INFILTRATION Flashcards
saturation of tissue cavities and cells by a supporting substance which is generally, but not always, the medium in which they are finally embedded.
Infiltration
Tissues are infiltrated by immersion in a substance such as_____.
Alternatively, tissues can be infiltrated with a solution of a substance dissolved in a solvent, for example_____ in ______, which solidifies on evaporation of the solvent to provide a firm mass suitable for sectioning.
wax
nitrocellulose in alcohol-ether
Ideally an infiltrating medium should be:
• soluble in processing fluids
• suitable for sectioning and ribboning
• molten between 30°C and 60°C
• translucent or transparent; colorless
• stable
• homogeneous
• capable of flattening after ribboning
• non-toxic
• odorless
• easy to handle
• inexpensive
In addition the properties of the infiltrating medium should approach those of the tissues to be sectioned with regard to (5) and should be harmless to the embedded material.
density, elasticity, plasticity, viscosity and adhesion
Infiltration may be done (3) method
manually
processed automatically
vacuum method
The main advantage of manual processing over automated methods lies in the flexibility of_______, conditions and schedule design to provide optimum processing for small batches of tissues.
reagent selection
The most common infiltration method is_____ infiltration.
Other methods include _____ and _____.
paraffin wax
celloidin infiltration and gelatin infiltration
is a polycrystalline mixture of solid hydrocarbons produced during the refining of coal and mineral oils
Paraffin wax
Wax hardness (viscosity) depends upon the _______of the components and the______
molecular weight
ambient temperature
Paraffin wax
_________ melt at higher temperatures than waxes comprised of lower molecular weight fractions.
High molecular weight mixtures
Paraffin wax
The wax is a solid at room temperature but melts at temperatures up to about_______
65°C or 70°C.
Paraffin wax can be purchased with melting points at different temperatures, the most common for histological use being about______, At its melting point it tends to be slightly viscous, but this decreases as the temperature is increased.
56°C–58°C
The traditional advice with paraffin wax is to use this about___ above its melting point.
2°C
To decrease viscosity and improve infiltration of the tissue, technologists often increase the temperature to above_______ in practice to decrease viscosity.
60°C or 65°C
improve ribboning:
prolong heating of paraffin wax at high temperatures or
use microcrystalline wax
Modiefied paraffin wax
• increase hardness:
Add stearic acid
Modified paraffin wax
decrease melting point:
add spermaceti or phenanthrene
Modified paraffin wax
• improve adhesion between specimen and wax (alter crystalline morphology):
ADD either of the ff:
0.5% ceresin
0.1-5% beeswax
rubber
asphalt
bayberry wax
phenanthrene
Early histological wax formulations have largely been replaced by uniform, high quality proprietary blends of histological paraffin waxes.
Additives recently incorporated in these waxes include the following:
- Piccolyte 115
- Plastic polymers
- Dimethyl sulphoxide (DMSO)
, a thermoplastic resin added at the rate of 5%-10% to the infiltrating wax, or to the final investing paraffin wax to improve tissue support for thin sectioning and facilitate flattening and expansion of sections on the waterbath.
Piccolyte 115
% of Piccolyte 115 added to paraffin wax
5%-10%
, added to improve adhesion, hardness and plasticity. Polymer waxes are incorporated in the majority of paraffin wax blends presently available.
Plastic polymers
Plastic polymer example
polyethylene wax
are incorporated in the majority of paraffin wax blends presently available.
Polymer waxes
added to proprietary blends of plastic polymer paraffin waxes that reduces infiltration times and facilitates thin sectioning.
Dimethyl sulphoxide (DMSO)
scavenges residual transition solvent and probably alters tissue permeability by substituting for or removing bound water thus improving infiltration.
Dimethyl sulphoxide (DMSO)
Some individuals who handle _______may experience an unpleasant and annoying oyster or garlic taste probably caused by _____metabolites.
DMSO-paraffin wax
DMSO
SUBSTITUTES FOR PARAFFIN WAX
Paraplast
Polyethylene glycols (PEG)
Carbowaxes
Ester waxes
A modern paraffin embedding medium for tissue specimens is
Paraplast®
highly purified paraffin with a melting point of 56ºC and formulated to give outstanding tissue infiltration and superior quality sections.
Paraplast®
Melting point of highly purified paraffin with a melting point of Paraplast
56C
Unlike some other tissue embedding products,______ is already doubly filtered, and there is no need for further filtration prior to use.
Paraplast
are water soluble media used for investigation of heat and solvent-labile lipids and proteins
Polyethylene glycols (PEG)
Used to overcome tissue shrinkage, damage and distortion inherent in the paraffin wax technique.
Polyethylene glycols (PEG)
The polyethylene glycols, or_____, are polymers of varying length.
In general they are less elastic, denser and somewhat harder than paraffin wax.
Carbowaxes
have low melting points, are hard at room temperature and have good adhesive properties.
Ester waxes
They are ideal for supporting and serially sectioning refractory hard, criticized material such as arthropods, and tissues which heat-harden excessively
Ester wax
used for simple investment of paraffin blocks to be sectioned under hot conditions and in double embedding with agar.
Ester wax
are no longer commercially available and must be prepared from the basic ingredients.
Ester waxes
Celloidin and Low Viscosity Nitrocellulose, mixtures of di- and tri-nitrocellulose, are composed of yellowish-white matted filaments with the appearance of_____.
raw cotton
is insoluble in water, but soluble in absolute ethanoldiethyl ether, amyl acetate, methyl benzoate, methyl salicylate and 2ethoxyethanol and is set by most hydrocarbon solvents.
Nitrocellulose
Nitrocellulose is insoluble in water, but soluble in (5)and is set by most hydrocarbon solvents.
absolute ethanoldiethyl ether
amyl acetate
methyl benzoate
methyl salicylate
2ethoxyethanol
T or F
Nitrocellulose is highly flammable,
True
Nitrocellulose must be kept_________ or as_________
alcohol-damped with n-butanol
8% solutions in ethanol-ether
solutions have a low tolerance of water and dehydration must be thorough
Celloidin
tolerates up to 6% of water, has superior penetration and final block hardness and is supplied in various grades of viscosity and nitrogen content.
low viscosity nitrocellulose (LVN)
________tissue processing techniques are generally employed for sectioning hard tissues such as bone, for topographical studies of central nervous system tissues, or for delicate embryonic material.
Tissues are processed at room temperature producing minimal and shrinkage and hardening.
Nitrocellulose
Infiltration sequence for specimens not more than 4mm thick.
1st paraffin wax bath - 30 mins
2nd paraffin wax bath - 30 mins
3rd paraffin wax bath - 45 mins
Process whereby the clearing agent is completely removed from the tissue and replaced by a medium that will completely fill all the tissue cavities and give a firm consistency to the specimen.
Infiltration
