Processing Flashcards
incomplete dehydration
accounts for the vast majority of processing problems and results in mushy tissue
alcohols
the most common dehydrating reagents
clearing agents
must be miscible with both the dehydration agent and the infiltration medium
primary purpose is to remove the alcohol used to dehydrate the tissue
prolonged clearing
results in hard, brittle tissues
xylene
most commonly used clearing agent, turns cloudy in the presence of water
toluene
does not over harden tissue as much as xylene
overdehydration
seen as microchatter in H&E stained slides, commonly seen in biopsy specimens, can be corrected by processing biopsies separately from other tissues and decreasing the amount of time in the dehydrating solutions
poor processing
evidenced by poor nuclear staining, often caused by water remaining in the tissue when it is placed in the clearing agent which results in poor clearing and infiltration, can also be caused by too much heat during processing
melting point
high melting point results in hard blocks that section thinly but ribbon poorly, while low melting point ribbons easily but doesn’t section as thin. 55-58C is common melting point
diagonal embedding
makes sectioning much easier
forceps metastasis
when fragments of tissue are transferred between blocks if the forceps aren’t wiped down between specimens
paraffin crystal size
smaller is better, allows for better support of the tissue
mushy in the middle
tissue was cut too thick during grossing and needs to be reprocessed
dehydrant
alcohol for removal of water
clearing agent
xylene to remove the alcohol and make the tissue receptive to paraffin, high index of refraction renders tissue transparent
infiltrating medium
paraffin infiltrates the tissue and gives it support
open tissue processor
rarely used because of reagent evaporation
closed tissue processor
preferred method
Ethanol (ethyl alcohol)
clear, colorless, flammable
fast, best dehydrant
hydrophilic
70-95-100% steps help reduce tissue shrinkage
Methanol (methyl alcohol)
clear, colorless, flammable, poisonous
rarely used
except for fixation of blood smears
Isopropanol (Isopropyl alcohol)
flammable, toxic
excellent substitute for ethanol
eosin can’t stain with it
doesn’t harden or shrink tissue as much as ethanol
Acetone
volatile, flammable dehydrates rapidly, cheap excessive shrinkage absorbs water when exposed to air hard to maintain solution levels in open air processors
xylene
most commonly used clearing agent
can overharden tissues
intolerant of water left in the tissue
flammable hazardous neurotoxin, do not pour down the sink
toluene
flammable, more volatile than xylene
does not overharden tissue as much as xylene