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HT 12 Exam 1

QuestionAnswer
surgery tissue is removed in surgery, in the physician's office or at postmortem
grossing the tissue is described and dissected, looking for abnormalities
fixation stabilization of proteins to prevent autolysis and putrefaction
processing dehydration-->clearing-->infiltration to embed the tissue in a solid medium firm to enable thin sections to be cut
embedding orientation of the tissue in infiltration medium and allowing it to soldify
microtomy the sectioning of tissue for examination by microscopy
cover-slipping a mounting medium
purpose of fixation a. Stabilization of proteins to prevent autolysis and putrefaction b. Maintain proper relationship between cells and extracellular material
Discuss the goal of tissue processing To embed the tissue in a solid medium firm enough to support the tissue and give it sufficient rigidity to enable thin sections to be cut, and yet soft enough no to damage the knife or tissue
Define and discuss dehydration the removal of free water from tissue
Define and discuss clearing to remove the dehydrating fluid from the tissue
Define and discuss infiltration Maintains structure of tissue for cutting
Define and describe embedding Fixing the tissue specimen firmly into the embedding medium, allowing it to harden-->ready to cut
Describe microtomy and the use of a microtome a. The sectioning of tissue for examination by microscopy b. Sections routinely cut at 4-6 microns (um)
Discuss staining as it applies to Histotechnology Tissue is stained to aid in light microscopy
Discuss cover-slipping Protection and for better viewing under the microscope
Pathology study of disease
Histology study of tissues
Cytology study of cells
Histologist a scientist who specializes in the study of tissues
Histotechnology a technical histology concerned especially with preparing and processing histological specimens
Histotechnician/histotechnologist a technician who specializes in histotechnology
Biopsy removal of living tissue for examination under the microscope
Autolysis cell breakdown
Putrefaction rotting degradations from micro organs
Postmortem an examination of a dead body to determine the cause of death
Miscible forming a homogeneous mixture when added together
Microtome an instrument used to cut a specimen, as of organic tissue, into thin sections for microscopic examination
define and describe embedding Orientation of the tissue in infiltration medium and allowing it to solidify. Also known as casting or blocking. Areas of interest should be embedded down.
Describe the proper specimen orientation when embedding SKIN and other structures with a wall (layers) Wall tissues must be embedded on edge (gallbladder, intestine, skin)
describe the proper specimen orientation when embedding tubular structures tubular structures (with a LUMEN) are embedded in cross section (on end)
Describe proper specimen orientation when embedding bone. Embedding at an angle. It allows knife edge to cut into a small area first and gradually cut through the remainder of the tissue (diagonal).
Describe double embedding Tissues embedded in a medium and then re-embedded in paraffin. Used to orientate difficult specimens.
Demonstrate the use of quality control in embedding Recording number of pieces and blocks submitted, plus any special instructions (stains, embedding instructions). Only embed one cassette at a time, ensure forceps are clean between embedding different tissues.
Define longitudinal section cut that is parallel to the longest dimension
Define transverse section a cut that is perpendicular to the longest dimension
Define oblique section any cut made between the longitudinal plane and the transverse plane
Define tangential section a cut that only touches or grazes the surface
Define sagittal section a cut that divides into left and right
Define lumen the space within an artery, vein, intestine or tube.
Describe proper embedding techniques of proper orientation, prevention of contamination and optimal tissue support. Clean the forceps between embedding of different tissues to prevent cross contamination. Areas of interest should be tampered so that the tissues lays flat down ensuring the entire specimen is cut. Cooling the blocks as rapidly as possible.
Define use of tampers, base molds and cassettes. Tampers are used to flatten tissue in the mold. Base molds form the shape of the block to be sectioned (reusable, disposable, and peel away). Cassettes hold and identify the specimen to be embedded.
Discuss the marking of specimens during grossing Area of interest can be marked with a notch or ink. Some facilities use "ink side up" while others use "ink side down." India ink, Davidson Marking, tissue dyes, silver nitrate, and tattoo ink.
Discuss optimal temperatures for the embedding center components. 2-4 C above melting point for the paraffin.
rotary microtome embedded tissue passes over the knife edge cutting the section. each turn advances the number of micrometers. used in cryostats. (most common)
sliding microtome block is held stationary and the knife travels over the specimen. block advances up to the knife at a preset distance. usually used for larger blocks in research
freezing microtome knife moves over the specimen on a stage, advancing upward after each cut. liquid CO2 supplied to the tissue for freezing. sections floated in distilled water
sledge microtome sample is placed into a fixed holder, which then moves backwards and forwards across a knife cuts sections of plastic, wood, and other hard materials cuts sections at 1-60 um biconcave knives
ultra microtome ultra thin sections of specimens are cut. uses glass/diamond knives. 0.5-2 um
rocking microtome 6-20 um. performs repetitive rocking motions while advancing towards a fixed knife. biconcave knives.
vibrating microtome accurately cut tissue without freezing or embedding. cutting using a vibrating blade. used for difficult biological samples. 30-50 um for live tissue. 10-500 um for fixed tissue. maintains cell morphology.
steel knives rust resistant, free from impurities and contain anti-corrosives. heat treated to harden the edge
disposable blades most common. superior edges. refined, thickened razor blades. high and low profile.
tungsten carbide nonmagnetic and 100x harder than hardened tool steel. excellent resistance to wear but are brittle because of extreme hardness.
glass knives used for ultramicrotomy. hard but brittle.
diamond knives expensive but extremely durable due to hardness. used for ultramicrotomy
bioconcave concave on both sides. used on rocking or sledge type microtomes. rarely used in U.S.
planoconcave concave on one side only. used for celloidin or plastic resin sectioning.
plano-wedge (wedge) used for paraffin sectioning on a rotary microtome, for cryostat sectioning. easiest knives to sharpen
chatter parallel tears or fractures in tissue
serial section all consecutive sections from a block are saved
levels (steps/step sections) block is trimmed, sections are cut and picked up, the more of the block is trimmed away, and more sections are cut and picked up
coarse trimming using a blade to manually trim away excess material
washboarding aka: udulations- parallel blinds of varied thickness
clearance angle the angle between a plane containing the end surface of a cutting tool and a plane passing through the cutting edge in the direction of cutting motion
microtome saftey always lock the handwheel, use of blade guard, do not rock the handwheel
how to prevent repetitive motion injury (RMI) taking breaks using an automated rotary microtome
identify problem and the appropriate corrective action: compression cause: knife too dull or dirty remedy: sharpen/replace blade cause: too little knife tilt remedy: increase knife angle cause: block too warm remedy: chill block
identify problem and the appropriate corrective action: split ribbons cause: dull knife remedy: sharpen knife/change blade cause: knife angle too little remedy: adjust knife tilt cause: warm room/paraffin too soft remedy: chill block
identify problem and the appropriate corrective action: length wise scratches cause: defect in the knife edge or hard particle in the tissue/dirty knife remedy: move or change blade
identify problem and the appropriate corrective action: gouging (overlapping) cause: knife too dull or dirty remedy: sharpen knife or replace cause: too little knife tilt remedy: increase knife angle
identify problem and the appropriate corrective action: crooked ribbons cause: horizontal edges of block not parallel remedy: trim block until parallel, reposition specimen holder cause: blade blunt in one area remedy: move blade or replace cause: tissue varies in consistency remedy: rotate block
identify problem and the appropriate corrective action: chatter cause: over dehydration remedy: rehydrate cause: too much knife tilt remedy: adjust knife angle cause: dull knife remedy: sharpen knife or replace blade
identify problem and the appropriate corrective action: lifting sections cause: dull knife remedy: sharpen or change blade
identify problem and the appropriate corrective action: udulations/washboarding cause: knife angle too great remedy: decrease knife tilt cause: loose block or knife remedy: tighten block or knife cause: hard tissue remedy: use very sharp knife
identify problem and the appropriate corrective action: holes in sections cause: block faced too aggressively remedy: face block less aggressively cause: excessive dehydration remedy: rehydrate
identify problem and the appropriate corrective action: lack of ribbon forming cause: dull knife remedy: replace knife cause: paraffin too hard remedy: warm paraffin, re-embed cause: too much knife tilt remedy: adjust knife angle cause: room temp too low remedy: adjust room temp, warm knife/block
identify problem and the appropriate corrective action: thick and thin sections cause: block or blade loose remedy: tighten cause: too little knife tilt remedy: increase clearance angle cause: wax too soft remedy: cool block or change to higher melting point wax
describe the effects of speed on microtomy whether fast or slow a constant speed is necessary to avoid uneven sections one revolution of the hand-wheel per second smooth even revolutions
describe "tricks" for obtaining optimal microtomy sections breathe on the block to provide heat, moisture, reduce static. softening paraffin embedded tissue by trimming the block until the tissue is exposed and then soaking the block in H2O or glycerol. tissues containing keratin can be soften by a depilatory
describe routine mircrotome maintenance keep covered when not in use. thorough cleaning after use. lubrication as recommended. routine preventative maintenance.
Optimal temperature for the flotation bath? 5-10 C below melting point of paraffin (which is 58 C)
Describe the need for distilled or deionized water deoionized water prevents static and contamination
Describe the use of additives and adhesives in the flotation bath Alcohol, detergent, photo-flo can be added to the flotation bath to lower the surface tension.
Identify the need for drying slides Water is not miscible with the solvent used for deparaffinizing slides for staining
Created by: deeloused