Chairside Investigations in

Oral Cancer & Oral

Potentially Malignant
By Aditya Jain Disorders (OPMD)
Final Year BDS
Roll No. 2

Under the guidance of Dr Sunita Gupta Ma’am & Dr Sujoy Ghosh Sir
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What we’ll learn

1.Overview
2.Oral cancer and OPMDs
3.Role of Chairside Investigations in Early
Detection of Oral Cancer
4.Chairside Diagnostic Aids:
- Vital Staining
- Cytopathologic Studies
2
- Light Based Detection Systems
OVERVIEW Despite advances in diagnosis and treatment,
survival rates remain stagnant at
approximately 50%, mainly due to late-stage
detection (Madhura et al., 2020).

It is the 3rd most common cancer, affecting

20 per 100,000 people and accounting for
nearly 30% of all cancers worldwide (Priya et
al., 2020)

The graph highlights the higher incidence (9.8) and

mortality (6.8) rates of oral cancer in males compared
to females (incidence: 5.2, mortality: 3.6), 3
emphasizing the need for early detection and
prevention efforts (Aggarwal, 2015).
Oral cancer and
OPMDs
• According to Burket’s, OSCC(oral squamous cell carcinoma) and
OPC(oropharyngeal cancer)represent the sixth most common
cancer worldwide.
• Causes of Oral Cancer:
1.The various causes of Oral Cancer are as follows
2.Using tobacco or smokeless tobacco
3.Drinking alcohol
4.Human papillomavirus infections
5.Sun exposure
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 Oral potentially
malignant
Disorders
The WHO listed several oral conditions as having the potential to
transform into oral cancer :
• Lichen planus
• Leukoplakia
• Discoid lupus erythematosus
• Inherited disorders
• Tobacco related lesions
• Erythroplakia
• Actinic cheilitis
• Oral submucous fibrosis 5

Reference: Burket’s Oral Medicine Textbook(13th

Oral cancer is often preceded by precursor lesions
known as oral potentially malignant disorders
(OPMDs), which, if detected early, can significantly
improve patient outcomes and enhance productive
longevity (Walsh et al., 2021).

Investigations of a patient with an oral lesion include:

Chairside Investigations

Radiographic Investigations

Laboratory Investigations

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Chairside
Investigations
• Diagnosing mouth cancer involves a biopsy, where a tissue
sample is surgically removed and examined in a lab, causing
pain and a delay in results.
• According to Sadawarte et al., chairside tests are simpler,
cost-effective, and faster but less precise, often used for
initial screening (2022).
• Clinical examination is the standard method for detecting
oral squamous cell carcinoma, with biopsy and
histopathology as the gold standard for confirmation.
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Chairside
Investigations

Vital Staining Cytopathology Light- Based

• Oral Systems
• Toluidine Blue
exfoliative
• Methylene • Chemiluminescen
cytology
Blue ce
• Brush biopsy
• Lugol’s Iodine (ViziLite)
• FNAC(Fine
• Rose Bengal • VELscope
needle
aspiration
cytology)
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 Vital Staining
Simple, economical, efficient and
relatively sensitive chair-side
diagnostic
technique.
• Enhances the characteristics of the
lesion.
• Helps identify satellite lesions and
clinically non-apparent lesion sites.
• Aids in the selection of the biopsy
site.

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 Toluidine
Blue
Composition:
As described by Mashberg et al,a 100 ml of 1% toluidine blue solution was prepared by
mixing 1 g toluidine blue powder, 10 ml 1% acetic acid, 4.19 ml absolute alcohol and 86 ml
distilled water with pH regulated at 4.5.

Chemistry and Principle:

• Acidophilic metachromatic dye, part of the thiazine group.
• High affinity for acidic compounds.
• Stains cells/tissues blue with greater nucleic acid content.

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 Toluidine
Blue
Interpretation:
• Positive: Dark blue/royal blue.
• Uncertain: Pale royal blue.
• Negative: Absence of color uptake.
• Digital color analysis: Helpful for accurate interpretation.

Applications:
• Screening for oral precancerous lesions and oral cancer.
• Post-treatment surveillance of oral potentially malignant disorders (OPMDs, oral precancer).

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 Dark royal blue TB stain(Ventral surface of
tongue suspected of a white lesion)

Before TB stain After TB stain

Accessed from “Toluidine blue uptake in
potentially malignant oral lesions in vivo: 12
Clinical and histological assessment” (Gandolfo
 Toluidine
Blue
Advantages :
• Inexpensive and simple to perform
• Helpful to evaluate free surgical margins

Disadvantages :
• Both false positive and false negative results can be seen
• Filiform papillae in the tongue retain the dye (False positive)
• Invaded underlying tissues do not take up the stain (False negative)

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 Lugol’s Iodine
Composition

Original Lugol’s solution consists of

• 5 g iodine (I2)
• 10 g potassium iodide (KI)
• 85 ml distilled water

Principle and Mechanism:

• Iodine reacts with glycogen in the cytoplasm, leading to a visible color change.
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• Malignant cells do not exhibit this iodine-starch reaction due to increased glycolysis and loss of
cellular differentiation.
Interpretation:
• Normal mucosa: Stains brown or mahogany.
• Dysplastic and cancerous tissues: Appear pale as they fail to absorb the stain.

(White arrow) shows

black to brown colour
areas after staining
indicating normal mucosa

The unstained
lesion(black arrow)
showed no uptake or pale
yellow discolouration

Accessed from “Clinical relevance of the utilization

of vital Lugol’s iodine staining in detection and 15
diagnosis of oral cancer and dysplasia” (Elimairi et
al., 2015).
 Lugol’s
iodine
Advantages :
• Inexpensive and simple to perform
• Widely available and less time consuming

Disadvantages :
• It is an irritant that damages normal epithelial cells
• Allergic reaction to iodine 16
Methylene Blue
Chemical Properties:
• Heterocyclic aromatic compound.
• Appears as a solid, odorless, dark-green powder at room temperature.
• Forms a blue solution when dissolved in water.

Mechanism:
• The uptake mechanism in epithelial tissue may resemble that of toluidine
blue.
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• Toluidine blue is hazardous if swallowed, unlike methylene blue.
Methylene Blue

Interpretation:

• The pattern of dye retention is assessed by intensity of stain retention on the

lesion
• Local,stippled,patchy,and deep blue stains are marked as positive
• Wide,shallow or faint blue stains are marked as negative

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 Accessed
from
“Methylene
Blue as a
Diagnostic Aid
in the Early
Detection of
Potentially
Malignant and
Malignant
Lesions of
Oral Mucosa ”
(Lejoy et al.,
2016).

1a. Clinical photograph 1b. Clinical photograph 1c. Photo micrograph

showing non showing dark uptake of showing moderate
homogeneous leukoplakia stain after application of dysplasia(10x) 19
on right buccal mucosa methylene blue dye
 Accessed from “Methylene blue as an early diagnostic marker
for oral precancer and cancer
” (Riaz et al., 2013).

Pre staining clinical Clinical photograph

photograph showing a showing a dark uptake of
white lesion on right methylene blue on right
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buccal mucosa buccal mucosa
 Methylene Blue

Comparison to Toluidine Blue:

• Less toxic to the human body.

• Cheaper and suitable for in vivo staining.
• Similar physiochemical structure to toluidine blue.

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Methylene blue
Advantages :
• Substitute for toluidine blue because of even more low cost
• Less toxic to human body

Disadvantages :
• Histopathology report still remains the gold standard for
confirmation of diagnosis

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Rose Bengal
• Rose bengal (RB), the 4,5,6,7-tetrachloro-20,40,50,70-tetraiodo-
derivative of fluorescein, has been widely used to diagnose various
ocular surface disorders
• It has been believed to stain desquamated ocular epithelial cells, dead
or degenerated cells but not healthy epithelial cells, or wherever there is
poor protection of the surface epithelium by the preocular tear film
rather than lack of cell vitality.(Fei Du et al 2007)

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Rose Bengal
Procedure
• The patients are asked to rinse their mouth with distilled water to clean the
lesions for 1 minute;
• 1% Rose Bengal solution is applied with a cotton tip for 2 minutes
• Patients are asked to rinse their mouth for one minute with distilled water to
remove excess RB solution
Interpretations
• Staining results are classified according to shade tabs
• Staining results of 3,4 are regarded as positive while results of 1,2 are regarded
as negative.
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 1.Mild staining with rose bengal of
homogeneous leukoplakia

1 2 2.Negative staining with rose bengal of

homogenous leukoplakia
3.Positive intense staining with rose
bengal of non homogeneous
3 4 leukoplakia
4.Positive very intense staining with
rose Bengal of Squamous cell
carcinoma

Accessed from “Rose Bengal staining –

diagnostic aid for potentially
malignant and malignant disorders: A pilot 25
study
” (Nitasha et al., 2013).
Oral Exfoliative Cytology

Overview:
• Introduced by George Papanicolaou in 1942 for early diagnosis of cancer and
precancerous lesions.
• The main concept behind cytological studies was to study the cells, which are exfoliating
due to pathologic or physiologic process of the body.

• Indications:
• Oral lesions that cannot be identified clinically with certainty.
• Lesions suspected to be benign.
• To diagnose carcinoma in situ or early-stage premalignant lesions (dysplasia).
• Lesions strongly suggestive of cancer based on clinical evaluation.

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Oral Exfoliative Cytology

Procedure:
• Remove debris from the lesion and scrape with a tongue blade.
• Smear collected cells evenly on a labeled glass slide.
• Fix the slide by immersion in alcohol/ether or spraying with commercial fixative.
• Send to a pathologist for evaluation.

Evaluation Results
• Negative for cancer.
• Suspicious for cancer.
• Positive for cancer.

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A.Sample taken from buccal B.Sample fixed with a spray
mucosa citofixer

Accessed from “Exfoliative cytology

of oral epithelial cells from patients
with type 2 diabetes:
cytomorphometric analysis.
” (Rivera et al., 2013). 28
Oral Exfoliative Cytology

Advantages:
• Painless, bloodless, non-invasive, quick, and
• economical.
• Suitable for patients contraindicated for biopsy.
• Useful for mass screening.
Disadvantages:
• Provides less information than histological slides.
• Positive results are reliable, but negative results may not be.
• Suitable only for epithelial cells.
• Supplementary tool, not a substitute for biopsy.
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Brush Biopsy

OralCDx® brush biopsy is a transepithelial system using computer-

assisted brushing to collect samples from mucosal lesions,
representing the superficial, intermediate, and basal epithelial layers.

Indications:
• Detects asymptomatic, precancerous red and white
dysplasias, chronic ulcers.
• Not suitable for fibromas, mucoceles, hemangiomas, or
pigmented lesions.
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Procedure:
• A specially designed brush collects epithelial cells by rotating 5-15 times on the
Brush biopsy
lesion with firm pressure.
• The collected tissue is placed on a dry slide and fixed with ethanol or another
fixative.
• Visualised under light microscope

Results:
• Positive: Suspicious for carcinoma or epithelial dysplasia.
• Atypical: Highly suspicious cellular morphology.
• Negative: No abnormalities detected.

An atypical or positive result would then be subjected to biopsy for a

definitive diagnosis
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Brush biopsy

Smear obtained from a potentially malignant disorder using a

cytobrush
Accessed from “ Evaluation of the Accuracy of Liquid-Based Oral Brush 32
Cytology in Screening for Oral Squamous Cell Carcinoma” (Deuerling et al.,
 Brush biopsy
Advantages:

• Simple, rapid, non-aggressive, and relatively painless.

Disadvantages:
• Cannot replace surgical biopsy.
• Limited value for detecting mucosal changes not visible to the naked
eye.

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Fine Needle Aspiration
Cytology

Overview:
• A technique for obtaining cells from a lesion using a thin bore needle.
• Smears are made for cytopathological diagnosis.

Principle:
• Based on the fact that tumor cells are less cohesive and can be easily aspirated.

Procedure:
• Material is obtained from a body cavity, cystic space, or fluid-containing lesion.
• Needles used: 18, 21, 23, or 25 gauge.

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 . (a) A metastatic small cell carcinoma exhibiting a crush artifact
and apoptosis of individual cells. (b) A metastatic keratinizing
squamous cell carcinoma. Nuclei are hyperchromatic (c) Syncytial
tissue fragments of adenocarcinoma cells. (d) Metastatic papillary
thyroid carcinoma showing intranuclear cytoplasmic inclusions
(arrows).
(a–d): Papanicolaou stain, 400× magnification.

35
Accesses from “improving detection
of precancerous and cancerous oral
lesions. Computer-assisted analysis
of the oral brush biopsy”. U.S.
Collaborative OralCDx Study Group.
Sciubba et al 1999

36
Chemiluminiscence
• CHEMILUMINESCENCE (VIZILITE), a diagnostic tool based on
the principle of chemiluminescence.

Principle
• The theory behind this technique is that the acetic acid removes
the glycoprotein barrier and slightly desiccates the oral mucosa;
the abnormal cells of the mucosa then absorb and reflect the
white/blue light in a different way with the respect to normal
epithelial cells.
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Vizilite Kit

• The kit contains 1% acetic acid solution (removes glycoprotein

barrier and slightly dessicates the mucosa)
• a capsule with an outer shell of flexible plastic
• an inner vial of fragile glass
• a retractor.

PROCEDURE:
• The patient must have a 1% acetic acid solution oral wash
• Followed by direct visual examination of the oral cavity using a blue-white light source.

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Chemiluminiscence
Activation of capsule
• Activation needs breakage of the glass vial by bending the capsule. This allow
the chemical products to react with it and produce a bluish- white light with a
wave length of 430-580 nm that lasts for around 10 min.

Interpretation
• Normal mucosa appears blue
• Abnormal mucosal areas reflect the light (due to more nuclear/ cytoplasmic
ratio in epithelial cells) and appear a more acetowhite with brighter, sharper
and more distinct margins

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 Buccal mucosa seen in incandescent After staining with acetic acid
light

Accessed from “Role of

Chemiluminescence examination as
non-invasive diagnostic
tool in early detection of
Leukoplakia”
Jain et al 2017

40
Aceto white borders delineated on
Indications
» It is used to diagnose leukoplakia and radiation mucositis.
» Identification of asymptomatic and clinically non-evident lesions.
» Diagnostic aid for the detection of oral cancer and premalignant
early lesions.

Advantages
» Vizilite has the advantage in that it is capable of delineating the
sharp borders between normal and abnormal oral mucosa and
often extended beyond the clinically identified outline.
» Malignant lesions could be recognized without use of any
adjunctive diagnostic tools.

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Velscope

• Visual enhanced light scope

• Based on Fluorescence Visualization Technology

Principle
• A specific wavelength of blue light, transmitted through a halide lamp is used, to excite tissue from the
epithelial surface, down through the basement membrane, stopping at the stroma.
• The lighted tissue, in turn, emanates a green fluorescence (sometimes referred to as autofluorescence).
• The emitted fluorescence is not visible to the naked eye, but the VELscope® hand piece filters out the
blue light, so that only the green fluorescence remains.
• Differences in the degree of green reveal possible abnormalities. 42
Workin
g

43
 Brush
Interpretation :
biopsy
• Under intense blue excitation light, normal oral mucosa emits a pale
green auto fluorescence
• Abnormal or suspicious tissue exhibits decreased levels of normal auto
fluorescence and appears dark by comparison to the surrounding
healthy tissue.

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 Lesion present on dorsum of Same lesion examined by
tongue VELscope

Accessed from “Early Diagnosis on Oral and Potentially Oral Malignant

Lesions: A Systematic Review on the VELscope®Fluorescence Method” 45
Cicciu et al 2019
 Crisply defined homogeneous Same lesion examined with VELscope
white plaque of lower labial demonstrating retained
mucosa and lip vermilion autoflourescence

Accessed from “ The role of Velscope in routine screening for

46
potentially malignant oral mucosal lesions” (McNamara et al
2012).
Indications:
• Soft tissue examination in diagnosis of suspicious oral lesions
Brush biopsy
• Determining surgical borders

Advantages:

• Takes only 1-2 mins and is painles and non invasive,with no stains or rinses required
• Determining surgical borders.
• Covers large surface of the oral cavity.

Disadvantages:
• Prolonged examination may cause discomfort
• Costly equipment

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References
• Burket’s oral medicine textbook(thirteenth edition)
• Cicciù, M., Cervino, G., Fiorillo, L., D’Amico, C., Oteri, G., Troiano, G., Zhurakivska, K., Lo Muzio, L., Herford, A. S., Crimi, S., Bianchi, A., Di
Stasio, D., Rullo, R., Laino, G., & Laino, L. (2019). Early diagnosis on oral and potentially oral malignant lesions: A systematic review on the
VELscope® fluorescence method. Dentistry Journal, 7(3), 93. https://doi.org/10.3390/dj7030093
• Jain, N., Nagarajappa, A. K., Bhasin, M., Pandya, D., & Tripathi, K. P. (2018). Role of chemiluminescence examination as non-invasive
diagnostic tool in early detection of Leukoplakia. Journal of Oral Biology and Craniofacial Research, 8(3), 177–181.
https://doi.org/10.1016/j.jobcr.2017.04.004
• SCIUBBA, J. J. (1999). Improving detection of precancerous and cancerous oral lesions. The Journal of the American Dental Association,
130(10), 1445–1457. https://doi.org/10.14219/jada.archive.1999.0055
• Akshatha, B., Shashikala, R., Indira, A., Manjunath, G., & rao, Ka. (2015). Role of micronucleus in oral exfoliative cytology. Journal of
Pharmacy And Bioallied Sciences, 7(6), 409. https://doi.org/10.4103/0975-7406.163472
• McNamara, K. K., Martin, B. D., Evans, E. W., & Kalmar, J. R. (2012). The role of Direct Visual Fluorescent Examination (VELscope) in routine
screening for potentially malignant oral mucosal lesions. Oral Surgery, Oral Medicine, Oral Pathology and Oral Radiology, 114(5), 636–643.
https://doi.org/10.1016/j.oooo.2012.07.484
• Abraham, L., Rai, A., Burde, K., & Naikmasur, V. (2016). Methylene blue as a diagnostic aid in the early detection of potentially malignant
and malignant lesions of oral mucosa. Ethiopian Journal of Health Sciences, 26(3), 201. https://doi.org/10.4314/ejhs.v26i3.2
• Seetharam, S., & Ramachandran, C. (1998). Fine needle aspiration cytology as a diagnostic test for oral squamous cell carcinoma. Oral
Diseases, 4(3), 180–186. https://doi.org/10.1111/j.1601-0825.1998.tb00276.x
• Petruzzi, M., Lucchese, A., Baldoni, E., Grassi, F. R., & Serpico, R. (2010). Use of lugol’s iodine in oral cancer diagnosis: An overview. Oral
Oncology, 46(11), 811–813. https://doi.org/10.1016/j.oraloncology.2010.07.013
• Strome, A., Kossatz, S., Zanoni, D. K., Rajadhyaksha, M., Patel, S., & Reiner, T. (2018). Current practice and emerging molecular imaging
technologies in oral cancer screening. Molecular Imaging, 17. https://doi.org/10.1177/1536012118808644
• Du, G., Li, C., Chen, H., Chen, X., Xiao, Q., Cao, Z., Shang, S., & Cai, X. (2007). Rose Bengal staining in detection of oral precancerous and
malignant lesions with colorimetric evaluation: A pilot study. International Journal of Cancer, 120(9), 1958–1963.
https://doi.org/10.1002/ijc.22467
• Ford, P. J., & Farah, C. S. (2013). Early detection and diagnosis of oral cancer: Strategies for improvement. Journal of Cancer Policy, 1(1–2).
https://doi.org/10.1016/j.jcpo.2013.04.002
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• Gandolfo, S., Pentenero, M., Broccoletti, R., Pagano, M., Carrozzo, M., & Scully, C. (2006). Toluidine blue uptake in potentially malignant
oral lesions in vivo: Clinical and histological assessment. Oral Oncology, 42(1), 88–94. https://doi.org/10.1016/j.oraloncology.2005.06.016
Thank You

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