The Experimental and Basic Sciences core of SERI comprises of all the research laboratory space and core equipment contained therein that facilitate and support the translational and basic ocular disease research within SERI. This research platform provides the core facilities available for all SERI researchers engaged in wet or bench laboratory research. The Experimental & Basic Sciences Research Platform is further divided into three separate facilities.
1. Experimental Microscopy, Molecular & Cell Biology
2. Proteomics
Head: Assoc Prof Wang Xiaomeng
3. Translational Ophthalmic Pathology & Immunology
(1) Experimental Microscopy, Molecular & Cell Biology includes all the tissue culture and -80 degree freezer rooms, laboratories dedicated to electrophysiology, chemistry and microbiology, as well as 33 general lab benches. Both specialised cell biology and general core lab equipment are part of this facility. This platform is supported by two laboratory managers who oversee daily laboratory operations, procure equipment, monitor utilisation of space, update risk assessments and ensure that biological and general workplace safety are in compliance with the rules and regulations of the SingHealth Institutional Biosafety Committee (SHSIBC).
(2) The Proteomics Facility, housed within a micro-analytical laboratory, is equipped with 2 HPLCs (nano- and high-flow) and 2 mass spectrometers (high-resolution TripleTOF and triple quadrupole) for proteomics, metabolomics and drug analyses. This facility provides 25 types of services, including gel protein identification, affinity purification of complex samples, relative/absolute quantitative proteomic analysis, identification/quantification of post-translational modifications, biomarker discovery/ validation, untargeted/targeted metabolomics, and pharmacokinetics/drug quantitation. New proteomics/metabolomics technologies to access the frontier “Omics” technologies are being developed.
(3) The Translational Ophthalmic Pathology & Immunology Facility works in synergy with the Experimental Microscopy and Imaging Facility to provide specialised non-routine histopathology services involving ophthalmic tissues. This facility is led by a trained ophthalmic pathologist, Dr Anita Chan. Non-routine histological services include but are not limited to specialised techniques for tissue processing for transmission electron microscopy, multiplexed immunohistochemistry and preparation of ocular microvasculature and retinal tissue whole mounts.
The Translational Ophthalmic Pathology & Immunology as well as The Experimental Microscopy and Imaging Facilities interact with the Department of Anatomical Pathology of the Singapore General Hospital and the Advanced Bio-imaging Core. They provide SERI researchers with access to a variety of advanced imaging equipment, as well as specialized services that help keep them abreast with advancing technology in this area.
Overview
The Proteomics Laboratory is a core platform in SERI and provides expertise and instrumentation in cutting-edge proteomics and metabolomics research. Established in 2004, the Ocular Proteomics Laboratory focuses on the application of proteomics to clinical samples from the eye to find new biomarkers and understand the disease. Additionally, we collaborate with other research groups within SERI and SNEC. We offer a broad range of proteomics/metabolomics and biological mass spectrometry (MS) services, including protein identification, quantitative proteomics (iTRAQ, high resolution – MRM, SWATH, etc.), characterisation of post-translational modifications (PTMs), MS-based non-targeted and targeted metabolomics and drug analysis. Recently, we have been asked to participate in the Human Proteome Organisation (HUPO) Eye Proteome project, which aims to establish a human eye proteome database.
Facilities
AB SCIEX TripleTOF 5600 Mass Spectrometer
Dionex UltiMate 3000 RSLCnano system
AB SCIEX API 2000 MS/MS system
Waters 2695 HPLC system
Waters Acquity UPLC I-Class system
Capabilities
Protein identification (identify proteins from a gel band, or whole proteome)
Quantitative proteomics and proteomic profiling (iTRAQ, mTRAQ, high-resolution MRM, SWATH, etc.)
Post-translational modification discovery (glycosylation, phosphorylation, acetylation, methylation, etc.)
MS-based metabolomics
Small molecule quantitation (drug level, targeted metabolomics, etc.)
Projects
One of our focuses is on tear fluid proteomics and metabolomics. We believe that tear fluid is a useful source for discovering biomarkers associated with the various components of the lacrimal functional unit, because of the close relationship of tears to the disease sites. Recent studies by our group showed that more than 1500 proteins and 60 small molecule metabolites were identified in healthy human tears using the latest proteomics and metabolomics technology. This comprehensive tear composition repertoire can be served as a background for future biomarker research of ocular diseases.
The current biomarker research projects which use tears as the source are:
Validation study of tear biomarkers for dry eye
Epidemiologic studies of tear proteomics in normal Chinese, Malay and Indian population
Tear proteomic profiles in post-LASIK (Laser in-situ keratomileusis) dry eye
Effect of glaucoma medication on tear protein profiles and use them as biomarkers to predict the risk of glaucoma surgical failure
Tear biomarkers in lacrimal gland tumor
Tear proteomic profiles in thyroid eye disease
Tear proteomic profiles in keratoconus
Other important proteomics/metabolomics projects include:
Integrated “Omics” in ocular important drug-resistance pathogens
Circulating biomarkers in diabetic retinopathy using a metabolomics approach
Molecular mechanism of myopia and atropine treatment in a mouse model using quantitative proteomics
Vitreous proteomic profile in diabetic retinopathy
To identify blood biomarker for responder vs non-responder of AMD patients towards anti-VEGF therapy
To identify novel aqueous humor biomarker for AMD/PCV
Inflammation markers in aqueous humor from PAS, PAC, and PACG
Publications
Chen L, Gao Y, Wang LZ, Ning Cheung N, Tan GSW, Cheung CMG, Beuerman RW, Wong TY, Chan ECY, Zhou L*. Recent Advances in the Applications of Metabolomics in Eye Research. Analytica Chimica Acta. 2018, accepted. (IF = 5.123). * Corresponding author.
Tong L, Zhou L, Beuerman RW, Simonyi S, Hollander DA, Stern ME. Effects of punctal occlusion on global tear proteins in patients with dry eye. Ocular Surface. 2017, S1542-0124(16)30285-3. (IF = 5.530).
Zhou L*, Beuerman RW. The power of tears: how tear proteomics research could revolutionize the clinic (Editorial). Expert Review of Proteomics. 2017, 14(3):189-191. (IF = 3.849). * Corresponding author.
Chen L, Cheng CY, Choi HW, Ikram MK, Sabanayagam C, Tan GS, Tian DC, Zhang L, Venkatesan G, Tai ES, Wang JJ, Mitchell P, Cheung CM, Beuerman RW, Zhou L*, Chan ECY*, Wong TY*. Plasma Metabonomic Profiling of Diabetic Retinopathy. Diabetes. 2016, Apr;65(4):1099-108. (IF = 8.474). * Corresponding author.
Chen L, Li J, Guo T, Ghosh S, Koh SK, Tian D, Zhang L, Jia D, Beuerman RW, Aebersold R, Chan EC, Zhou L*. Global Metabonomic and Proteomic Analysis of Human Conjunctival Epithelial Cells (IOBA-NHC) in Response to Hyperosmotic Stress. Journal of Proteome Research. 2015, 14, 3982-3995. (IF = 5.001). *Corresponding author.
Tong L, Zhou XY, Jylha A, Aapola U, Liu DN, Koh SK, Tian D, Quah J, Uusitalo H, Beuerman RW, Zhou L*. Quantitation of 47 Human Tear Proteins using High Resolution Multiple Reaction Monitoring (HR-MRM) Based-Mass Spectrometry. Journal of Proteomics. 2015, 115, 36-48. (IF =5.080). *Corresponding author.
Barathi VA, Chaurasia SS, Poidinger M, Koh SK, Tian D, Ho C, Iuvone MP, Beuerman RW, Zhou L*. Involvement of GABA transporters in Atropine-treated Myopic Retina as revealed by iTRAQ Quantitative Proteomics. Journal of Proteome Research. 2014, Nov. 7; 13(11): 4647-4658. (IF = 5.001). *Corresponding author.
Zhou L*, Wei Ruihua, Zhao Ping, Koh Siew Kwan, Beuerman Roger W, Ding Chuanqing, Proteomic Analysis Revealed the Altered Tear Protein Profile in a Rabbit Model of Sjögren’s Syndrome-Associated Dry Eye. Proteomics, 2013, 13(16), 2469-2481. *Corresponding author.
Zhou L, Beuerman RW. Tear analysis in ocular surface diseases. Progress in Retinal and Eye Research, 2012, 31(6), 527-550. (IF = 11.653).
Zhou L*, Zhao SZ, Koh SK, Chen L, Vaz C, Tanavde V, Li XR, Beuerman RW. In-depth analysis of the human tear proteome. Journal of Proteomics, 2012, 75(13), 3877-3885. (IF = 5.080). *Corresponding author.
Chen L, Zhou L*, Chan E, Neo J, Beuerman, RW, Characterization of The Human Tear Metabolome by LC-MS/MS. Journal of Proteome Research. 2011, Oct 7;10(10):4876-82. (IF = 5.617). *Corresponding author.
Wong TT, Zhou L, Li J, Tong L, Zhao SZ, Li XR, Yu SJ, Koh SK, Beuerman RW, Proteomic profiling of inflammatory signaling molecules in the tears of patients on chronic glaucoma medication, Invest. Ophthalmol. Vis. Sci. 2011, Sep 22;52(10):7385-91. (IF = 3.933).
Zhou L, Liu SP, Li J, Ong LB, Guo L, Wohland T, Tang C, Lakshminarayanan R, Mavinahalli J, Verma C, Beuerman RW. The structural parameters to antimicrobial activity, human epithelia cell cytotoxicity and killing mechanism of 10-AA synthetic analogues and covalent dimer derived from hBD3 C-terminal. Amino Acids. 2011 Jan;40(1):123-33. (IF = 4.106).
Zhou L, Beuerman RW, Chew AP, Koh SK, Cafaro TA, Urrets-Zavalia EA, Urrets-Zavalia JA, Li SFY, Serra HM. Quantitative Analysis of N-linked Glycoproteins in Tear Fluid of Climatic Droplet Keratopathy by Glycopeptide Capture and iTRAQ. Journal of Proteome Research, 2009, 8(4), 1992-2003. (IF = 5.737).
Zhou L, Beuerman RW, Chan CM, Zhao SZ, Li XR, Yang H, Tong L, Liu SP, Stern ME, Tan DTH. Identification of Tear Fluid Biomarkers in Dry Eye Syndrome Using iTRAQ Quantitative Proteomics. Journal of Proteome Research, 2009, 8(11), 4889-4905. (IF = 5.737).
Head
Assoc Prof Wang Xiaomeng
Experimental Microscopy, Molecular & Cell Biology
The Experimental Microscopy, Molecular & Cell Biology Research Platform provides the infrastructure and a range of essential and sophisticated core equipment to facilitate researchers in Singapore Eye Research Institute/Singapore National Eye Centre in their basic to advanced molecular and cell biology studies.
Our laboratory space occupies two floors, on the 11th and 12th floors of The Academia. There are 33 general laboratory benches as well as rooms dedicated to the following:
Chemistry
Microbiology
Gel electrophoresis
Dark room/ X-ray processor
Tissue culture
Core equipment includes:
absorbance/ fluorescence/ luminescence plate readers
autoclaves
Bio-Plex 200 system
BD FACSVerse™ flow cytometer
ChemiDoc Imager
circular dichroism spectroscopy
Essen IncuCyte zoom system
Guava Easycyte flow cytometer
Instron universal material testing system
real-time PCR instruments
droplet digital PCR system
medical X-Ray processor
microvolume spectrophotometers
water filter system
xCELLigence RTCA systems
Manpower
This research platform is managed by the SERI laboratory administration team and supported by scientists, research officers and laboratory executives to ensure optimal performance of all facilities and equipment. We comply with the rules and regulations of the SingHealth Institutional Biosafety Committee (SHSIBC).
Molecular and cellular experimentation is fundamental to understanding the mechanisms of eye diseases, as well as drugs for therapy. The projects facilitated by this platform are multidisciplinary, diverse and vary in both breadth and depth. Besides supporting experimental research, this platform also offers assistance for assessing other resources within the SingHealth campus and beyond. We collaborate extensively with other academic institutions within Singapore. We also welcome collaboration with industry partners interested in eye-related research or the exploration of innovative molecular and cell technologies. Our industry collaborators include AYOXXA Biosystems (Cologne, Germany) and Santen Pharmaceutical Co., Ltd. (Osaka, Japan).
Examples of ongoing research areas include:
Corneal endothelial cultures for ocular transplantation
Functional genomics
Therapeutics and drug delivery systems for dry eye, ocular surgery, etc.
Anti-microbial therapeutics and biomaterial
Stem cell therapy for retinal diseases
Epidemiology and biomarkers of retinal diseases
Translational Ophthalmic Pathology & Immunology
The Translational Ophthalmic Pathology & Immunology Research Platform is a core platform that specializes in molecular pathology for the study of the pathology of ocular disorders. In our laboratory we have the latest DEPArray NXt © machine which has the capacity to sort single cells from fresh and fixed tissues, as well as paraffin embedded tissues for further downstream analysis. Tissues may be liquid based or solid tissue. This technology is superior to routine flow cytometry as it is able to handle single cells. The major disadvantage in working with ocular tissues is the small specimen size and limited cellular quantity. This technology is crucial for advancing molecular capabilities in our ophthalmic related research.
The research platform is led by Dr Anita Chan who is a practicing ophthalmologist, as well as a trained ophthalmic pathologist. In our team, we have expertise in analyzing human and animal related histology for clinical and research studies respectively.
In addition to molecular pathology, we also offer advance histological services including:
Multiplex immunofluorescence imaging
Immunofluorescence imaging
Immunohistochemistry
Electron microscopy
Our research platform works closely with the other core units in SERI (ocular genetics, proteomics, translational pre-clinical model research platform, as well as the microscopy imaging unit) to provide a comprehensive approach for ophthalmic research studies.
We currently have several industry research collaborations ongoing, including work with Santen Pharmaceutical and Menarini Biomarkers.
Other studies include the use of DEPARRay to study iris tissue in exfoliation syndrome.
Dr Anita Chan (PI)
Team members
Mona Wang
Myoe Naing Lynn
Collaborators
Dr Lim Tong Seng (Co-PI, MBS collaboration)
Dr Tan Wei Jian (Post doc, MBS collaboration)
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