FAULTS IN TISSUE PROCESSING AND SECTIONING Flashcards
reason of brittle and hard tissue
prolonged fixation
prolonged dehydration
prolonged clearing
prolonged paraffin infiltration in overheated paraffin oven
drying out of tissue before actual fixation
failure to give special handling of tissues when treated with routine processing methods
remedy in brittle and hard tissue
Tissues may be softened by soaking in a small dish or bowl containing water with detergent, phenol or Molliflex.
causes of clearing agent turns milky as soon as tissue is placed on it
water not completely removed due to incomplete dehydration
remedy for the clearing agent that turns milky as soon as tissue is placed in it
Repeat dehydration with absolute alcohol, then clear again
give the reason
On trimming, tissue smells of clearing agent
Clearing agent not completely removed due to insufficient impregnation
give the remedy
tissue smells of clearing agent
Block is trimmed down nearest to the tissue, remaining wax is melted on embedding oven and paraffin impregnation is repeated, changing the paraffin at least once before blocking
give the reason
Tissue is opaque, section–cutting is difficult due to presence of alcohol
Insufficient clearing
give the remedy
Tissue is opaque, section–cutting is difficult due to presence of alcohol
Repeat clearing; if object has already been embedded, prolong clearing up to 12 hours then re–embed
reason for
Tissue shrinks away from wax when trimmed
Insufficient dehydration, therefore incomplete clearing and impregnation
remedy
Tissue shrinks away from wax when trimmed
repeat the whole procedure
reason
Tissue is soft when block is trimmed
incomplete fixation
remedy for
Tissue is soft when block is trimmed \
repeat fixation
reason for
Airholes found on tissue during trimming
incomplete impregnation
remedy for
Airholes found on tissue during trimming
repeat impregnation
reason for
On trimming, wax appears crystalline
contaminated wax
remedy for
On trimming, wax appears crystalline
re-embed in freshly filtered wax
reason for
Paraffin block after cooling is moist and crumbles
insufficient impregnation
remedy for
Paraffin block after cooling is moist and crumbles
repeat paraffin impregnation then re-embed
what are the faults in tissue processing
brittle or hard tissue
clearing agent turns milky as soon as tissue is placed in it
on trimming, tissue smells clearing agent
tissue is opaque, section-cutting is difficult due to presence of alcohol
tissue shrinks away from wax when trimmed
tissue is soft when block is trimmed
airholes found on tissue during trimming
on trimming, wax appears crystalline
paraffin block after cooling is moist and crumbles
reason
sections are too thick
wrong micrometer setting
remedy if the sections are too thick
microtome needs recalibration
give the reason
sections fall to form ribbons
surfaces and edges of the block are not parallel
horizontal surface of the block is not parallel to the knife
paraffin wax is too hard
knife is tilted too much
sections are too thick
knife is dull
give the reason if the sections roll up on cutting so that they adhere and get broken against the knife edge
knife is blunt
tilt of knife is too great
knife edge is dirty
remedy if the sections roll up on cutting so that they adhere and get broken against the knife edge
sharpen the knife
reduce the tilt
clean the knife edge
sections fall to form ribbons
reason: Surfaces and edges of the block are not parallel
remedy:
re trim the block
sections fall to form ribbons
reason: horizontal surface of the block is not parallel to the knife
remedy:
re adjust and re orient the block
sections fall to form ribbons
reason: paraffin wax is too hard
remedy:
coat horizontal edges of the block with wax of lower melting point
sections fall to form ribbons
reason: knife is tilted too much
remedy:
reduce the tilt
sections fall to form ribbons
reason: section that are too thick
remedy:
readjust the thickness of the sections
sections fall to form ribbons
reason: dull knife
remedy:
hone and strop
give the reason
ribbon is curved, crooked, or uneven, instead of straight
blunt or dull spot on the knife producing an irregular knife edge
edges of the block are not parallel nut round or wedge shaped
knife is not parallel to the block
ribbon is curved, crooked, or uneve, instead of straight
reason : ribbon is curved, crooked, or uneven, instead of straight
remedy :
adjust the knife so that knife edge will present a uniformly sharp edge to the black or sharpen it
ribbon is curved, crooked, or uneve, instead of straight
reason : edges of the block are not parallel but round or wedge shaped
remedy :
retrim the block
ribbon is curved, crooked, or uneve, instead of straight
reason : knife is not parallel to the block
remedy :
readjust the knife and block
reason why the sections turned compressed, wrinkled, or jammed
knife is blunt or dull
paraffin block is warm and soft
knife edge is coated with paraffin
sections are too thin
microtome set screw loose
tilt or knife is vertical
sections turned compressed, wrinkled, or jammed
reason: knife is blunt or dull
remedy:
resharpen the knife
sections turned compressed, wrinkled, or jammed
reason: paraffin block is warm and soft
remedy:
cool the block on ice water until firm
sections turned compressed, wrinkled, or jammed
reason: knife edge is coated with paraffin
remedy:
clean the knife edge
sections turned compressed, wrinkled, or jammed
reason: sections are too thin
remedy:
re adjust thickness of the section
sections turned compressed, wrinkled, or jammed
reason: microtome set screw is loose
remedy:
tighten the scree
sections turned compressed, wrinkled, or jammed
reason: tilt of knife is too vertical
remedy:
reduce the tilt
reason why the sections turned squashed (width of each section is less than of the block)
bevel knife is lost due to incorrect sharpening
remedy for the sections that turned squashed (width of each section is less than of the block)
re sharpen using knife back or automatic knife sharpener
reason why there’s a hole formed in the section
bubble or dirt formed in the embedding medium
tissue is not processed properly and will not form a section
under-processed portion of tissue bursts on contact with warm water
hard spot in tissue due to calcium
A hole is formed in
the section
reason: bubble or dirt formed in the embedding medium
remedy
re embed in freshlt filtered wax if necessary
A hole is formed in
the section
reason: tissue is not processed properly and will not form a section
remedy :
re process tissue
A hole is formed in
the section
reason: under processed portion of tissue bursts on contact with warm water
remedy:
re process tissue
A hole is formed in
the section
reason: hard spot in tissue due to calcium
remedy :
once embedded in paraffin wax, decalcification is impractical; use a base-sledge microtome with a wedge knife
reason for section of unequal thickness are produced
tilt of knife is too great or bevel is not cleared, hence object is compressed against the knife edge
clamp set screw on knife or block holder is loose
blocks are too large
blocks are too hard
reason for section of unequal thickness are produced
reason: tilt of knife is too great or bevel is not cleared, hence object is compressed against the knife edge
remedy:
reduce the tilt
reason for section of unequal thickness are produced
reason: clamp set screw on knife or block holder is loose
remedy:
tighten the screw
reason for section of unequal thickness are produced
reason: blocks are too large
remedy:
cut blocks into smaller fragments
reason for section of unequal thickness are produced
reason: blocks are too large
remedy:
cut blocks into smaller fragments
reason for section of unequal thickness are produced
reason: blocks are too hard
remedy:
soften the blocks in detergent or phenol
reason why
sections adhere to the knife or other parts of the microtome
static electricity due to low atmospheric humidity
knife edge is dirty
knife edge is dull
knife till is too great
Sections adhere to the
knife or other parts of
the microtome
reason: static electricity due to low atmospheric humidity
remedy:
breathe out or blow gently on the block and knife to break up static electricity, or boil water in the room to increase humidity
reason for ribbon split or lengthwise vertical scratches seen on sections
nicks or damage on the knife edge
dirty embedding
knife edge is dirty
tilt of knife too great
ribbon split or lengthwise vertical scratches seen on sections
reason: nicks or damage on the knfe edge
remedy:
sharpen the knife
ribbon split or lengthwise vertical scratches seen on sections
reason: dirty embedding
remedy:
re-embed in freshly filtered wax
ribbon split or lengthwise vertical scratches seen on sections
reason: dirty knife edge
remedy:
clean with xylene
ribbon split or lengthwise vertical scratches seen on sections
reason: tilt of knife too great
remedy:
reduce the tilt
reasons for sections are lifted from the knife on upstrokes
knife tilt is too great
knife is dull
paraffin is too soft or room temperature is warm
reasons for sections are lifted from the knife on upstrokes
reason: parafin is too sofy or room temperature is warm
remedy:
cool paraffin wax in ice water
reason for the resistance felt on the lower part of the section during cutting
tilt of knife is too small,
paraffin block is therefore compressed against the based of the knife towards the end of the stroke
remedy for the resistance felt on the lower part of the section during cutting
increase the tilt of the knife
reasons why there’s horizontal or parallel lines or furrows across the section (chatters) are seen
knife edge vibrations due to hardness of tissue
tilt of knife is too great
remedy for knife edge vibrations due to hardness of tissue
treat with phenol during processing or collodioniz
reason for
Section cut is
sometimes thin,
sometimes thick
knife is blunt
knife is not clamped properly
knife or block holder is loose
knife tilt is too small that block is compressed by bevel and section is not cut
reason
Knife makes a hard
metallic scraping or
ringing sound on
backstroke, when
section is cut
tilt of knife is too slanted or too big
tissue is too hard
knife is too thin
reason why Frozen tissue
crumbles and comes
off the block holder
when cut
freezintg is not adequate
Frozen tissue chips
into fragments when
cut
tissue is frozen too much
