Histo Lec Module 2 Unit 5 Flashcards
process where alcohol or dehydrating agent is removed and replaced by a substance that will dissolve the wax
clearing or dealcoholization
process where alcohol or dehydrating agent is removed and replaced by a substance that will dissolve the wax
clearing or dealcoholization
clearing agent imparts optical property due to its
high refractive index
volume of clearing agent
40 times the tissue volume
clearing time for a small biopsy
1 hr
clearing time for larger tissues
3 changes with 60 minutes each
end point of clearing
transparent tissue
prolonged exposure to clearing ageng makes tissue
brittle, more difficult to cut
wax that is more readily replaced by paraffin has ___ boiling point
lower
clearing agent that causes tissue shrinkage and hardening but is affordable and acts rapidly
xylene
better at preserving tissue structure and does not garden tissue, but is slower acting and can cause tissue shrinkage
toluene
xylene can clear within
15 -30 mins
acts rapidly, does not harden or shrink tissue but is carcinogenic and highly flammable
benzene
used for tough tissues like skin, fibroid and decalcified tissues because they can process thick tissues and large spcimens. it is however slower and not transparent
chloroform
used for processing CNS tissues and cytological studies of smooth muscle and skin because it causes mininal shrinkage, can be used at indefinite time without causing damage. it does not produce uniform quality however, and is hard to eliminate from tissues
cedarwood oil
not recommended for routin clearing but used for embryos, insects and delicate specimens. it can clear 70% alcohok without shrinkage and hardening
aniline oil
most used xylene replacement
lemonene
used for celloidin sections, and has slow, gentle, and non hardening action
terpenes
has lowest hazard rating among all clearing agents and can be used in place of xylene. but it can breakdown compounds if not pure or stable
orange-oil based
toxic clearing agent that is now restricted by the governement
chlorinated hydrocarbon
it is efficient, nonhazardous, less expensive, causes less shrinkage, does not lose quality or detail, but solidifies at lower temperature
coconut oil
it produces good tissue, is nontoxic, non flammable, biodegradable, economic, easy to handle and readily available
bleached palm oil
causes minimum shrinkage but it dissolves celloidin, produces brittle tissue which tend to be adulterated
clove oil
similar to chloroform, dangerous to inhale on prolonged exposure but cheapaer
carbon tetrachloride
superior to ordinary clearing agents because it perofroms 2 processes at a time which shortens total processing time, and is non toxic
tetrahydrofuran
used when time is important because tossues may be embedded 4 hours after fixation due to its miscibility in both water and paraffin. it causes greater shrinkage hiwever and have toxic fumes
dioxane
process by which clearing agent is removed and replaced by a medium that completely fills tissue cavities and give firm consistency to tissues
infiltration or impregnation
clearing agent that takes longer time to infiltrate
cedarwood oil
accelerates the infiltration process
vacuum
simplest, most common embedding medium
paraffin wax
most common melting point of paraffin wax for histological use
56 - 58 degrees Celsius
recommended use of paraffin wax
2 celsius above melting point
pressure applied within a vacuum embedding iven
760 mmHg
vacuum embedding redices embedding time by
25-75%
mixture of highly purified paraffin and synthetic plastic polymers
paraplast
melting point of paraplast
56-57 C
permits large dense tissue blocks such as bines and brain to be cut easily
paraplast
less brittle and less compressible paraffin substitute
embeddol
lower melting point but harder than paraffin
ester wax
melting point of ester wax
46-48 degrees
soluble in 95% alcohol meaning that it can be used without clearing
ester wax
most used water soluble wax
carbowax
suitable for enxyme histochemical studies
carbowax
proprietart blends of plastic plymer paraffin waxes
dimethyl sulphoxide
scavenges residual transition solvent and alters tissue permeability by substituting or removing bound water
dimethyl sulphoxide
suitable for specimens with large hollow cavities, hard dense tissues, laegre tissues and soft tissues with mixed consistency
celloidin
celloidin is supplied as
thin (2%)
medium (4%)
thick (8%)
solutions dissolved in equal parts ether and alcohol
three types of celloidin infiltration
wet mount, dry mount, low viscosity nitrocellulose
difference between wet and dry mount celloidin
wet - 70% alcohol for bines teeth and large brain sections
dry- Gilsons mixture of chloroform and cedarwood oil for whole eyes
some advantages of cellodin impregnation
rubbery consistency, does not require heat, less shrinkage and distortion
some disadvantages of celloidin
slow infilitration
difficult to cut thin sections
cannot make serial sections
difficult to get photomicrographs
rarely infiltration medium used except when dehydration is avoided
gelatin
used for frozen sections
gelatin
thickness of tissues in gelatin infiltration
2-3 mm
prevents growth of mold in gelatin
1% phenol
other terms for embedding
casting blocking
impregnated tissue is positioned in a nold that is allowed to solidify
embedding
if the embedding medium is too soft
torn or shredded sections
embedding medium too hard
brittle tissues that will shatter
arrangement of tissue in a precise position during embedding, microtomy, and before staining
orientation
4 types of mold
Leukharts embedding mold
Compound embedding unit
Plastic embedding ring
disposable embedding molds
two L shaped strios of heavy brass or metal
Leukharts embedding mold
can be adjusted to the size of the specimen
leukhart
series of interlocking plates resting on flat metal base
compound embedding unit
special stainless steel base mold and fitted with block holder
plastic embedding rings and base mold
equipped with warm plate, cold plate, and white plastic cassette molds with snap on lid
tissue tek
three types of disposable embedding molds
peel away
plastic ice trays
paper biats
tissues are first infiltrated with supporting medium such as agar or nitrocellulose then infiltrated a second time with paraffin
double embedding
obsolete due to difficulty of tissue samples to adhere to slides and limiting of tissue expansion
agar as preembedding medium
three types of plastic embedding medium
epoxy
polyester
acrylic
hydrophobic, may reduce antigenicity, and carcinogenic
epoxy
large molecules so slowly infiltrates
bisphenol A Araldite
mixture of isomers that has lower viscosity
glycerol (epon)
purena di filtrae the fastest
cyclohexene dioxide (spurr)
made for EM but now seldom used
polyester
extremely hydrophilic, tough when dehydrated, and used for TEM and preferable to use when high resolution light microscopy is needed
glycol methacrylate (GMA)
where the tissue is held into position
block holdet
for actual cutting of the sections
knife carrier and knife
to line up tissue block in proper position with the knifex adjusting proper thickness for successive sections
pawl, ratchet feed wheel and adjustment screws
hard sharpening
honing
removal of nicks on the knife edge to remove blemishes
coarse honing
grinding the cutting edge of the knife on a stone tk acquire even edge
honing proper
for manual sharpening when cutting edge has been rendered blunt or nicked and usually gives best results
belgium yellow
gives more polishing effect then belgium yellow
arkansas
much coarser than the first 2 types and used only for badly nicked knives followed by either of the two
fine carborundum
direction of honing
heel to toe
to poliish and sharpen the cutting edge
stroping
used in stroping
horse leather
