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Saliva serves many biological functions in the oral mucosa such as solubilizing dry food, initiating starch digestion, oral lubrication and maintaining oral hygiene. It can also be used for the detection of different biomarkers such as hormones, antibodies and enzymes

Saliva is produced by 3 salivary glands; the sublingual, submandibular and parotid glands. A fine mesh of capillaries surrounds a network of ducts from salivary glands. Blood vessels crowd around the lobed structures that make saliva and transfer salts, proteins, hormones and circulating genomic materials from the blood circulation into the secretory cells of the salivary glands

The measurement of salivary biomarkers has several advantages :

• Collection of saliva is non-invasive, convenient, and generally easily accepted by patients, including young children and the elderly
• It avoids painful blood taking and multiple and repeated sampling is more readily accepted
• Collection of saliva samples does not require a trained phlebotomist, making self-collection possible

The Salivary Biomarkers Research Group (SBRG) was formed in March 2001 to conduct research on saliva as an alternative diagnostic fluid for clinical management and bio-behavioral research

• To assess the role of salivary biomarkers in various diseases
• To explore the use of saliva as an alternative diagnostic fluid
• To establish the use of salivary biomarkers in clinical settings
• To establish the use of salivary biomarkers as an objective measurement of stress

Subjects are asked in advance not to eat or drink (except water) one hour before saliva collection to minimize possible food debris, interference by acidic food and stimulation of salivation

Subjects are asked to fill in a questionnaire to ensure test validity e.g. occurrence of upper respiratory tract infection in the past one week (which may affect markers of mucosal immunity), use of sublingual hormones (may affect cortisol profile), use of corticosteroids (may affect inflammatory markers assessment)

Saliva collection can be done either with assistance from investigators or can be self collected

Pictures depicting field conditions where saliva collection is possible; (a) during medical examination (b) at the worksite (c) sampling student cohorts (d,f) during psycho-physiological experiments (e) sampling pre-schoolers (g) sampling infants

Various collection devices (a) Tube to collect passive drool (b) Salivettes® (c) Sorbettes® (d) Oragene saliva collection kit

Several saliva collection devices are available. They vary from collection of passive drool, to collection using devices such as cotton pledgets, sterile braided cotton ropes, filter paper, or hydrocellulose microsponges

BIOMARKERS THAT WE TEST @ SBRG

Cortisol Immunoglobulin A Lysozyme Chromogranin A Alpha-amylase Eosinophil Cationic Protein Epidermal Growth Factor

Melatonin Acetylcholinesterase Cotinine Nitrate / Nitrite Testosterone Blood contamination Anti-oxidants - Total antioxidant - Catalase - Glutathione Peroxidase - Superoxide Dismutase - Ascorbic Acid - Uric Acid

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Studies with other departments in the Yong Loo Lin School of Medicine

A. Department of Epidemiology and Public Health

To study burnout and salivary biomarkers (immunoglobulin A and cortisol) among medical house-officers
To study the change in salivary physiological stress markers (cortisol and alpha-amylase) associated with the use of an automated massage chair
To study the relationship between serum, erythrocyte and saliva acetyl cholinesterase (AChE) levels as to explore the use of saliva AChE as potential biomarker for organophosphate pesticide exposure
To investigate the response of salivary alpha amylase activity to a potent “real life” stressor, inferred from passing or failing an oral examination
To examine the response of salivary alpha-amylase to an acute stressor – being subjected to a venepuncture as well on a control day

B. Department of Otolaryngology To study the impact of Stress on diurnal patterns of cortisol, symptoms and Underlying Immunologic Mechanisms of Allergic Rhinitis

C. Department of Medicine To determine the effect of hypobaric hypoxic conditions in a simulated flight at 8000 feet on stress and immune markers in healthy volunteers
To determine the extent of the effect that sleep deprivation has (versus hypobaric hypoxic) on stress and immune markers in healthy volunteers

D. Department of Anaesthesia To investigate the relationship between stress ( cortisol and alpha-amylase as outcome measurements) on learning in crisis management scenarios among medical students using a human patient simulator

E. Department of Paediatrics To determine whether salivary eosinophillic cationic protein and exhaled nitric oxide concentration levels of children with asthma and allergies are different as compared to those without asthma

Studies with other faculties in the National University of Singapore

A. Faculty of Dentistry, Department of Preventive Dentistry To study periodontal disease salivary biomarkers in adult diabetics as compared with healthy controls

B. School of Arts and Social Sciences, Department of Psychology To study the effect of physical exercise on mean diurnal salivary cortisol and cognition, depressive and anxiety symptoms in two groups of young healthy adults over a period of 3 months
To study the effect of physical exercise and stretching on mean diurnal salivary cortisol and cognition, depressive and anxiety symptoms in older healthy adults over a period of 3 months

C. School of Design and Environment Department of Building To investigate the relationship of salivary biomarkers (IgA, cortisol, α-amylase) and various indoor climatic environments To identify useful salivary biomarkers of indoor environmental quality that can be utilised for objective medical surveillance of office-based workers (Our collaborator, Dr Willem Henry Cahyahdi, was awarded the prestigious Ralph G Nevins Award for this work) To investigate the relationship between salivary biomarkers and climatic environments in childcare centres To examine effects of ozone initiated chemistry on sick building syndrome symptom intensity, perception, work performance and salivary biomarkers

D. Business School, Department of Management and Organisation To study how testosterone is associated with status and collective efficacy in a group and the effects of a mismatch between testosterone and status within a group on the collective efficacy of the group

Studies with other organisations in Singapore

A. Singapore Sports School To investigate the changes of salivary IgA and pattern of illness of swimmers in response to altitude training To investigate the immune pattern (salivary IgA) of swimmers preparing for the Beijing Olympics
B Singapore Ngee Ann Polytechnic To study the 24-hour circadian rhythm of antioxidants in saliva in a group of healthy subjects To examine the association of antioxidant levels in blood and saliva To observe the effect of physical exercise on salivary antioxidant status

Collaborative studies with overseas universities

A. The University of Hong Kong Department of Nursing Studies To evaluate the effectiveness of a proactive multi-step family smoking intervention delivered by nurses to non-smoking mothers and smoking fathers to protect the infants and mothers from second hand smoke exposure and to help the father to quit smoking by measuring salivary cotinine, a marker of tabacco smoking exposure
B. University of Malaya, Department of Social and Preventive Medicine To investigate teachers self-perceived stress and salivary IgA and cortisol among teachers in Kuala Lumpur

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Seated (L- R): Dr Cheong SK, Dr Gerald Koh, Prof David Koh, A/P Chia SE, Dr Wong TY Standing: Andrew Wee, Shin Ni (Intern), Vivian Ng, Pei En, Ivy (Intern), Shaomin (Intern)

Salivary Biomarkers Research Group Publications

Ng V, Koh D, Chan G, Ong HY, Ong CN. Stress and salivary IgA among female nurses. Proceedings of an International Expert Meeting on Women at Work. 1997, Research Report 20: 196-203

Ng V, Koh D, Chan G, Ong HY, Chia SE, Ong CN. Are salivary IgA and lysozyme biomarkers of stress among nurses? J Occup Environ Med 1999; 41 (10):920-927

Ng V, Yang Y, Koh D, Lee CY, Chan G, Fang D, Goh SH, Anantharaman V, Chia SE. Work-related stress and salivary biomarkers among female nurses in emergency department. In Book of Abstracts: 26th International Congress on Occupational Health, 27th August - 1st September 2000, Singapore

Mok BYY, Lim LP, Yang Y, Koh D. A pilot study of salivary immunoglobulin A in disabled adults. J Dent Res 2000; 79:5, 1327

Yang Y, Koh D, Ng V, Lee FCY, Chan G, Dong F, Chia SE. Salivary cortisol levels and work-related stress among emergency department nurses. J Occup Environ Med 2001; 43:1011-1018

Ng V, Koh D, Mok B, Lim LP, Yang Y, Chia SE. Stressful life events of dental students and a salivary biomarker. Int J Immunopathol and Pharmacol 2002; 15:2(S) 40

Koh D, Ng V, Chua LH, Yang Y and Chia SE. Exposure to inorganic lead and mucosal immunity. Int J Immunopathol and Pharmacol 2002; 15:2(S) 27

Ng V, Koh D, Mok B, Choo S, Lim LP. Salivary Biomarkers associated with Academic Examination Stress. In Proceedings: 17th. Asian Conference on Occupational Health, Taipei. Nov 2-4 2002

Koh D. The use of salivary biomarkers in occupational health. In Proceedings: 17th. Asian Conference on Occupational Health, Taipei. Nov 2-4 2002

Yang Y, Koh D, Ng V, Lee CY, Chan G, Dong F, Goh SH, Anantharaman V, Chia SE. Self-perceived work-related stress and the relationship with salivary IgA and lysozyme among emergency department nurses. Occup Environ Med 2002; 59:836-841

Koh D, Yang Y, Ng V, Chia SE. Stress, mucosal immunity, upper respiratory tract infections and sickness absence. J Occup Environ Med 2002; 44(11): 987-988

Koh D, Ng V, Chua LH, Yang Y, Ong HY, Chia SE. Can salivary lead be used for biological monitoring of lead exposed individuals? Occup Environ Med 2003; 60: 696-698

Ng V, Koh D, Mok B, Chia SE, Lim LP. Salivary biomarkers associated with academic assessment stress among dental undergraduates. J Dent Educ 2003; 67(10): 1091-1094

Ng V, Koh D, Chia SE. Examination stress, salivary cortisol and academic performance.Psychol Rep 2003; 93:1133-1134

Ng V, Koh D, Fu QY, Chia SE. Effects of storage time on stability of salivary immunoglobulin A and lysozyme. Clin Chim Acta 2003; 338: 131-134

Ng V, Koh D, Mok B, Lim LP, Yang Y, Chia SE. Stressful life events of dental students and salivary immunoglobulin A. Int J Immunopathol and Pharmacol 2004; 17,2(S):49-56

Koh D, Yang Y, Khoo L, Nyunt SZ, Ng V, Goh CL. Salivary immunoglobulin A and lysozyme in patients with psoriasis. Ann Acad Med S'pore 2004; 33:307-310

Ng D, Koh D, Choo S, Ng V, Fu Q. Effect of storage conditions on the extraction of PCR-quality genomic DNA from saliva. Clinica Chimica Acta 2004; 343(1-2): 191-4. (Corresponding author – D Koh)

Ng DPK, Koh D, Choo S, Chia KS. Viability of saliva as an alternative source of genomic DNA for genetic epidemiological studies. Ann Acad Med S’pore 2005; 34 (Suppl) 9 : S244.

Ng V, Koh D, Mok B, Lim LP, Chia SE. Stress, coping strategies and salivary immunoglobulin A of dental undergraduates in Singapore. J of UOEH 2006 May; 28: 160-170

Ng DPK, Koh D, Choo S, Chia KS. Saliva as a viable alternative source of human genomic DNA in genetic epidemiology. Clinica Chimica Acta 2006; 367, 81-85. 

Koh G, Shek L, Kee J, Wee A, Ng V, Lee BW, Koh D. Serum ECP correlates with asthma severity and salivary ECP correlates with co-existence of allergic rhinitis in children. Ann Acad Med Singapore 2006;35(10):S1

Koh D, Koh G. The use of salivary biomarkers in occupational and environmental medicine. Occup Environ Med 2007; 64: 202-210

Koh G, Shek L, Goh D, Van Bever H, Koh D. Eosinophil cationic protein: Is it useful in asthma? A systematic review. Respiratory Medicine 2007;101:696-705

Koh D, Koh G. The confounding effects of intra-oral metals in salivary biomarkers - Authors’ reply. Occup Environ Med 2007;64:856

Ng V, Koh D. The Use of salivary biomarkers in occupational stress Studies. Malaysia Journal of Public Health Medicine 2008; 8 (Suppl 1): 12

Ng V, Koh D, Wee A. Salivary Alpha Amylase levels under conditions of extreme examination stress. Psychol Rep 2008; 103:455-458

Wong TY, Koh D, Wee A, Ng V, Koh YT, Sum Z, Koh G. The effect of cotton-based collection methods on eosinophil cationic protein (ECP) concentrations detected in saliva. J Asthma Allergy 2008; 1:45-48

Ng V, Koh D, Wee A, Chia SE. Salivary acetylcholinerase as a biomarker for organophophate exposure. Occup Med 2009; 59:120-122

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