Journal of Dento-Medical Science and Research (Vol.

1/Jan-June 2013)

Saliva : An Alternative Biological Fluid for Clinical Applications

Chamindie Punyadeera, PhD

Saliva Translational Research Group, The University of Queensland Diamantina Institute, Translational Research Institute Pty. Ltd.
37 Kent Street, Woolloongabba, Queensland, Australia, 4102

In recent years, saliva has gained recognition as an important

diagnostic fluid just as for blood and urine. It is now evident
that saliva contains many of the same biomolecules that are
commonly measured in other body fluids. As an example,
approximately, 30% of the proteins that are found in saliva
are also found in blood1 highlighting the diagnostic potential
of saliva. Saliva tests are advantageous in comparison to
blood tests due to simple collection being noninvasive,
simple, and inexpensive, with minimal risk of contracting
infectious organisms such as HPV, HCV and HIV by the
healthcare professional. In addition saliva is an ideal biofluid
for children because of no issues with compliance.2 Saliva
tests can be done in the comfort of one's home, thus
eliminating the need for a visit to the clinic or hospital.

Saliva is produced by 3 major salivary glands

(submandibular, sublingual and parotid) and about 400 minor
salivary glands and intraoral sources such as oral mucosa
or gingiva. In general, a healthy adult produces 500-1500
ml of saliva per day, at a rate of approximately 0.5 ml/min15
but several physiological and pathological conditions can
modify saliva production quantitatively and qualitatively e.g.,
smell and taste stimulation, chewing, psychological and
hormonal status, drugs, age, hereditary influences and oral
hygiene.16 Secreted saliva can be classified into unstimulated
resting saliva or stimulated saliva (acid or mechanically)
and the salivary composition is affected based on the
stimulation type.17, 18, 19, 20

The first article relating to saliva was published in 1827

(Data source: Web of Knowledge)3 and the author observed
changes in saliva when exposed to mercury. Since this
publication, this field has grown exponentially with the
launch of a large number of successful commercial
products. In the past, primarily, dental healthcare
professionals used saliva for either research or diagnostic
purposes but now many biologists, chemists, statisticians,
bioinformaticians, engineers and clinicians are working
closely to develop novel innovative diagnostic tools using
saliva.4,5 Dental researchers in the past have used the
buffering capacity and the bacterial content of saliva to
assess a person's risk of developing tooth decay.6

Saliva is a plasma ultra filtrate and contains proteins

either synthesized in situ in the salivary glands or derived
from blood. It contains biomarkers derived from serum,
gingival crevicular fluid, and mucosal transudate. Saliva is
produced in the acinar cells and acinar cells are connected
to the vasculature which enables molecular transportation
from blood into saliva. Salivary components may originate
entirely from the salivary glands or may be derived from
the blood by passive diffusion or active transport.16 To date,
researchers have identified 2,340 proteins in saliva.21, 22 Just
like the plasma proteome, the saliva proteome has a large
dynamic range and it is important to suppress this dynamic
range in order to enable low abundant proteins of diagnostic
potential.

There is growing evidence that an individual's oral

hygiene is an important factor regulating systemic
inflammation, ultimately leading to the development of
coronary events. Today, the scientific and technological
advances in biochemistry, microbiology, and immunology
are paving the way for the discovery of new biomarkers in
saliva that can be used to detect systemic illnesses. My
group and others have utilized saliva to diagnose systemic
diseases such as ischemic heart disease and heart failure,7,8,9
renal function,10 and a large number of cancers.11,12,13,14

Early pioneers of oral diagnostics are two companies

based in the U.S. and include Epitope, Inc. and Saliva
Diagnostic Systems, Inc. They have both commercialized
saliva collection devices in the early 1990s, and in 1996 the
Food and Drug Administration (FDA) approved Epitope's
Orasure HIV test, the first test to use an oral fluid to test for
an infectious disease. More recently, FDA has approved the
first over-the-counter salivary HIV test which enables people
to test themselves in the privacy of their homes for the HIV
virus. The OraQuick HIV test, which takes only 15 minutes

Journal of Dento-Medical Science and Research (Vol.1/Jan-June 2013)

from start to finish, detects the presence of HIV antibodies

in saliva via a mouth swab.
Several companies outside the U.S. have commercial
tests to detect drugs-of-abuse in a spit sample, including
Cozart Biosciences, Securetec and Mavand. Some of these
companies send their kits via regular mail to customers,
allowing individuals to collect their own saliva either in a
cup or with a swab and then send the sample to a laboratory
for analysis. Other tests target DNA in saliva. Canada-based
DNA Genotek was the first company to commercialize a
saliva collection tool for genotyping based on polymerase
chain reaction (PCR), microarrays, and sequencing. Oral
DNA Labs, recently divested from Quest Diagnostics, also
offers two salivary tests in the U.S. in its CLIA-approved
testing facility. My PerioPath is a DNA test that determines
the risk of periodontal infections by detecting bacterial
pathogens in saliva. OraRisk HPV is a salivary test that
determines an individual's risk of developing HPV-related
oral cancers. It identifies various HPV genotypes, including
HPV 8, 11, 16 and 18. Furthermore Oasis Diagnostics
(Vancouver, U.S.) has commercialized a large number of
saliva collection devices for research and clinical
applications.

0.1-0.001 of the levels found in blood; therefore, very

sensitive detection technology is required to develop effective
tests; (e) another impediment is the lack of information about
baseline levels or reference ranges of molecules in saliva
within a healthy control population. This information is
crucial to discriminating disease-specific changes; (f) to be
clinically useful, there must be reliable correlations between
the levels of the target substance in saliva and in blood or
plasma. For example, we know that salivary diagnostics
are not well suited for measuring glucose levels because
blood and salivary levels of this analyte are poorly correlated.
This could be the case for other analytes as well.
The key parties responsible for translating salivary
research from a laboratory setting to clinical practice,
including scientists, regulatory agencies, and third parties
such as insurance companies, will need to work together to
determine how new saliva diagnostics are adopted by the
health care community. But undoubtedly, a saliva swab test
in the privacy of one's home or at the general practitioners
office will become a reality for diagnosing systemic diseases.
References
1.

Yan, W, et al. Systematic comparison of the human saliva and

plasma proteomes. Proteomics Clin Appl 2009, 3(1):
pp. 116-134.

2.

Pfaffe, T, et al. Diagnostic potential of saliva: Current state and

future applications. Clin Chem 2011, 57(5): pp. 675-87.

3.

Bostock, J. Observations on the saliva during the action of mercury

upon the system. Medicochirurgical transactions 1827, 13(Pt 1):
pp. 73-87.

4.

Choi, M. Saliva diagnostics integrate dentistry into general and

preventive health care. Int J Prosthodont 2010, 23(3): p. 189.

5.

Chiappin, S, et al. Saliva specimen: A new laboratory tool for

diagnostic and basic investigation. Clinica Chimica Acta 2007,
383(1-2): pp. 30-40.

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Lawrence, H. Salivary markers of systemic disease: Noninvasive

diagnosis of disease and monitoring of general health. J Can Dent
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Christodoulides N, FP, Miller CS, et al. Lab-on-a-chip methods

for point-of-care measurements of salivary biomarkers of
periodontitis. Ann N Y Acad Sci 2007, pp. 1098: 411-28.

8.

Punyadeera, C, et al. One-step homogeneous C-reactive protein

assay for saliva. J Immunol Methods 2011, 373(1-2): pp. 19-25.

9.

Foo, JY, et al. NT-ProBNP levels in saliva and its clinical relevance
to heart failure. PLoS One 2012, 7(10): p. e 48452.

Future Outlook
As our knowledge of the biomolecules present in saliva
grows, the potential applications for oral and systemic
disease diagnosis will expand. While the scientific link
between salivary biomarkers and oral diseases is clear, more
studies are needed to delineate the mechanisms by which
saliva reflects other systemic diseases. Furthermore, before
saliva can become widely recognized as a reliable diagnostic
fluid, we need to understand a number of important variables
that may alter delivery secretion.
The expansion of salivary diagnostics is hampered by
several factors: (a) the lack of standardized saliva collection
methods make the widespread adoption of salivary
diagnostics more challenging. In one study, researchers
reported that the OraSure saliva collection device detects
hepatitis C virus with greater sensitivity than the Salivette
device;23 (b) saliva composition is highly variable and thus
it is important to collect multiple saliva samples to reduce
inter and intra individual variability. Research should also
focus on how molecules vary diurnally. For example, we
know that salivary growth hormone levels are higher in the
morning than during the day, which could also be the case
for other biomarkers; (c) it is also important to understand
the mechanisms of biomolecular transport from blood into
saliva and the factors governing these processes and these
biomolecules may undergo modifications due to intraoral
conditions; (d) analytes in saliva are usually present at only

10. Blicharz TM, RD, Bowden M, Hayman RB, DiCesare C, Bhatia

JS, et al. Use of colorimetric test strips for monitoring the effect
of hemodialysis on salivary nitrite and uric acid in patients with
end-stage renal disease: A proof of principle. Clin Chem 2008,
54: pp. 1473- 80.

Journal of Dento-Medical Science and Research (Vol.1/Jan-June 2013)

11. Wong, DT. Oral cancer and saliva diagnostics, in National Oral
Health Conference 2007, Denver CO.
12. Ovchinnikov, DA, et al. Tumor-suppressor gene promoter
Hypermethylation in saliva of head and neck cancer patients.
Transl Oncol 2012, 5(5): pp. 321-6.
13. Streckfus, CF, et al. Breast cancer related proteins are present in
saliva and are modulated secondary to ductal carcinoma in situ of
the breast. Cancer Investigation 2008, 26(2): pp. 159-167.
14. Bigler, LR, CF Streckfus and WP Dubinsky. Salivary biomarkers
for the detection of malignant tumors that are remote from the
oral cavity. Clinics in Laboratory Medicine 2009, 29(1):
pp. 71-85.
15. Chicharro JL, LA, Perez M, Vaquero AF, Urena R. Saliva
composition and exercise. Sports Med 1998, 26: pp. 17-27.

18. Mohammed R, LCJ, Cooper-White J, Dimesky G, Punyadeera

C. The impact of saliva collection and processing methods on
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20. Navazesh, M and SK Kumar. Measuring salivary flow: Challenges
and opportunities. J Am Dent Assoc 2008, 139 Suppl:
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21. Bandhakavi, S., et al. A dynamic range compression and threedimensional peptide fractionation analysis platform expands
proteome coverage and the diagnostic potential of whole saliva.
J Proteome Res 2009, 8(12): pp. 5590-600.

16. Aps, JK and LC Martens. Review: The physiology of saliva and

transfer of drugs into saliva. Forensic Sci Int 2005, 150
(2-3): pp. 119-31.

22. Schulz, BL, J Cooper-White and CK Punyadeera. Saliva proteome

research: Current status and future outlook. Crit Rev Biotechnol,
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17. Topkas, E, et al. Evaluation of saliva collection devices for the

analysis of proteins. Clin Chim Acta 2012, 413(13-14):
pp. 1066-70.

23. JuddA, PJ, Hickman M, et al. Evaluation of a modified commercial

assay in detecting antibody to hepatitis C virus in oral fluids and
dried blood spots. J Med Virol 2003, 71: pp. 49-55.