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Common Artifacts and Remedies in Histopathology A
Common Artifacts and Remedies in Histopathology A
Common Artifacts and Remedies in Histopathology A
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Abstract
The essence of preparing a biological specimen for microscopic studies is to get
adequate and accurate medical information that is a true representation of the
specimen, be it for research or diagnostic purposes. An artifact can be defined as
unrelated, self-colored artificial feature found in tissue sections. This has been an
age long cause of misinterpretation leading to misdiagnosis to microscopists. The
processing of specimen for medical information is subject to a procedure that
results in a tissue fit for diagnosis and interpretation. The procedures themselves
are subject to human and material errors and the result is an artifact that in the
least may interfere with adequate diagnosis or at the most render the tissue so
distorted as to be undiagnosable. The need to recognize these artifacts and attempt
to overcome them is the single biggest challenge in the histopathology Laboratory.
This article focused on identifying artifacts and their potential cause so that
misinterpretation and difficulty in diagnosis can be overcome, and help
microscopist to come into definite diagnosis.
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EKUNDINA AND EZE ARTIFACTS
Examples of pre-fixation artifact include: electrocautery[ Meghana and Ahmedmujib
i. Artifacts due to surface preparation (2007), (Seoane, 2004). This causes the surface
The excision margins of fresh surgical epithelium to be forced through the connective
specimens are sometimes marked with coloring tissue, producing small “pseudocysts” and also
agents to allow appropriate orientation of the causes loss of cytoplasmic and nuclear features
specimen and assessment of these margins (Camacho et al., 2008).
microscopically Krishnanand et al., (2010).
Biopsy area is also prepared by using tincture
iodine. Reagents commonly used for surface
marking include Indian ink, Silver nitrate,
Alcian blue and Alcian green.
REMEDY
These artifacts can be prevented by handling
Plate 1: Artifacts due to surface preparation and
tattoo pigmentation: margin of a skin biopsy marked the specimen gently, avoiding applying
with silver nitrate. The reagent stains the soft tissues unnecessary force and holding the tissue in
and has penetrated into the dermis an area away from the lesional tissue.
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EKUNDINA AND EZE ARTIFACTS
REMEDY
Artifact due to contaminations can be
prevented by careful manipulation of
specimen at every step of preparation and
cleaning dissecting board after treating each
Plate 4 (Artifacts due to contamination). Starch specimen.
granules: Starch granules in a typical starch
granuloma stained with H&E and Polarized light
FIXATION ARTIFACTS
(b) Specimen-specimen contamination Although fixation is necessary to avoid
artifact probably occurs during dissection diffusion of soluble tissue components and
where tissue from a previous specimen is decomposition, it by itself constitutes a major
transferred via the instruments used (such as cause of artifact. If the procedure is not carried
scalpel blades) or from fragments which out under optimal conditions, if fixative does
remain on the dissecting board surface. not have proper access to the tissues, or because
of the nature and quality of the particular
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EKUNDINA AND EZE ARTIFACTS
reagent used, artifact can occur. (McInnes,
2005:Weir, 1976). The most commonly used
fixative is 10% formalin. The concentration of
the formalin, contamination and prolong
fixation time leads to difficulty in sectioning of
the specimen. Fixation artifacts arise due to
formalin, mercuric chloride and picric acid used
in various fixative agents which causes Brown- Plate. 8 streaming of glycogen, Seen in hepatic tissue
Black granular and yellow stains distributed
randomly throughout the tissues. (Samar et al., iii. Diffusion artifacts: Materials may
2014) sometimes diffuse out of the tissue. Apart
from large molecules, small molecules like
inorganic ions and biogenic amines can be
lost from tissue. For example; when cut end
of adrenal gland tissue are placed in iodate
for iodate reaction, late cholamines can be
seen leaving the tissue as a red cloud of
aminochromes. (Samar et al., 2014)
STAINING ARTIFACTS
i. Artifacts due to residual wax:
Plate 10 water bubble trapped in section Residual wax in a section will prevent
Artifacts in water bath: Air bubble entrapment;
penetration of both aqueous and alcoholic dye
Section of lymph node showing circular darkly
stained areas with pale centers and surrounding radial solutions leaving areas totally devoid of stain.
cracks. These faults indicate collapsed bubble Traces of residual wax have a subtle effect on
artifact; H&E Stain. nuclear staining producing small patches in
sections where nuclei appear muddy and lack
iii. Artifacts related to oven/hot plate: detail (Krishnanand et al., 2010; Wynnchuk,
1990)
The temperature setting of the oven/hot plate
should be approximately that of the melting
point of paraffin. This type of artifact arises due
to increase in temperature beyond melting point
of paraffin. If the oven is too hot there may be
distortion to the cells causing dark pyknotic
nuclei or nuclear bubbling. Cells appear to be
completely devoid of nuclear details. Plate 12 Artifact due to residual wax
Thick coat of section adhesive will take the stain Plate .13 Artifacts due to incomplete staining
and background stain may be detected resulting
in irregular, poor quality sections. Mounted, MOUNTING ARTIFACTS
unstained section left uncovered can be During mounting, air bubble entrapment,
contaminated with various materials such as residual water and excessive use of mounting
media will bring artifactural changes. These
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EKUNDINA AND EZE ARTIFACTS
changes can be avoided if proper mounting Camacho FA, Jornet PL, Torres M J, Orduna A
technique is incorporated. (2008). Analysis of the histopathological
artifacts in punch biopsies of the normal oral
MICROSCOPY ARTIFACTS mucosa. Med Oral Patol Oral Cir Bucal.
Dust particles, impurities which can be internal 13(10):E636-9. .
or external present on slide will bring
artifactural changes. Fatty films are observed Ficarra G, Clintock B, Hansen L (1987).
due to unclean lenses or greasy deposits on Artifacts created during oral biopsy procedures.
eyepieces due to eyelashes will results in foggy J Craniomaxillofac Surg. 15(1):34-7.
appearance.
Grizzle WE, Stockard C, Billings P (2001). The
Remedy effects of tissue processing variables other than
Ensure that microscopes are well kept and fixation on histochemical staining and
regularly maintained immunohistochemical detection of antigens. J
Histotechnology. 24(3):213–219.
Section Preservation Artifacts Krishnanand P, Kamath V, Nagaraja A. (2010).
This include majorly drying artifact and Artefacts in Oral Mucosal Biopsies A Review.
bleaching artifact Orofac Sci, 2(1); 57-62.
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