Aquitania, Mary Christelle G. Date Submitted: UST Medical Technology Intern Histolopathology

Faults occurring during Trimming/Cutting of Paraffin Blocks

Trimming is a process wherein the paraffin block is exposed for actual cutting after when the wax is solidified and removed from the mold. The sides, top and bottom of the tissue block are trimmed until leveled perfectly and all sides are parallel to each other. An old knife or blade may is used for this procedure yet still must be relatively sharp to avoid tissue block damages. After coarse trimming, the block is then placed in the microtome for final trimming and cutting. To have a fine thin trimming, setting the thickness adjuster at 15mm or by advancing the block using coarse feed mechanism is done, according to Gregorios’ Histopathologic Techniques. The block is clamped on the chuck must be retracted enough to ensure that knife does not touch the block on initial down stroke. The surface is trimmed first until the tissue surface is partly exposed. Regular cutting rhythm should be maintained. Thin ribbons are taken out at a time to prevent the block from cracking. Factor considered for the cutting rate of the tissue blocks are the type of tissue, size of the slick, and the model of the block. For routine histologic procedures, usually sections are cut between 4-6μ.

However, during the processing and cutting of the section, several circumstances and difficulties may be encountered. A medical technologist should be alert taking note of faults, which if not immediately remedied, shall cause entire failure of process, poor sectioning, and ultimately improper evaluation of the tissue in question. The most commonly encountered difficulties observed during tissue processing with its reason and remedy is placed in the table.

FAULTS REASON REMEDY

Brittle or hard tissue

Prolonged fixation

Tissue may be softened by soaking in a small dish containing water with

detergent, phenol or Molliflex

Prolonged dehydrationProlonged clearing

Prolonged paraffin infiltrationOverheated paraffin oven

Drying out of tissue before actual fixation

Clearing agent turns milky as soon as tissue is placed in it

Water not completely removed (incomplete

dehydration)

Repeat dehydration with absolute alcohol, then repeat

clearing

Upon trimming, tissue smells of clearing agent

Clearing agent is not completely removed due to insufficient impregnation

Blocked is trimmed down nearest to the tissue. The

remaining wax id melted on embedding oven and paraffin

impregnation is repeated, changing the paraffin at least

once before blocking.

Tissue is opaque, section cutting is difficult due to the presence of alcohol

Insufficient clearing

Repeat clearing; id object has already been embedded,

prolong oven and paraffin impregnations repeated,

changing the paraffin at least once before blocking

Tissue shrinks away from wax when trimmed

Insufficient dehydration, therefore incomplete clearing

and impregnationRepeat the whole procedure

Tissue is soft when block is trimmed

Incomplete impregnation Repeat whole procedure

Air holes on tissue during trimming

Incomplete impregnation Repeat impregnation

On trimming, wax appears crystalline

Contaminated waxRe-embed in freshly filtrated

waxBlock not cooled rapidly

enoughParaffin block, after cooling, is moist and crumbles

Insufficient paraffin impregnation

Repeat paraffin impregnation, then re-embed

Faults Observed during Section-Cutting

FAULTS REASON REMEDY

Sections fail to form ribbons

Surfaces and edges of the block are not parallel

Re-trim the block

Horizontal surface of the block is not parallel to the

knife

Re-adjust and re-orient the block

Paraffin wax is too hardCoat horizontal edges of the

block with wax of lower melting point

Knife is tilted to much Reduce the tilt

Sections are too tickReadjust the thickness of the

sectionsHone and strop

Sections roll up on cutting so that they adhere and get broken against the knife edge

Knife is blunt Sharpen the knifeTilt of knife is too great Reduce the tile

Knife edge is dirty Clean the knife edgeRibbon is curved, crooked or uneven instead of straight

Blunt of dull spot on the knife, producing an irregular knife

edge

Adjust the knife so that the knife will present a uniformly

sharp edge to the block, or sharpen

Edges of the block are not parallel but round wedge

shape

Re-trim the block

Knife is not parallel to the block

Readjust knife and block

Paraffin is impureRepeat impregnation using

pure wax

Sections are compressed, wrinkled or jammed

Knife is blunt or dull Re-sharpen the knifeParaffin block Is warm and

softCool the block on ice water

until firmKnife edge is coated with

paraffinClean the knife edge

Sections are too thin Readjust thickness of sectionMicrotome set screw is loose Tighten the screw

Tilt of knife is too vertical Reduce the tiltSections are squashed (width of each section is less than that of block)

Bevel of knife is lost due to incorrect sharpening

Re-sharpen, using a knife back or automatic knife sharpener

A hole is formed in the section

Bubble or dirt formed in the embedding medium

Re-embed in freshly filtered wax if necessary

Hard spot in tissue due to calcium

Once embedded in paraffin wax, decalcification is

impractical; use a base-sledge microtome with a wedge knife

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

Reduce the tilt

Or blockholder is looseBlocks are too large

Tighten the screw

Block are too hardCut blocks into smaller

fragmentsSoften the blocks in detergent

or phenol

Sections adhere to the knife or other parts of the microtome

Static electricity due to low atmospheric humidity

Breather out or blow gently on the block and knife to breakup static electricity, or boil water

in the room to increase the humidity

Knife edge is dirty Clean the knife edgeKnife edge is dull Sharpen the knife

Knife tilt is too great Reduce the tilt

Ribbon is split or lengthwise vertical scratches are seen on sections

Nicks or damage on the knife edge

Sharpen the knife

Dirty embedding Re-embed in filtered waxKnife edge is dirty Clean knife edge with xylene

Tilt of knife is too great Reduce the tiltSections are lifted from the Knife tilt is too great Reduce the tilt

knife on upstrokesKnife is dull Sharpen the knife

Paraffin is too soft or room temperature is warm

Cool paraffin wax in ice water

Resistance is felt on the lower part of the section during cutting

Tilt of knife is too small, paraffin block is therefore

compressed against the base of the knife towards the end of

stroke

Increase the tilt

Horizontal or parallel lines or furrows across the section (“Chatters”) are seen, forming thin and thick zones

Knife edge vibrate due to hardness of tissue

Treat with phenol during processing or collodionize

Tilt of knife is too great Reduce the tilt

Section cut is sometimes thin, sometimes thick

Knife is blunt Sharpen knifeKnife is not clamped properly Adjust the knife

Tilt of knife is too great Reduce the tilt

Knife or block holder is looseTighten adjusting and locking

screwsKnife tilt is too small that

block is compressed by bevel and section is not cut

Increase the tilt

Knife makes a hard metallic scrapping or ringing sound on backstroke, when section is cut

Tilt of knife is too slanted or too big

Readjust the angulation of the knife

Tissue is too hardTake fresh block treated with

phenol during processingKnife blade is too thin Change the knife

Frozen tissue crumbles and comes off the block holder when cut

Freezing is not adequate Refreeze the tissue block

Frozen tissue chips into fragments when cut

Tissue is frozen too hard Warm the tissue with fingers