Biopsy - An Overview

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ESSENTIAL DENTAL KNOWLEDGE
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J Can Dent Assoc 2012;78:c75
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Oral Soft-Tissue Biopsy: An Overview

Sylvie-Louise Avon DMD, MSc, PhD, FRCD(C);
Hagen B.E. Klieb DMD, MSc, FRCD(C)
Abstract
Types of Oral Lesions
The oral cavity is lined with strati-
fied squamous epithelium overlying
mesenchymal tissues. Benign and
Proper management of an oral mucosal lesion begins with diagnosis, and the gold malignant neoplasms may originate
standard for diagnosing disease, oral or otherwise, is tissue biopsy. The oral environ- from any of these tissues. Lesions of
ment, which is moist and confined, poses challenges for collecting a viable tissue epithelial origin are typically white
sample that will be suitable for diagnosis. These challenges are further compounded or red. Hyperplasia of the epithelium
by the myriad of biopsy techniques and devices now available. In addition, certain and accumulation of keratin produce
oral subsites are subject to diagnostic pitfalls that necessitate modifications of tech-
a white lesion (Fig. 1), whereas epi-
nique. This article provides an overview of the oral soft-tissue biopsy and highlights
thelial atrophy allows greater visual-
some potential pitfalls.
ization of the underlying vasculature,
which leads to a red appearance. The
epithelial integrity may be disrupted
(a process known as ulceration), and
resultant tissue proliferation may
B
iopsy is the removal of a tissue be exuberant, producing a papil-
sample from a living body lary or verruciform configuration.
with the objective of providing Representative biopsy of an epithelial
the pathologist with a representa- lesion must include the full epithelial
tive, viable specimen for histopatho- thickness with some supporting con-
logic interpretation and diagnosis.1 nective tissue to allow assessment for
This approach is used for all tissues invasive carcinoma but also to provide
physical support for the specimen.
of the body, including those of the
There are no strict size criteria, but
oral cavity, where a wide spectrum
tiny samples impede processing and
of disease processes may present. The
interpretation of the specimen.1 For a
dental clinician should be aware of thin plaque, a sample only a few milli-
the various biopsy techniques that are metres deep will typically suffice, and
available for the oral tissues, as well as there is no need to extend deep into
the challenges specific to these tissues. the connective tissues or musculature.
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Figure 1: Thick, homogenous leukoplakia Figure 2: Benign salivary gland neoplasm Figure 3: Asymptomatic ulcerated erythrol-
affecting the posterior buccal mucosa is (pleomorphic adenoma) presenting as a eukoplakia diffusely affecting the right
characterized by hyperplasia of the epithel- dome-shaped submucosal palatal swelling lateral tongue. Representative incisional
ium and accumulation of keratin, produ- with a surface of intact pink mucosa. biopsies were obtained and showed,
cing a white lesion. Histologic assessment from left to right, areas of thick keratosis,
showed epithelial hyperplasia, hyperkerato- erythema and induration.
sis and mild dysplasia.
However, a sample of greater depth will be needed expertise in the diagnosis and management of oral
if there is exuberant epithelial thickening due to disease is always a possibility.
epithelial hyperplasia, elongation of the rete ridges
or accumulation of keratin, as may be the case in Indications for Biopsy
verrucous carcinoma or verrucous hyperplasia. Biopsy is essential if there is any clinical sus-
Lesions of mesenchymal origin arise below the picion of malignancy, such as an enlarging mass,
epithelium and appear as dome-shaped swellings chronic ulceration, tissue friability, induration
(Fig. 2). The overlying epithelium is often intact, on palpation or persistence of mucosal changes
but its coloration reflects the contents of the lesion: despite removal of local irritants.2-4 New or enlar-
red–purple if vascular, blue if mucinous or yellow ging pigmented lesions, especially those with an
if adipose, lymphoid or neural. The representa- irregular border and nonhomogenous colora-
tive biopsy sample must be of sufficient depth, but tion, should be biopsied to exclude mucosal mel-
the appropriate depth varies from one lesion to anoma. Entities that appear to be clinically benign
another, depending on the thickness and location or reactive (e.g., pyogenic granuloma or muco-
of the mass. cele) may be excised for esthetic or functional rea-
sons, but the tissues should be submitted for histo-
Preliminary Examination of the Lesion logic analysis to confirm the clinical impression.
A comprehensive dental examination will Lichen planus, mucous membrane pemphigoid,
include both visual and tactile assessment of the pemphigus vulgaris and other immune-mediated
oral soft tissues. Once an oral lesion has been disorders may present with widespread mucosal
identified, the clinician should undertake a stan- erythema and ulceration, and biopsy is essential
dard protocol, beginning with eliciting and docu- for definitive diagnosis.
menting the pertinent history, including duration,
Contraindications for Biopsy
any antecedent event, symptoms and changes in
appearance, as well as prior diagnostic and thera- Significant hemorrhage may accompany biopsy
peutic measures. The location, size, colour, and of a vascular lesion, and caution should therefore
consistency or texture of the lesion should be be exercised in the biopsy of any lesion with red,
documented, which may be facilitated by pho- purple or blue coloration or with blanching or
tography. The differential diagnosis will guide pulsation on palpation. Location of the lesion in
management decisions, including the decision to an esthetic region (e.g., vermilion border of the
obtain a biopsy sample. Referral to a clinician with lip) is not a strict contraindication, but referral
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e f g
Figure 4. Homogenous leukoplakia affecting the right lateral and ventral areas of the tongue (a). Incisional biopsy is used to
sample a thick, keratotic region at the anterior aspect of the tongue. Infiltration of local anesthetic (b) is followed by tracing of
an ellipse (c and d). The anterior edge of the ellipse is gently raised with tissue forceps, which allows detachment of a canoe-
shaped sample (e and f). Hemostasis is achieved with single interrupted sutures (g).
to a specialist should be considered in such cases. Biopsy Techniques

Similarly, certain oral subsites, such as the floor
of the mouth, may be challenging to access, dif- Scalpel biopsy, for both incisional and exci-
ficult to provide hemostasis and risks damage to sional procedures, is the most common tech-
nique and generally produces the most satisfactory
anatomic structures (e.g., submandibular duct). It
samples.
is unwise to proceed if one is uncomfortable with
either the surgical procedure or the prospect of Incisional Biopsy
relaying devastating results to the patient. Incisional biopsy provides a representative
Standard biopsy procedure may need to be sample of tissue for diagnostic purposes. It is the
modified and medical clearance obtained in the method of choice when the differential diagnosis
case of medically compromised patients, including includes malignancy. Its accuracy is relative, since
those with severe or poorly controlled systemic by nature it does not allow study of the entire
diseases such as coronary artery disease, renal or lesion. Should the clinician be uncertain about
hepatic impairment, and various endocrinopathies the most appropriate site from which to obtain the
and immune-compromised states. Clearance from biopsy sample, referral to a specialist is necessary. 5
the primary medical caregiver and baseline sero- The clinician should attempt to sample the
logic studies should be considered for patients tissue that has been most severely and signifi-
with significant risk of hemorrhage due to under- cantly affected. In cases of epithelial dysplasia, the
lying bleeding diatheses related to anticoagu- severity of the epithelial changes or the presence
lant therapy, thrombocytopenia or any of various of carcinoma will be correlated with the clinical
inherited coagulopathies (e.g., hemophilia or Von appearance. For example, a thin white plaque is
Willebrand disease). Patients undergoing bisphos- less likely to harbour high-grade dysplasia than a
phonate therapy or radiotherapy may be predis- thick white plaque or erythematous, ulcerated and
posed to osteonecrosis if the biopsy procedure indurated regions. Multiple biopsy samples may
exposes bone. be required if the lesion is extensive or shows a
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Figure 5. Pink nodule affecting the buccal Figure 6. Thin, homogenous leukoplakia Figure 7: Widespread, thick leukoplakia
mucosa and located adjacent to the affecting the lateral and ventral areas of affecting the floor of the mouth and the
occlusal plane is consistent with an irritation the tongue is suitable for incisional punch mandibular gingiva is excised with a laser to
fibroma. Elliptical excisional biopsy facili- biopsy. The punch biopsy device is applied help minimize hemorrhage and discomfort
tates primary-intention healing. with a downward and twisting motion. (photograph courtesy of Dr. S. Trembley).
variety of clinical presentations1,2,5 (Fig. 3). Classic Punch Biopsy

teaching includes selecting a site at the periphery Punch biopsy may be used for either incisional
of the lesion to ensure inclusion of healthy tissue biopsy or excision of a small lesion at an access-
in the sample. However, the principle guiding site ible site. The lateral tongue and buccal mucosa
selection should be acquisition of the most rep- are appropriate sites for punch biopsy, as it must
resentative sample; an attempt to include tissue be feasible for the device to approach the mucosal
from the periphery may inadvertently lead to surface perpendicularly. The punch is placed on
underdiagnosis.1,5 the lesional tissue, and a downward, twisting
The technique used for incisional biopsy is motion is applied (Fig. 6). The tissue core is then
usually straightforward and is illustrated step-by- severed at the base with curved scissors. The cir-
step (Fig. 4a to 4g). An elliptical incision, with a cular wound makes approximating the edges more
length-to-width ratio of 3:1, is made with a size
difficult than is the case with an elliptical shape.
15 scalpel blade. The elliptical shape facilitates
Punch biopsy is not appropriate for vesiculobul-
primary-intention closure. The inferior incision is
lous diseases, as the twisting action would detach
made first, so that hemorrhage does not obscure
the epithelium and prevent proper assessment of
the surgical field. The anterior tip of the ellipse is
the interface between epithelium and connective
gently lifted with tissue forceps, and the base is
tissue that is necessary for subclassification of such
severed.
lesions.
Excisional Biopsy
Electrosurgery and Laser Techniques
Excisional biopsy is the complete removal of
Electrosurgery and laser techniques produce
a lesion for functional and aesthetic purposes,
as well as to confirm the clinical diagnosis. This thermal artifacts that may hamper histologic
is appropriate only if the lesion is almost cer- interpretation; accordingly, these methods should
tainly benign.6 The size, accessibility and regional be used with caution for diagnostic biopsy or when
anatomy of the lesion must all be considered. information from the margins is required. Lasers
Small, pedunculated, exophytic lesions in access- may be of great value, however, in managing a
ible areas are excellent candidates for excisional wound left by scalpel biopsy in areas of the mouth
biopsy. An ellipse is traced around the lesion, with where closure is difficult or inappropriate. A laser
the blade angled toward the centre of the lesion produces a zone of thermal coagulation smaller
(Fig. 5). This produces a wedge-shaped specimen than that of electrosurgery, but still, a 0.5-mm
that is deepest under the centre of the lesion and margin should be maintained between the cut and
leaves a wound that is simple to close. the representative area to be sampled. This tech-
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nique may produce good local hemostasis and

minimal postoperative discomfort (Fig. 7). Biopsy Procedure
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Adjunctive Techniques Armamentarium DES CONNAISSANCES
Brush biopsy has been advocated as a screening

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The minimal requirements are as follows: l’Association dentaire canadienne
modality for innocuous lesions that may otherwise • blade handle and no. 15 blade
not be sampled. With this method, a stiff brush • fine tissue forceps (preferably Adson forceps)
is used to collect cells from all epithelial layers • syringe and local anesthetic
• retractor appropriate for the site
through application of firm pressure with a rota-
• sutures, if needed
tional movement. Pinpoint bleeding indicates suf-
• needle driver
ficient depth of cell collection. 3,4 The sample is • curved scissors
transferred to a glass slide and sent to the labora- • hemostatic agents
tory for analysis. 3,4,7 If atypical cells are found, (silver nitrate or absorb-
conventional biopsy is also required. Routine use able gelatin sponge)
of brush biopsy remains debatable, especially • gauze sponges
given the accessibility of oral lesions for conven- • specimen bottle
containing 10% neutral
tional biopsy.2
buffered formalin
Adjunctive diagnostic aids, including nuclear
• biopsy data sheet Figure 8. Basic armamentarium for biopsy.
stains and light sources, are marketed to assist in
the detection of early cancerous changes. Staining
with a metachromatic dye (toluidine blue) can help
to define subtle erythroplakia, which facilitates
complete excision. Pilot studies with autofluor- may result in recession and ultimately esthetic
escence, to delineate field changes around high- defects and exposure of the root.
grade dysplasia and cancers and thereby guide Biopsy of the lip may be accompanied by brisk
excision, have been promising, but the potential hemorrhage, and the mobility of the labial tis-
for effects on disease recurrence requires further sues further complicates what would otherwise be
study.8 a simple procedure. Local hemostasis and tissue
stabilization are managed better if the assistant
Special Considerations and Pitfalls firmly grips the lip by placing the thumb and index
finger on either side of the lesion. Although biopsy
Rapid proliferation of a carcinoma can out-
is unlikely to produce a major deformity, esthetics
pace the nutrient supply, resulting in necrosis and
may be compromised, especially if an incision
ulceration. Biopsy of a sample from the centre of
crosses the vermilion border. Local paresthesia
such an ulcer, or its base, results in a nonspecific
may occur if nerves are severed.
diagnosis. Rather, the adjacent intact mucosa,
Tongue lesions present similar difficulties. An
often configured as a raised border, should be the
assistant can stabilize the tongue by wrapping the
biopsy target. Similarly, for vesiculobullous dis-
tip with gauze. For closure of tongue wounds,
orders, there may be widespread ulceration, but
deep sutures should be placed when indicated, and
diagnosis necessitates sampling intact mucosa in
mucosal sutures should be placed fairly close to
proximity to the ulceration. Extensive epithelial
each other. Inadequate suturing will quickly lead
sloughing can make this type of biopsy extremely
to reopening of the wound, with resultant hemor-
challenging.
rhage and delayed healing.
The bound tissues of the gingiva and hard
palate preclude the use of sutures in these areas. Biopsy Procedure
Healing is by secondary intention, and denuded
bone and associated discomfort may persist for sev- Armamentarium
eral weeks. Palatal biopsy must take into account The minimal requirements to perform the
the underlying vascular anatomy. Gingival biopsy biopsy procedure are shown in Fig. 8.
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Figure 9: (a) Crush artifact, whereby excessive compressive force has produced an impression of the teeth of the tissue
forceps within the tissue. Cautery artifact (b) caused by a laser device distorts the cytologic detail around the perimeter of the
specimen. Formation of ice crystals (c) distorts the tissue architecture in a sample that became frozen while being shipped to
the pathologist.
Consent be used with care to prevent loss of the specimen

Verbal and written informed consent should and suction-induced artifact. Gauze mounted on a
be obtained before any biopsy. The surgical details hemostat is sufficient in most cases.
should be discussed with the patient, as well as Tissue artifacts may originate from a number
potential complications (usually discomfort, of sources, but all interfere with histologic inter-
swelling, infection and hemorrhage). Surgery adja- pretation. Crush artifact is common and is typ-
cent to nerves can result in transient or permanent ically due to inappropriate compression from
paresthesia. Esthetic considerations are applicable forceps. Often the forceps teeth will leave an
to the gingival esthetic zone and the vermilion impression in the tissue (Fig. 9a). Thermal arti-
border of the lip. Procedures in the floor of the fact accompanying use of laser or electrosurgery
mouth can damage or obstruct the salivary ducts. devices obscures cytologic details around the per-
Reactive lesions may recur, and re-excision may be imeter of the specimen (Fig. 9b) and is especially
required. problematic when information about the status of
the margin is required. In colder climates, tissues
Anesthesia may freeze during mailing; freezing severely dis-
Administration of local anesthetic is generally torts the histologic architecture (Fig. 9c).
simple and straightforward. Lidocaine 2% com- On occasion, it may be necessary to specify
bined with epinephrine (for local hemostasis), at the orientation of the specimen if information
a ratio of 1:100 000, is commonly used. Typically, about the completeness of excision is required,
less than one carpule, or 1.8 mL, is required. The for example, when excising high-grade epithelial
local anesthetic should be infiltrated deep to or in dysplasia. Orientation is accomplished by placing
a field around the biopsy site. Intralesional admin- one or several sutures on known margins. If the
istration of anesthetic should be avoided, as it may specimen is thin, it is advisable to place it on
distort the tissue or produce artifact. Regional a piece of paper, with the connective tissue side
block may also be used, although the benefit of down, for at least 1 minute to ensure that the
epinephrine is lost. sample stays flat during fixation. When more than
one specimen is collected, the specimens should be
Handling of the Specimen clearly distinguished either by placing identifying
The specimen must be gently grasped with for- sutures in each specimen or by submitting sep-
ceps or secured with a traction suture.1 Toothed arate specimen containers.1
forceps are acceptable if care is taken and the area The specimen should be placed in 10% neu-
grasped is away from the primary site of interest tral buffered formalin with at least 20 times the
(see Fig. 4e). Suction devices, if required, should volume of the sample to avoid improper fixation
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or autolysis. Water, saline, alcohol, surface dis- and wrapped in sufficient absorbent material, such
infectant, local anesthetic solution and mouth
rinse cannot fix tissue properly and should not be
as paper towel, in case of leakage. The wrapped
container should then be placed in a sealed plastic
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used for this purpose. The fixative should not be bag, which is then placed in a rigid outer container Publié par
changed or diluted.1 that can be secured by adhesive tape. The package l’Association dentaire canadienne
For definitive diagnosis of vesiculobullous dis- should indicate “Human Tissue” or “Pathological
orders, tissue is obtained for conventional biopsy Specimen” and “Fragile” or “Handle With Care,”
examination as well as direct immunofluorescence with the complete name and address of the sender.
studies. The latter require tissue that has been
Follow-up and Reporting of Biopsy Result to
stored in Michel’s solution. The sample must be
the Patient
received by the lab within 48 hours as the solution
Patients should be seen 1 to 2 weeks postopera-
is water-based and does not preserve the tissue.
tively to ensure healing and to discuss the results
Hemostasis of the biopsy. It is the responsibility of the clinician
Small biopsy wounds may be closed with a few (not the assistant or secretary) to explain the diag-
single interrupted sutures using resorbable plain nosis and any further management if necessary.
gut. Scaffolding agents, such as absorbable gelatin If the microscopic diagnosis is inconsistent with
sponge, are useful for gingival and palatal biopsies, the clinical impression, the clinician is strongly
where the bound mucosa precludes placement of advised to discuss any concerns directly with the
sutures. A denture or preconstructed acrylic base pathologist.
or stent can be helpful to protect palatal surgical
sites. Small wounds in the floor of the mouth heal Conclusion
well without primary closure, and hemostasis may Tissue biopsy is an indispensable tool, as
be achieved with a chemical cautery agent such proper management of oral mucosal disease begins
as silver nitrate. Hemostasis may also be achieved with diagnosis. Although a wide variety of biopsy
with a laser or electrosurgery unit. techniques and devices exist, the ultimate under-
lying goal is to obtain a representative tissue
Postoperative Instructions
sample to facilitate histologic interpretation.
Standard postoperative instructions should
When in doubt, the patient should be referred to
be provided to the patient, including generic
a specialist with expertise in the diagnosis and
guidelines for eating, brushing, pain control and
management of oral diseases, such as an oral
bleeding (Appendix 1). Emergency contact infor-
pathologist or oral surgeon. a
mation should be made available.
Submission of Biopsy The Authors

The specimen should always be accompanied
by pertinent clinical information, including the
Dr. Avon is an oral medicine and oral pathology
patient’s demographic data, clinical appearance,
specialist and professor, faculty of dentistry,
location of the lesion and any relevant medical his- Laval University, Quebec City, Quebec.
tory. Including a colour photograph of the lesion
can be helpful.1 Dr. Klieb is a staff oral pathologist at the
Materials should be sent by courier to mini- Sunnybrook Health Sciences Centre, a
University of Toronto-affiliated hospital, with
mize delay in diagnosis and to prevent freezing cross appointments in the departments of dent-
artifact, which may occur if the specimen is left istry and anatomic pathology.
in a mailbox or transported without temperature Correspondence to: Dr. Hagen Klieb, Sunnybrook Health
regulation. If the specimen is to be sent through Sciences Centre, Suite H126, 2075 Bayview Avenue, Toronto,
the post, precise details of the regulations gov- Ontario, M4N 3M5. Email: hagen.klieb@sunnybrook.ca
erning pathological specimens, available from the
The authors have no declared financial interests.
post office, should be followed. Most regulations
require a primary container that is tightly sealed This article has been peer reviewed.
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screening for oral squamous cell carcinomas. Supplement to

References
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Aug/Sept 2010 isssue of Dispatch magazine.
1. Melrose RJ, Handlers JP, Kerpel S, Summerlin DJ, Tomich CJ, 5. Poh CF, Samson Ng, Berean KW, Williams PM, Rosin MP,
American Academy of Oral and Maxillifacial Pathology. The Zhang L. Biopsy and histopathologic diagnosis of oral premalig-
DES CONNAISSANCES
use of biopsy in dental practice. The position of the American nant and malignant lesions. J Can Dent Assoc. 2008;74(3):283-8.
Academy of Oral and Maxillofacial Pathology. Gen Dent. 6. Oliver RJ, Sloan P, Pemberton MN. Oral Publiébiopsies:
par methods
2007;55(5):457-61. and applications. Brit Dent J. 2004;196(6):329-33.
2. Mashberg A, Samit A. Early diagnosis of asymptomatic 7. Patton LL. The effectiveness of community-based visual
oral and oropharyngeal squamous cancers. CA Cancer J Clin. screening and utility of adjunctive diagnostic aids in the early
1995;45(6):328-51. detection of oral cancer. Oral Oncol. 2003;39(7):708-23.
3. Driemel O, Kunkel M, Hullmann M, von Eggeling F, Müller- 8. Poh CF, Zhang L, Anderson DW, Durham JS, Williams PM,
Richter U, Kosmehl H, et al. Diagnosis of oral squamous cell Priddy RW, et al. Fluorescence visualization detection of field
carcinoma and its precursor lesions. J Dtsch Dermatol Ges. alterations in tumor margins of oral cancer patients. Clin Cancer
2007;5(12):1095-100. Res. 2006;12(22):6716-22.
4. Collaborators from Royal College of Dental Surgeons of
Ontario. Evidence-based clinical recommendations regarding
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Appendix 1:
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Postbiopsy instructions for the patient Publié par

Biopsy is a minor surgical intervention, but some basic instructions are important for you to know and apply.
Bleeding: Some minor bleeding is expected. Your saliva may have a pinkish colour during the first day. If bleeding is heavy, you may
apply pressure for several minutes by placing gauze (or even a moist tea bag) over the site of surgery.
Diet: Soft food and warm or cool beverages are acceptable. Avoid biting the biopsy site. Also avoid hot foods and beverages, which
may scald the tissue.
Swelling: Soft-tissue swelling may occur and generally peaks after 48 hours. Apply a cold compress over the area if the swelling
is bothersome.
Sutures: The sutures (“stitches”) will dissolve in about 7 days. Do not attempt to remove the sutures by yourself. Occasionally,
a suture may pop off earlier than expected. This is usually not a problem, so long as there is no bleeding.
Analgesics (painkillers): Prescription pain medications are seldom needed. If you are in pain, take the same over-the-counter
medication that you would take for a headache, such as acetaminophen (Tylenol).
If you have any questions, you may contact us at: ___________________________________________________________________
If there is an emergency, you may contact emergency services at: ______________________________________________________
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ESSENTIAL DENTAL KNOWLEDGE

Oral Soft-Tissue Biopsy: An Overview

to a specialist should be considered in such cases. Biopsy Techniques

variety of clinical presentations1,2,5 (Fig. 3). Classic Punch Biopsy

nique may produce good local hemostasis and

Brush biopsy has been advocated as a screening

Consent be used with care to prevent loss of the specimen

Submission of Biopsy The Authors

screening for oral squamous cell carcinomas. Supplement to

Postbiopsy instructions for the patient Publié par

If you have any questions, you may contact us at: ___________________________________________________________________

If there is an emergency, you may contact emergency services at: ______________________________________________________

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