WAX IMPREGNATION Flashcards
Process of replacing the clearing agent with the infiltrating medium to fill all tissue cavities, providing firm consistency for easy handling and thin section cutting
Impregnation (Infiltration)
Process where impregnated tissue is positioned in a mold with an embedding medium and allowed to solidify
Embedding (Casting)
Most common embedding medium; allows easy cutting of serial sections without distortion; very rapid (prepared within 24 hours); suitable for many staining procedures
Paraffin
Melting point for routine work; should not exceed this temperature to avoid tissue damage
56°C
Overheating paraffin (>60°C) causes these issues
Brittleness, shrinkage, and hardening
Type of tissue for which paraffin embedding is not recommended
Fatty tissues
Manual processing requires four changes of paraffin, with this interval between each change
15 minutes
Two types of automatic tissue processing
Tissue transfer (dip and dunk) and fluid transfer (enclosed type)
Specimens are transferred from container to container for processing
Tissue transfer
Specimens are held in a single process chamber or retort where fluids are pumped in and out
Fluid transfer
Number of stations in an autotechnicon
12 stations
Thermostat setting for wax bath in autotechnicon
3°C higher than the melting point of wax
Histopathologic technique steps performed by autotechnicon
Fix,dehydrate, clear, infiltrate
First two stations in autotechnicon
10% formalin
Stations 3-6 in autotechnicon
Increasing grades of 70-95% ethanol
Stations 7-8 in autotechnicon
Acetone
Stations 9-10 in autotechnicon
Chloroform or xylene
Stations 11-12 in autotechnicon
Liquid paraffin
Main advantage of autotechnicon
Constant agitation with vertical oscillation
Process of wax impregnation under negative atmospheric pressure inside an embedding oven
Vacuum embedding
Vacuum embedding advantages
Hastens removal of air bubbles and clearing agent; promotes rapid penetration of impregnating medium
Fastest method for urgent biopsies, delicate tissues, and CNS tissues
Vacuum embedding
Mixture of paraffin and synthetic plastic polymer
DMSO
Melting point of DMSO
56-57°C
Substitute similar to Paraplast
Embeddol
Melting point of Embeddol
56-58°C
Semisynthetic tissue embedding material for eyes
Biobloid
Material used for tissue mat
Rubber
Properties of ester wax
Melting point: —
46-48°C, harder than paraffin, water-insoluble but soluble in 95% ethanol
Microtome used for ester wax sectioning
Sliding/base sledge microtome
Water-soluble waxes
Polyethylene glycols
Melting point range of polyethylene glycols
38-42°C or 45-56°C
Most common water-soluble wax
Carbowax
Properties of Carbowax
Miscible and soluble with water, hygroscopic, no need for clearing and dehydration
Uses of Carbowax
Used for enzyme histochemistry
Disadvantages of Carbowax
Hygroscopic, dissolves fishing out, crumbles during sectioning
Sectioning condition for Carbowax infiltrated tissue
Fish out immediately
Composition of Pearse wax
Composed of diethylene glycol, distilled water, and 40% formalin
Composition of Blank and McCarthy wax
Equal parts of 0.02% gelatin and 0.02% potassium dichromate
Purified form of nitrocellulose
Celloidin
Nitrocellulose is preferred for
Nervous tissue (brain), spx with large hollow cavities
Celloidin composition
2%, 4%, or 8% dissolved in equal parts of ether and alcohol
Celloidin advantages
Less shrinkage, less tissue distortion
Processing time for celloidin
Very slow (days to weeks)
Celloidin storage for bones, brain sections, teeth
70% alcohol for storage
Celloidin storage for whole eye sections
Gilson’s mixture (equal parts of chloroform and cedarwood oil)
Type of celloidin used for bones, brain, teeth
Wet celloidin
Type of celloidin used for whole eye sections
Dry celloidin
Preferred method for embedding tissue blocks
Nitrocellulose method (Low Viscosity Nitrocellulose)
Solubility of low viscosity nitrocellulose
Soluble in equal concentrations of ether and alcohol
Advantages of low viscosity nitrocellulose
Produces harder tissue block, allows thinner sections
Additive to prevent tissue cracking in nitrocellulose
Plasticizer (castor oil or oleum ricini)
Safety concern for nitrocellulose
Explosive
Rarely used; mostly for histochemical and enzyme studies, frozen sections; water soluble
Gelatin
Does not require clearing and dehydration
Gelatin
Tissue thickness for processing tissues impregnated with gelatin should be less than
2-3 mm
Gelatin uses ____ to prevent mold
1% phenol
Gelatin impregnating medium volume should be at least ___ the volume of tissue
25x