Review

A Review of Biopsy in Dentistry: Principles, Techniques, and

Considerations

Massoumeh Zargaran

Department of Oral and Maxillofacial Pathology, Dental Research Center, Faculty of Dentistry,
Hamadan University of Medical Sciences, Hamadan, Iran

Received 22 September 2013 and Accepted 2 January 2014

Abstract Introduction
Biopsy is a useful technique for definitive diagnosis Any lesion appearing in oral cavity should be
of abnormal oral conditions and it must be performed carefully evaluated (1) while clinical diagnosis must be
carefully since any error may result in histopathologic confirmed throughout different procedures (2).
misidentification of the lesion. Present study evaluated In this context, microscopic analysis is accepted as a
the most relevant literatures published between 1982 gold standard procedure to diagnose most of lesions (3),
and 2012 in order to review existed guidelines of correct so the exact identification of their nature would be
biopsy performance using scientific database sources feasible only via histopathologic evaluation of tissue
such as PubMed, MEDLINE, IMEMR, and Chemical specimens taken from the patient by a biopsy technique
Abstract. The aim of this study was to increase (1,4). In addition, early diagnosis of some oral lesions,
knowledge of dental practitioners especially general especially various malignancies, is only possible by
dentists about factors which may influence optimization taking biopsy specimens, which are important to
of oral biopsy. improve patient prognosis (1). Nowadays, oral cancer is
considered as a major health problem with increasing
Key words: Biopsy, dentistry, histopathology, oral incidence and mortality rates in various parts of the
lesion, specimen. world. Rapid and early diagnosis of oral cancer makes it
possible to apply various conservative treatment
modalities and shorter period of care with more
favorable prognosis (5). Therefore, a delay in diagnosis
or referral might have a great effect on mortality and
morbidity associated with oral cancer (6). Dental
practitioners have an important role in this context (7).
The majority of researchers believe that proper
knowledge, ability and sufficient expertise in relation to
the recognition of symptoms and signs of malignant and
premalignant lesions in clinical examinations, carrying
out a biopsy procedure and its submission for
histopathologic evaluation are associated with early
diagnosis (7,8). In fact, more and widespread use of
biopsies by dental practitioners will most probably
decrease the number of legal actions due to belated
diagnosis or lack of diagnosis of a lesion (9). The
application of biopsies may also provide the correct
diagnosis of oral lesions and some systemic diseases
--------------------------------------------------------- which decreases patients’ problems and improve
Zargaran M. A Review of Biopsy in Dentistry: Principles, professional credibility of dentists (4).
Techniques, and Considerations. J Dent Mater Tech 2014; Considering the importance of obtaining a biopsy all
3(2): 47-54. the dental practitioners, including general dental

Zargaran JDMT, Volume 3, Number 2, June 2014 47

practitioners, should be able to take biopsies (2). Obtaining biopsies in general dental office has
However, it is not very common for dental practitioners several advantages for patients. Considering the rapport
to take biopsies due to lack of sufficient knowledge and between the patient and the dentist, the patient will have
awareness (1). Knowledge about the nature of a lesion is a better feeling. In addition, the patient will cut down on
only one step in the pathway toward a correct diagnosis the number of commutes and will spend less time
and proper management of the patient. Other steps of waiting for the biopsy to be taken and for receiving its
this pathway include the selection of the type of biopsy, report. On the other hand, fewer patients will be referred
correct implementation of the biopsy technique, proper to specially centers (14).
fixation of the specimen obtained, transport and Based on suggestions made by Oliver, biopsies
submission to the laboratory (10,11) and transfer of permissible for general dentists have been listed.
adequate and complete clinical information to the Fibroepithelial polyp, pyogenic granuloma and epulis
pathologist by general dental practitioners, which are all are lesions which can undergo biopsy techniques by
necessary to achieve the best results (11,12). general dental practitioners. In relation to mucoceles, if
The principal aim of obtaining a biopsy is to provide care is exercised, they can be biopsied. Highly
a specimen for histopathologic diagnosis of lesions (12). experienced dentists can biopsy mucosal lichen planus.
Any error during biopsy procedure may lead to However, general dental practitioners should refrain
misidentification of the lesions by pathologists (4). from taking biopsies from chronic ulcers, squamous cell
Hence, the procedure should be carried out carefully carcinomas and small and large salivary gland tumors
and correctly in order to provide an appropriate and should immediately refer the patient to a hospital.
specimen for pathological analysis (12). Regarding to Patients with leukoplakia, erythroplakia, blistering
the importance of awareness and knowledge of dental lesions, such as pemphigus, and granulomatous
practitioners especially general dentists on biopsies diseases, too, should be referred to a hospital (15).
issue (4); the aim of this study was to review indications After a decision is made to obtain a biopsy
and contraindications of biopsies, correct sampling specimen, the dentist should have the necessary skills
techniques, and correct stage, transfer and submission and expertise and the necessary instruments. In addition,
techniques of biopsy specimens to the laboratory in the dentist should be experienced in establishing a
order to prevent any errors in the diagnosis of lesions rapport with the patient, which is especially important
because arriving at a correct histopathologic when the dentist is to inform the patient of a malignant
interpretation depends on correct and precise lesion (14).
implementation of treatment procedures, promoting Indications for Biopsy
health and survival of patients. Biopsies are not confined to the diagnosis of
Definition malignant tumors; they are widely used to determine the
The process of obtaining a biopsy specimen from the nature of various lesions (2). Biopsies should be
oral mucosa is a simple surgical technique and general obtained from the three following:
dental practitioners are able to take biopsies from some - Lip and oral lesions those do not originate from
oral cavity lesions in order to establish a diagnosis local irrigative factors such as trauma or inflammation.
(12,13). Biopsy specimens are usually taken before - Lesions which persist for more than 2 week after
initiating the treatment procedures as the most accurate the exclusion of local irritants.
technique to reach a definitive diagnosis. The term - Lesions suspected of malignancies (16).
“biopsy” is a combination of two Greek words of “bios” Generally, oral lesions should be evaluated in
meaning “life” and “opsis” meaning “vision”. relation to their rapport with local irritating factors,
Technically, biopsy is defined as taking a tissue which should be eliminated after they are identified, and
specimen from a living organism in order to evaluate it the patient should be followed for 15-20 days. If the
under a microscope, with a diagnostic purpose. By using lesions persist after this period of time, biopsies and
this technique, the histologic characteristics of the lesion histopathologic evaluations are necessary (1).
are determined, it is differentiated from other lesions Osseous lesions with malignant features on X-rays
and its extent is recognized, based on which a treatment should be evaluated from histopathologic point of view.
plan can be prepared. The results of biopsy can help to It is necessary to histopathologically evaluate all the
determine course of the disease, prediction of recovery, maxillary cysts, especially keratocysts. All the osseous
recurrence or progression of lesions. In addition, the lesions associated with pain, paresthesia and other signs
results of biopsy procedures are invaluable in forensic and symptoms and rapidly expanding osseous lesions
medicine as hard evidence (1,9). should undergo a biopsy procedure. Biopsies are
Biopsies Permissible for General Dental indicated for all the oral mucosal lesions which have
Practitioners undergone significant and persistent color changes (very
white, red or pigmented) or changes in the appearance,

48 JDMT, Volume 3, Number 2, June 2014 Biopsy

such as ulcerations or proliferation, and there is a deep 2. Indirect: for deep lesions located under normal
palpable hard mass (1). Biopsies are necessary to tissues
evaluate precancerous mucosal lesions and lesions such *Biopsy technique: based on the technique
as lichen planus and leukoplakia in areas showing deployed, biopsies are classified into two groups:
atrophy and erosion (16). Identification of some  Incisional: In this biopsy technique a part of
systemic conditions, such as lupus, amyloidosis, the tissue representing the lesion is taken along with a
scleroderma and Sjögren syndrome, is possible by part of the healthy tissue (22). Incisional biopsy is used
obtaining biopsy specimens from the oral tissues for when it is not possible to remove the whole lesion, such
definitive diagnosis. It is possible to diagnose Sjögren as lichen planus or a diffuse white patch. In addition, in
syndrome by taking biopsy specimens from the minor cases in which the clinical diagnosis is unknown, the
salivary glands on the lips. In addition, biopsies are incisional biopsy is the technique of choice. In this
useful to confirm some oral lesions resulting from technique, biopsies should be taken with an adequate
certain infections such as syphilitic lesions and depth (at least 3-4 mm in depth) to include the basement
tuberculosis. Other uses of biopsies include diagnosis of membrane and the underlying connective tissue. The
blister lesions in mucocutaneous conditions with the length of the sample depends on the location of
involvement of oral mucosa, such as pemphigus sampling. However, as a general suggestion, the length
vulgaris and cicatricial pemphigoid (1). of the specimen is usually three times its width. The
Contraindications of Biopsy specimen should include some healthy tissue too, so that
Biopsy is contraindicated in patients with severe it can be used as a criterion for comparison with the
medical conditions and in individuals with systemic lesion (14). If the lesion is extensive, multiple biopsies
disorders where a biopsy procedure will deteriorate the are taken, which are placed in separate containers.
patient’s medical status or will lead to complications When these samples are submitted to the laboratory for
(16). In cases in which the lesion is situated in very deep histopathologic evaluation an outline image of the
locations or in an area with difficult access, in which lesion is drawn as the location of each sample is marked
taking a biopsy will result in complications or injury to on the image (23). In addition, several specimens are
adjacent structures, biopsy should be avoided and the taken in cases in which it is difficult to remove the
patient should be referred to a surgeon. Hemangioma is whole lesion due to its large size or complex position.
considered a contraindication for biopsy due to the risk Furthermore, this technique is useful in the diagnosis of
of massive and persistent hemorrhage (17). systemic conditions (1). Incisional biopsy is used for
In patients receiving intravenous bisphosphonates if lesions suspected of malignancy or in precancerous
possible biopsy should be avoided (18) because they are lesions. It is also used for chronic ulcers, squamous cell
at risk of bisphosphonate-related osteonecrosis of the carcinoma, leukoplakia, erythroplakia, lichen planus and
jaw (BRONJ) (19,20). The BRONJ phenomenon is a bullous lesions (14). There is controversy over the use
serious condition triggered by trauma following of incisional biopsy in malignant lesions because some
invasive dental procedures (19,21). Biopsies are not researchers believe that this procedure might lead to the
recommended in multiple neurofibromas due to the risk invasion of malignant cells via bloodstream in the
of transformation to neurosarcoma and in major salivary surgical wound created in the lesion site, increasing the
gland tumors. In such cases biopsies should be obtained risk of metastasis (24).
by a specialized surgeon in order to prevent traumas to -Excisional: Excisional biopsy refers to the
adjacent anatomic structures or spread of tumor cells, complete removal of a lesion or a suspected area (22)
which compromise patient prognosis. Biopsies are not which contains diagnostic and therapeutic aims as well
necessary in lesions with known etiologies, i.e. trauma (1). In this technique the lesion is completely removed
or inflammation, or normal anatomic variations, such as along with a margin of surrounding normal tissue and
physiologic pigmentation of the gingiva, geographic deep healthy tissue to ensure safety (25). This technique
tongue, linea alba, extostoses and tongue indentation is used for small lesions (with a diameter of less than 1
(1). cm), which are probably benign. The majority of
Different Types of Biopsy biopsies in the general dental field are excisional. The
Biopsies are classified based on lesion technique is used for small benign lesions, including
characteristics, the technique and the materials used, fibroepithelial polyps, papilomas, fibromas, mucoceles,
duration and location of the lesion, preparation of the hyperplastic tissues and pyogenic granulomas (14). In
lesion and the aim of the biopsy procedure. biopsies of some tumors such as hemangiomas and
*Lesion characteristics: Biopsies are classified into melanomas the lesion should be completely and
two types based on lesion characteristics (1): extensively excised to prevent severe hemorrhage and
1. Direct: for superficial lesions with easy access metastatic spread of the tumor (23).

Zargaran JDMT, Volume 3, Number 2, June 2014 49

 *Biopsy instruments: A number of cutting mucosal tissues and generally it is used for differential
instruments, including scalpel blades, punches and B diagnosis, evaluation of treatment outcomes or the
forceps, are used to take biopsies. Taking a biopsy from lesion progression follow up (1).
the oral mucosa with a punch is a rapid, easy, safe and Processing of Biopsy Specimens
cheap technique and can be used in the majority of oral Depending on the type of preparation specimens
cavity areas. Cylindrical samples measuring 2-8 mm in might be frozen or placed in paraffin or methacrylate.
diameter can be obtained using this instrument. Small The specimens can also be evaluated without
wounds created by a punch do not require suture preparation in a fresh manner. Freeze specimens are
whereas big wounds do (1). A punch can be used to take frozen by dry ice and processed without fixation.
several biopsies from different locations in one session; Freezing at -40 to -60°C makes the tissue specimen firm
it inflicts less stress and anxiety on the patient compared and it can be cut using a microtome. Placement of the
to a scalpel blade (26). It should be pointed out that specimens in paraffin is the most commonly used
taking biopsies with a punch is confined to superficial technique (1) whereas the sample is fixed in 10%
epidermal and mesenchymal lesions and has no neutral buffered formalin (22), processed and embedded
application for taking biopsies from deep lesions. In in paraffin. Finally, thin sections are cut, stained and
addition, it is not advisable to use it in areas with rich evaluated under a light microscope (1).
vascular or nervous plexuses. It is difficult to take Hard tissue specimens are placed in methacrylate
biopsies from the soft palate, maxillary tuberosity and and evaluated by an electron microscope, which helps
the oral cavity floor with a punch due to the mobility of differentiate normal cells from tumor cells based on
these areas (27). The majority of oral pathologists morphologic characteristics (1). In evaluating fresh
believe that incision with a scalpel blade results in less samples, the tissue specimen is immediately transferred
injuries (28). In addition, lasers and electrocautery to the laboratory and evaluated by immunofluorescence
should not be used to take biopsies (29) because they techniques. The specimens are moistened with saline
produce histologic artifacts due to the heat produced solution and are prevented from drying. The specimens
during the use of these instruments (30,31). should be kept at 2-8°C and care should be taken to
The Time to Take Biopsies keep them from freezing during the interval from being
The samples can be taken and processed during taken to being transferred to the laboratory (1).
surgery by freezing (intraoperative biopsy) or in a Evaluation of fresh specimens by immunofluorescence
separate session by fixation (extraoperative biopsy). An is useful for diagnosing mucocutaneous conditions with
intraoperative biopsy makes it possible to reach a rapid an autoimmune origin such as pemphigus, pemphigoid,
histopathologic diagnosis but the obtained sections have erythema multiform and lichen planus (1).
a no ideal quality with troublesome microscopic The Aim of Taking Biopsies
evaluation (1). The pathologist can only report a Biopsies are taken for diagnostic, therapeutic or
positive, negative or suspected result. However, it has research purposes. Research biopsies are usually taken
the advantage of a report in a short time by the to study new medical conditions and are carried out on
pathologist, which can be sent to the operating room. laboratory animals and in some rare cases on man (1).
Therefore, the surgeon can make the necessary decision. General Principles of Biopsies
This technique is used in cases which are suspected of There are some similar general principles for
malignancy but it is not always reliable and the surgical biopsies regardless of the type of biopsy (14). It has
operation should be postponed, when there is doubt, been recommended that before taking any biopsies
until the definitive diagnosis is provided using written consent be taken from all the patients, with the
evaluation of the samples processed through extra- reason for prescribing a biopsy procedure and details of
operative technique. An important use of intra-operative possible risks being mentioned in the consent form. The
biopsy is the margins of malignant tumors assessment risks are mainly related to the location of surgery. For
during surgery so more extensive incisions can be made example, paresthesia may occur in the tongue and lips
if the lesion is extra-widespread (1). (15). The biopsy should be obtained from the most
Sampling and processing in a separate session is a prominent part of the lesion and in cases in which the
time-consuming method. However, this technique yields lesion has different clinical views in different areas,
sections with a higher quality and easier histopathologic several biopsies should be taken. Necrotic tissues and
evaluations compared to intraoperative biopsy (1). lacerated tissues should not be biopsied because such
Location of Biopsy specimens yield little diagnostic data (14). The
Based on the area involved, specimens can be taken instruments needed for biopsy are usually the same as
from different oral cavity locations, including salivary those used in surgery, including mirrors, various
glands, lymph nodes and other head and neck tissues. It forceps, surgical blades, syringes for anesthesia etc (Fig.
is easy to obtain a biopsy specimen from the oral cavity 1). (1). Similar to any other surgical procedure, it is

50 JDMT, Volume 3, Number 2, June 2014 Biopsy

absolutely necessary to observe the principles of
disinfection for instruments and the biopsy location and
sterile gauze pieces and gloves should be used. After
making sure that all the instruments needed are ready
and available, an anesthetic agent is administered for
anesthesia. Anesthetic agents containing
vasoconstrictors are used to decrease hemorrhage.
Injections should be made at a distance of 3-4 mm from
the area under question (1) in order to prevent damage
to the specimens (14). It is advisable to use elliptical
incisions in order to take tissue specimens (15). Wounds
created by such incisions, which have been properly
designed, are easily sutured and closed (32). In addition,
care should be exercised to place incisions away from Figure 2. Artificial suprabasal cleft and acantholytic
blood vessels and major nerves as far as possible and it epithelial cells which mimic pemphigus (sample fixed
is better to place them parallel to blood vessels and in saline solution)
nerves to minimize possible injuries (15). During
removal of a tissue sample, it can be stabilized by a
suture or forceps, if necessary (15). Of course, care
should be taken not to tear or compress the biopsy
specimen (1). If several tissue specimens are produced The biopsy specimen should be placed in a sufficient
during sampling, all the segments should be sent to the amount of the fixating agent; at least 10 times of tissue
laboratory. No selections should be made in order to specimen’s volume (14,15,22). Each biopsy specimen
send only the selected segments to the laboratory for should be sent in a separate container and if more than
histopathologic evaluation (22). one specimen is placed in one container, each one
The biopsy specimen should immediately be placed should be clearly marked (for example by using a
in a fixating solution (14,22) because tissue autolysis suture) (Fig. 3). Explaining and describing the shape of
takes place rapidly after resection and taking the each specimen is not a reliable technique because the
specimen due to the disruption of blood supply to the shape of the specimen will probably change after fixing
tissue specimen (29). The best and most commonly used (15). The container which the specimen is placed in
fixative is 10% neutral buffered formalin (22). Other should have a wide opening. The specimen becomes
solutions such as water, saline solution and alcohol are hard and rigid after being fixed; therefore, it might be
not suitable alternatives for formalin, as they cause difficult to retrieve it from a container with a narrow
severe and destructive changes in epithelial structures opening (34), resulting in damage to the specimen (29).
etc (Fig. 2) (33). Plastic containers with screw caps are recommended for
the transfer of biopsy specimens (29). Glass containers
may break and injure operators. In addition, glass
fragments might penetrate into the tissue specimen and
damage the microtome during preparation of tissue
sections. (Fig. 4) (34). Data such as the first name and
family name of the patient (15), the clinician’s name
and the area which underwent the biopsy procedure
should be written on a label on the container (15,29).
The laboratory might not accept specimens which do
not have any labels and return them to the clinician for
solving the problem (34).

Figure 1. Some of instruments needed for biopsy

Zargaran JDMT, Volume 3, Number 2, June 2014 51

 immunohistochemical staining techniques (34). The
evaluation of lesion radiographic images is usually
useful for correct and proper classification of the disease
and its diagnosis (9).
Complications of Oral Cavity Biopsies
In the most cases the incidence of complications are
minimized by applying meticulous surgical techniques,
although hemorrhage is possible to occur during the first
24 hours after sampling due to clot disruption or
secondary to loosening of the suture. An uncommon
problem in biopsies is wound dehiscence, which usually
occurs 5-8 days after sampling; such a complication is
attributed to infection or tight sutures, the latter of
Figure 3. Samples of tongue (left – labeled with suture)
which leads to local obstruction of blood vessels.
and buccal mucosa (right)
Another complication is paresthesia which might persist
for several hours or months (1,25).
The majority of patients do not have severe pain
after a biopsy procedure and in cases in which pain
relief is necessary; analgesics bring about proper pain
relief as the majority of patients experience pain relief
after 3 days. The patients who have gone through biopsy
procedure need to follow standard postoperative oral
surgery instructions (15).

Conclusion
Considering the importance of biopsy for definitive
diagnosis and management of oral lesions, dentists
Figure 4. Appropriate (left) and inappropriate (right) should be able to take biopsies while they must be
biopsy containers aware of variety of factors influencing histopathologic
interpretation of an oral biopsy specimen to prevent
misidentification of the lesions .This review article is to
increase knowledge of dental practitioners about
Data Sent to the Pathologist principles, techniques and considerations of biopsy.
The biopsy specimen should be accompanied with a
complete patient history and particulars (name, age,
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Corresponding Author:
Massoumeh Zargaran
Faculty of Dentistry
Hamadan University of Medical Sciences, Hamadan, Iran
Tel: +988118381063
Fax: +988118381085
E-mail: mm.zargaran@yahoo.com & massoumehzargaran@gmail.com

54 JDMT, Volume 3, Number 2, June 2014 Biopsy