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Artifacts in Histologic Sections

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Bayan Sa

on 15 December 2013

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Transcript of Artifacts in Histologic Sections

Artifacts in Histologic Sections
design by Dóri Sirály for Prezi
Artfacts
Artifacts
are structures or features in tissue that interfere with normal histological examination. These are not always present in normal tissue and can come from outside sources. Artifacts interfere with histology by changing the tissues appearance and hiding structures.

pre-fixtion
Tissue-Processing ArtifactsPoor Processing
Extensive loss of architectural detail and clarity within loose connective tissue.

These include
too short a processing cycle
inappropriate choice of reagents
use of exhausted reagents
an error in the processing machine .
Section mounting
various form of mechanical damage produced during section cutting and flotation , together with a range of contaminants from a variety of source , are commonly encountered in sections
Staining artifacts
Prevent the penetration of both aqueous and alcoholic dye solution leaving area totally devoid of stain
Pre histology:

The structure and features of animal cells or tissues are being incorporated to the tissues.  For instance freckles that are found in skin samples and ink from tattoos are pre histological type.
Post histology:

Artifacts can result from tissue processing. Processing commonly leads to changes like shrinkage, washing out of particular cellular components, color changes in different tissues types and alterations of the structures in the tissue.

A common example is mercury pigment left behind after using Zenker's fixative to fix a section
Microtome :
 
is a mechanical instrument used to cut biological specimens into very thin segments for microscopic examination
Microtome
Knife lines (vertical striations in section)
Fine cracks or micro-chatter
Coarse chatter
• Clamping mechanism not securely locked
• Very hard or large specimen
• Poor processing
• Insufficient clearance angle
• Sectioning too rapidly
• Worn microtome
• Tissue over-processed
• Block too cold
• Cutting too fast
• Clamping mechanism not securely
locked
• Clearance angle needs adjustment
• Damaged knife or blade used
• Poor processing
• Hard material such as calcium in block
• Debris in unfiltered wax
• Buffer salts precipitated in specimens
Bubbles Under Section
Air bubbles trapped in a section after flotation and mounting can collapse on drying leaving zones which crack and fail to adhere properly to the slide. These regions often display altered staining . The bubbles may be caused by poor flotation technique where a section is dropped rather than pulled gently across the water surface. Alternatively, bubbles already present in the bath can be dislodged by the slide and rise up under the section. Freshly boiled water used in flotation baths is less likely to produce bubbles.

Section of lymph node showing circular darkly stained areas with pale centers and surrounding radial cracks. These faults indicate collapsed bubble artifact ; H&E
Holes from roughing
This effect may occur when sectioning lymph node and other very cellular organs. Excessively rough trimming pulls tissue fragments from the block face and these appear as holes in subsequent thin sections . The effect can usually be seen when floating sections on the water bath.
Roughing effect in a section of
Lymph node . The parallel alignment
Seen in this particular example is
Not always evident.
Sneeze artifact
Squamous cell contamination on sections is not uncommon and usually originates from fingers or scalp and, less commonly, sneezes or coughs
A sneeze-generated deposit ( contaminant )
which contains Squamous cell , cell debris , some bacteria and an amorphous background that stains weakly with hematoxylin
Displacement of components
Displacement of cancellous bone in
a trephine section
stained for reticulin .
Residual wax
Incomplete staining
Incomplete staining with one dye in multi-step procedure may result from inadequately filled staining dish
Stain deposit
May arise from undissolved stain, stain precipitate or any solid component in unfiltered staining solution.
Contaminated staining solution
Contamination of staining solution by microorganism

Mucus contamination
Zonal Fixation
It occurs when there are differing degrees of fixation at different levels within the specimen.

causes:
Insufficient time in fixative
Too large specimen
It forms when acid formalin reacts with hemoglobin.

It is most often seen in:
tissues rich in blood such as spleen.
tissues which have had prolonged fixation.

The artifact can be prevented by :
using buffered formalin solution
restricting fixation time.
If the fixative solution in which the tissues are sitting is grossly brown to red, then place the tissues in new fixative.


The artifact can be removed by:
treating the section with saturated alcoholic picric acid solution prior to staining
Fault due to poor processing
1-
Fault
: the tissue is shrunken away from wax
Cause
: insufficient dehydration
Remedy
: reprocess
2-
Fault:
the tissue is too soft when block is trimmed
Cause:
Insufficient fixation
Remedy
: reprocess

3-
Fault:
specimen crumbles and drops out of the wax leaving a rim of wax as a section
Cause:
-insufficient infiltration
-overheated paraffin bath causing tissue to become hard and brittle
Remedy
: -re infiltrate and re-embed
-service the paraffin bath

4-
Fault
: tissue is dried out or mummified
Cause
: mechanical failure of tissue processing machine or a basket was out of balance and hung up
Remedy
: place the specimen in the following rehydration solution for 18-4 hrs
Sodium carbonate - 1.0gm
dist. Water - 70.0ml
Absolute ethyl alcohol - 30.0ml
Re hydrate the reprocess
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