Microtomy

49問 • 2年前

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It is an instrument capable of cutting section at a predetermined thickness by sliding block into a cutting tool which is fixed and attached to the machine

Microtome

What are the three essential parts of a microtome?

-Block holder -Knife carrier and knife -Pawl, Ratchet feed wheel and adjustment screws

It is where the tissue is held

Block holder

It is responsible for the actual cutting of the tissues

Knife carrier and knife

It line up the tissue block in proper position with knife, adjusting the proper thickness of the tissue for successive sections

Pawl, Ratchet feed wheel and adjustment screws

What are the diff kinds of microtomes?

-Rocking microtome -Rotatory microtome -Sliding microtome -Freezing microtome -Cryostat microtome -Ultrathin microtome

For cutting serial sections of large blocks of paraffin embedded tissues

Rocking microtome

For cutting paraffin embedded sections.

Rotary microtome

For cutting celloidin embedded sections.

Sliding microtome

For cutting unembedded frozen sections.

Freezing microtome

For cutting frozen sections

Cryostat or cold microtome

For cutting sections for Electron Microscopy.

Ultrathin microtome

Invented by Paldwell Trefall in 1881

Rocking(cambridge) Microtome

Invented by Minot in 1885-86 to cut paraffin embedded

Rotatory microtome

Developed by Adams in 1789.

Sliding microtome

Invented by Queckett in 1848.

Freezing microtome

- Simplest -Consists of a heavy base and two arms • Lower arm resting on pivots and supporting column, and attached to the micrometer screw • Upper arm, connected to a lever by a piece of nylon thread -A section is cut as the tissue passes to the knife edge in a slightly curved plane, in 10-12 um thickness - Not currently favored by most labs due to restrictions in size of tissue block that can be cut and difficult of reorienting the block

Rocking microtome

- Most common in laboratories -Electrically driven are available and can be ideally used to produce ribbons for serial sections - The knife is fixed in a horizontal position. - Sections are cut between 3 and 5 um using paraffin wax. -Knife and block holder are brought together by upward and vertical motions - Heavier and more stable. - Heavier knife is used so there is less vibration.

Rotatory microtome

There are two types of this microtome: 1)Base-Sledge Microtome, consists of two movable pillars holding the adjustable clamps, allowing the knife to set at an angle for cutting; Favored in laboratories where very hard or large blocks are usually sectioned; Angle of knife is adjustable and knife used is 24 cm long. 2)Standard sliding microtome, knife moves backward and forward; more dangerous due to the movement of knife which makes it difficult to attach knife guards; Required to be slow and steady

Sliding microtome

Consists of two movable pillars holding the adjustable clamps, allowing the knife to set at an angle for cutting; Favored in laboratories where very hard or large blocks are usually sectioned; Angle of knife is adjustable and knife used is 24 cm long.

Base-Sledge Microtome

Knife moves backward and forward; more dangerous due to the movement of knife which makes it difficult to attach knife guards; Required to be slow and steady

Standard sliding microtome

-Stage for block holder is hollow and perforated around its perimeter, attached to a reinforce flexible lead pipe thru which CO2 passes from a cylinder. - Used to cut undehydrated thin to semi-thin sections -Required in rapid diagnosis, histological demo of fat, neurological structures are to be studied, and sensitive tissue studies. - Equipped with a stage upon which tissue can be quickly frozen using either liquid CO2 or a low temperature recirculating coolant - Cutting action, knife is moved whilst the tissue block remains static

Freezing microtome

-Refrigerated apparatus used for freezing the tissue into the block holder to the correct degree of hardness that allows for easier and faster sectioning - Cold chamber maintains at temperature between -5 to -30 C (average is -20 C) by an adjustable thermostat, capable of freezing fresh tissues with 2-3 minutes, and cutting sections of 4 um with ease. - Commonly used for rapid preparation of urgent tissue biopsies for intraoperative diagnosis

Cryostat microtome

It is commonly used for rapid preparation of urgent tissue biopsies for intraoperative diagnosis

Cryostat microtome

-Equipped with a glass or gem grade diamond knife that is used to cut very thin sections (typically about 60 to 100 nm) - Sections are stained with an aqueous solution of an appropriate heavy metal salt and examined with a transmission electron microscope (TEM) - Semi-thin sections are generally 0.5 to 1 um thick and mounted on a glass slide. - Stained to locate areas of interest -Thin sectioning for the TEM is often done with a gem quality diamond knife

Ultrathin microtome

After carefully drying the machine and knife holder, the parts should be wiped with _______. Prolonged and continuous application of the painted parts with xylene should, however, be avoided since this reagent is capable of removing the paint.

Xylol

What are the diff types of Microtome knives?

-Plane- concave knife -Biconcave knife -Plane-wedge knife

-One side of the knife is flat while the other is concave. Less concave slides recommended for cutting celloidin-embedded tissue blocks. For sliding microtome - Base-sledge, rotary or rocking microtome

Plane- concave knife

- Usually 120 mm in length - Rotary microtome

Biconcave knife

-Usually 100 mm in length - Used in base-sledge type or sliding microtome

Plane-wedge knife

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

The perfect and optimum cutting angle is obtained when the sides of the wedge knife are inclined at an angle of about __ degrees.

Bevel angel is normally about _________.

27 to 32

-Involves the removal of gross nicks on the knife edge (Coarse Honing) to remove blemishes, and grinding the cutting edge of the knife on a stone (Honing proper) to acquire an even edge -Makes use of a hone, natural sharpening stone or hard grinding surface (carborundum), which serves to remove nicks and irregularities.

Honing(Hard Sharpening)

For manual sharpening when cutting edge has been rendered blunt or nicked. This type usually gives the best result

Belgium Yellow

What are the several types of hones that may be used in Honing?

-Belgium yellow -Arkansas -Fine caborundum

Gives more polishing effect than Belgium yellow

Arkansas

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.

Fine carborundum

It may also be used for final polishing

Diamantine

It 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 rotating glass wheel which is being driven by a mechanical device.

Mechanical honing

-"burr" formed during honing is removed and the cutting edge of the knife is polished - Purpose, to polish and sharpen the cutting edge, while that of honing is to remove the irregularities from knife.

Stropping

Sharpening (honing) and polishing (stropping) are no longer common practice in most modern laboratories because of the availability of these.

Disposable blades

-Generally used for trimming and semi-thin sectioning of tissue blocks for electron microscopy. They are prepared from commercially available 40 x 2.5 cm plate glass that have been washed with detergent, rinsed in distilled water and alcohol, dried with lint-free paper. - Should be prepare and stored in dust-free boxes with lids to avoid contamination

Glass knives

- Used to cut any type of resin block for electron microscopy. - Brittle, expensive but very durable

Diamond knives

- Thermostatically controlled type is preferable. -Temperature of water should be between 5 and 10 C below paraffin wax melting point

Water bath

-Small drying ovens are now available incorporating a fan, especially designed for drying tissue section on slides. - For delicate tissues, lower drying temp. is recommended to avoid splitting and cracking

Drying oven or hot plate

- Fine pointed or curved and squirrel hair brush -These tools are needed for handling sections during cutting, and for removing folds and creases on the sections during "floating out" in water bath

Forceps

-For routine work, 76 x 25 mm slides that are 1.0 to 1.2 mm thick are usually preferred because they do not break easily.

Clean slides

-Made on the assumption that regular sides have been used. -Larger size of slides are used for sections of eyes or CNS tissues when these will not fit on the regular ones.

Slide rack

Translation(Pre-finals)

MAHATHIR ALPHA · 55問 · 2年前

Translation(Pre-finals)