knives are usually provided with backs, to maintain the correct bevel angle throughout honing.
Detachable handles may be attached to the knife during sharpening.
Plane-wedge and plane-concave
The angle formed between the cutting edges is known as the “______”, normally about________
Bevel angle
27° to 32°
Too______ cutting edges are likely to become dull easily, while too____ edges are likely to produce nicks or jagged edges and irregularities on the knife edge, thereby producing tears or striation on the tissue sections during cutting.
soft
hard
A good cutting edge must be able to cut good sections from a paraffin wax block about_______ thick, without any serration noted on examination.
2-3 microns
________may be used for partially calcified materials, paraffin and frozen sections.
They are readily replaced when dull, and produce similarly good tissue sections as those cut with microtome knives
Safety razor blades
unsatisfactory for sections less than 10 µ
Safety razor blades
Theoretically, the perfect and optimum cutting angle is obtained when the sides of the wedge knife are inclined at an angle of about_____, causing maximum penetration of the tissues and minimizing distortion
15°
To prevent uneven sections, or alternate thin and thick sections, the knife should be inclined with a ______ clearance angle from the cutting plane so that the cutting facet will not compress the block during the process of cutting
5-10°
True or False
The cutting edge must be thinner than the section being cut.
True
A good cutting edge must be sharp enough to cut good sections from a paraffin wax block at____ thick without causing serrations.
4 µ
OTHER KNIVES AND BLADES
Disposable blades
Glass knives/ Ralph knives
Diamond knives
Safety razor blades
• Widely used now because cheaper;
• Honing and stropping are no longer common practices;
Disposable Blades
Disposable Blades
Coated with_______ (for ease of ribboning).
polytetrafluoroethylene
• For ultrathin microtomes.
Glass Knives/ Ralph Knives
• For resin blocks on ultrathin microtomes;
• Brittle and expensive
Diamond knives
For partially calcified materials, paraffin, and frozen sections.
Safety razor blades
Safety Razor Blades
Unsatisfactory for sections less than____
10 micro.
Badly Nicked knives with blunted ends have to undergo_____ in order to ensure optimum sectioning of tissue blocks.
sharpening
Sharpening of the knives involves 2 stages, namely:
Honing
Stropping
HONING
Removing of gross nicks
To acquire an even edge
HEEL to TOE
STROPPING
Removal of burr/irregularities
Final polishing of the knife edge
TOE to HEEL
Hard sharpening
Honing
Honing involves the removal of gross nicks on the knife edge (_______) to remove blemishes
grinding the cutting edge of the knife on a stone (_______) to acquire an even edge
Coarse Honing
Honing Proper
The degree of sharpness is proportional to the fineness of the abrasive used in sharpening
Honing
This procedure makes use of a______, a natural sharpening stone or hard grinding surface (carborundum), which serves to remove nicks and irregularities on the knife edges.
hone
Several types of hones may be used:
Belgium yellow
Arkansas
Fine carborundum
- for manual sharpening when cutting edge has been rendered blunt or nicked.
This type usually gives the best result.
MOST COMMON
Belgium Yellow
- gives more polishing effect than the Belgium Yellow.
Arkansas
- is much coarser than the first two types and is used only for badly nicked knives followed by either one of the first two knife sharpeners.
EXCELLENT
Fine carborundum
The surface of the hone is wiped clean with a soft cloth moistened with_____ in order to remove the scattered small particles of stones and metal
xylene
Honing lubricants
• Soapy water
• Mineral oil
• Clove oil
• Xylene
• Liquid paraffin
Honing
The knife is fitted to its corresponding back, placed on one end of the hone, and with the cutting knife edge first,
the “____” (handle end) is drawn obliquely or diagonally towards the operator on the stone until the “____” (head portion) is reached.
heel
toe
The knife is then turned over, and the other surface is again drawn forward, EDGE FIRST, with a HEEL TO TOE direction.
Honing
In the case of the Minot or plane-wedge knife, the knife is turned over so as to sharpen the other surface every_____ strokes
I0-20
Honing
For plane-concave knives, only the_____ surface should be rubbed on the Hone.
concave
Honing
______may also be used for final polishing
Diamantine
Mechanical honing with machines may make use of a vibrating frosted glass plate or a wheel driven by an electrical motor.
The knife is pressed against the flat side of a rotating glass wheel which is being driven by a mechanical device.
Approximately____\ double strokes are given each side of the knife to which very gentle pressure is applied.
The use of knife sharpening machines, although quite expensive, is usually time-saving and produce well sharpened knives with uniform bevels
30
The hone should be long enough (about______) to allow the whole length of the knife edge to be sharpened in a single stroke and wide enough to sufficiently support and prevent the rocking of the knife.
8” x 3”
is the process whereby the “burr” formed during honing is removed and the cutting edge of the knife is polished.
Stropping
The purpose of______ is to polish and sharpen the cutting edge, while that of______ is to remove the irregularities from the knife.
stropping
honing
If the knife has become dull and blunt, but is free from nicks or teeth, it is usually only necessary to____ it.
For delicate work, the knife is____ before every object is sectioned.
strop; stropped
A paddle strop made up of the best quality_______, firmly attached to a solid back, in order to prevent sagging is preferred
horse leather
e procedure is the reverse of honing.
Stropping
The knife is first fitted with its appropriate knife back, then laid obliquely on the strop and with the cutting edge behind, (EDGE LAST) is pushed backward and drawn forward in a TOE TO HEEL direction
Stropping
In stropping
Around______ double strokes are usually required.
40-120
Stropping
…applied on the back of the horse leather;
Not _____because it tends to blister and the leather destroys.
Vegetable oil (e.g. castor oil)
mineral oil
The strop should not be used for at least ______ after treatment.
Too much oil will make the stropping surface slippery and will render the procedure unsatisfactory.
To remove excessive oil from the strop, its surface is scraped with a_____
24-48 hours
blunt instrument, e.g. the back of the knife.
Wax must not be allowed to come in contact with the strop. With an applicator, the used knife blade should be washed and flushed with_____
xylene
Disposable blades
They have a sharp cutting edge that can cut_____ thick sections with ease.
2-4 µ
are also now available that can attach to some blade holders and are particularly suitable for use in the cryostat.
Magnetic knives
are generally used for trimming and semi-thin sectioning of tissue blocks for electron microscopy.
Glass knives
are used to cut any type of resin block for electron microscopy.
When supplied by manufacturers, they are already mounted in a metal block designed to fit directly into the knife holder of the ultrathin microtome
Diamond knives
.
Used for adhesion of the tissue to be examined to the slide.
.
For: Brain sections, decalcified tissue, and when using strong alkali at time of staining.
ADHESIVES
ADHESIVES examples
• Mayer’s Egg Albumin
• Dried albumin
• Starch paste
• Plasma
• 1% Gelatin
• 1% Methyl Cellulose
• Poly-L Lysine
• Sodium silicate
• Resins
