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Fulltext Biotin-Streptavidin Competition Mediates Sensitive Detection of Biomolecules in Enzyme Linked Immunosorbent Assay

Lakshmipriya T, Gopinath SC, Tang TH

PLoS One, 2016;11(3):e0151153.
PMID: 26954237 DOI: 10.1371/journal.pone.0151153

Enzyme Linked Immunosorbent Assay (ELISA) is the gold standard assay for detecting and identifying biomolecules using antibodies as the probe. Improving ELISA is crucial for detecting disease-causing agents and facilitating diagnosis at the early stages of disease. Biotinylated antibody and streptavidin-conjugated horse radish peroxide (streptavidin-HRP) often are used with ELISA to enhance the detection of various kinds of targets. In the present study, we used a competition-based strategy in which we pre-mixed free biotin with streptavidin-HRP to generate high-performance system, as free biotin occupies some of the biotin binding sites on streptavidin, thereby providing more chances for streptavidin-HRP to bind with biotinylated antibody. ESAT-6, which is a protein secreted early during tuberculosis infection, was used as the model target. We found that 8 fM of free biotin mixed with streptavidin-HRP anchored the higher detection level of ESAT-6 by four-fold compared with detection without free biotin (only streptavidin-HRP), and the limit of detection of the new method was 250 pM. These results suggest that biotin-streptavidin competition can be used to improve the diagnosis of analytes in other types of sensors.
Fulltext Aptamer-Antibody Complementation On Multiwalled Carbon Nanotube-Gold Transduced Dielectrode Surfaces To Detect Pandemic Swine Influenza Virus

Wang F, Gopinath SC, Lakshmipriya T

Int J Nanomedicine, 2019;14:8469-8481.
PMID: 31695375 DOI: 10.2147/IJN.S219976

BACKGROUND: A pandemic influenza viral strain, influenza A/California/07/2009 (pdmH1N1), has been considered to be a potential issue that needs to be controlled to avoid the seasonal emergence of mutated strains.
MATERIALS AND METHODS: In this study, aptamer-antibody complementation was implemented on a multiwalled carbon nanotube-gold conjugated sensing surface with a dielectrode to detect pandemic pdmH1N1. Preliminary biomolecular and dielectrode surface analyses were performed by molecular and microscopic methods. A stable anti-pdmH1N1 aptamer sequence interacted with hemagglutinin (HA) and was compared with the antibody interaction. Both aptamer and antibody attachments on the surface as the basic molecule attained the saturation at nanomolar levels.
RESULTS: Aptamers were found to have higher affinity and electric response than antibodies against HA of pdmH1N1. Linear regression with aptamer-HA interaction displays sensitivity in the range of 10 fM, whereas antibody-HA interaction shows a 100-fold lower level (1 pM). When sandwich-based detection of aptamer-HA-antibody and antibody-HA-aptamer was performed, a higher response of current was observed in both cases. Moreover, the detection strategy with aptamer clearly discriminated the closely related HA of influenza B/Tokyo/53/99 and influenza A/Panama/2007/1999 (H3N2).
CONCLUSION: The high performance of the abovementioned detection methods was supported by the apparent specificity and reproducibility by the demonstrated sensing system.
Aptamers as a replacement for antibodies in enzyme-linked immunosorbent assay

Toh SY, Citartan M, Gopinath SC, Tang TH

Biosens Bioelectron, 2015 Feb 15;64:392-403.
PMID: 25278480 DOI: 10.1016/j.bios.2014.09.026

The application of antibodies in enzyme-linked immunosorbent assay (ELISA) is the basis of this diagnostic technique which is designed to detect a potpourri of complex target molecules such as cell surface antigens, allergens, and food contaminants. However, development of the systematic evolution of Ligands by Exponential Enrichment (SELEX) method, which can generate a nucleic acid-based probe (aptamer) that possess numerous advantages compared to antibodies, offers the possibility of using aptamers as an alternative molecular recognition element in ELISA. Compared to antibodies, aptamers are smaller in size, can be easily modified, are cheaper to produce, and can be generated against a wide array of target molecules. The application of aptamers in ELISA gives rise to an ELISA-derived assay called enzyme-linked apta-sorbent assay (ELASA). As with the ELISA method, ELASA can be used in several different configurations, including direct, indirect, and sandwich assays. This review provides an overview of the strategies involved in aptamer-based ELASA.
Label-free methods of reporting biomolecular interactions by optical biosensors

Citartan M, Gopinath SC, Tominaga J, Tang TH

Analyst, 2013 Jul 7;138(13):3576-92.
PMID: 23646346 DOI: 10.1039/c3an36828a

Reporting biomolecular interactions has become part and parcel of many applications of science towards an in-depth understanding of disease and gene regulation. Apart from that, in diagnostic applications where biomolecules (antibodies and aptamers) are vastly applied, meticulous monitoring of biomolecular interaction is vital for clear-cut diagnosis. Several currently available methods of analyzing the interaction of the ligands with the appropriate analytes are aided by labeling using fluorescence or luminescence techniques. However, labeling is cumbersome and can occupy important binding sites of interactive molecules to be labeled, which may interfere with the conformational changes of the molecules and increase non-specificity. Optical-based sensing can provide an alternative way as a label-free procedure for monitoring biomolecular interactions. Optical sensors affiliated with different operating principles, including surface plasmon changes, scattering and interferometry, can impart a huge impact for in-house and point-of-care applications. This optical-based biosensing permits real-time monitoring, obviating the use of hazardous labeling molecules such as radioactive tags. Herein, label-free ways of reporting biomolecular interactions by various optical biosensors were gleaned.
Fulltext Titanium Dioxide Nanoparticle-Based Interdigitated Electrodes: A Novel Current to Voltage DNA Biosensor Recognizes E. coli O157:H7

Nadzirah Sh, Azizah N, Hashim U, Gopinath SC, Kashif M

PLoS One, 2015;10(10):e0139766.
PMID: 26445455 DOI: 10.1371/journal.pone.0139766

Nanoparticle-mediated bio-sensing promoted the development of novel sensors in the front of medical diagnosis. In the present study, we have generated and examined the potential of titanium dioxide (TiO2) crystalline nanoparticles with aluminium interdigitated electrode biosensor to specifically detect single-stranded E.coli O157:H7 DNA. The performance of this novel DNA biosensor was measured the electrical current response using a picoammeter. The sensor surface was chemically functionalized with (3-aminopropyl) triethoxysilane (APTES) to provide contact between the organic and inorganic surfaces of a single-stranded DNA probe and TiO2 nanoparticles while maintaining the sensing system's physical characteristics. The complement of the target DNA of E. coli O157:H7 to the carboxylate-probe DNA could be translated into electrical signals and confirmed by the increased conductivity in the current-to-voltage curves. The specificity experiments indicate that the biosensor can discriminate between the complementary sequences from the base-mismatched and the non-complementary sequences. After duplex formation, the complementary target sequence can be quantified over a wide range with a detection limit of 1.0 x 10(-13)M. With target DNA from the lysed E. coli O157:H7, we could attain similar sensitivity. Stability of DNA immobilized surface was calculated with the relative standard deviation (4.6%), displayed the retaining with 99% of its original response current until 6 months. This high-performance interdigitated DNA biosensor with high sensitivity, stability and non-fouling on a novel sensing platform is suitable for a wide range of biomolecular interactive analyses.
Fulltext Variations in Spontaneous Assembly and Disassembly of Molecules on Unmodified Gold Nanoparticles

Jin NZ, Anniebell S, Gopinath SC, Chen Y

Nanoscale Res Lett, 2016 Dec;11(1):399.
PMID: 27637891 DOI: 10.1186/s11671-016-1615-2

Electrostatic attraction, covalent binding, and hydrophobic absorption are spontaneous processes to assemble and disassemble the molecules of gold nanoparticles (GNP). This dynamic change can be performed in the presence of ions, such as NaCl or charged molecules. Current research encompasses the GNP in mediating non-biofouling and investigating the molecular attachment and detachment. Experiments were performed with different sizes of GNP and polymers. As a proof of concept, poly(ethylene glycol)-b-poly(acrylic acid), called PEG-PAAc, attachment and binding events between factor IX and factor IX-bp from snake venom were demonstrated, and the variations with these molecular attachment on GNP were shown. Optimal concentration of NaCl for GNP aggregation was 250 mM, and the optimal size of GNP used was 30 nm. The polymer PEG-PAAc (1 mg/ml) has a strong affinity to the GNP as indicated by the dispersed GNP. The concentration of 5800 nM of factor IX was proved to be optimal for dispersion of GNP, and at least 100 nM of factor IX-bp was needed to remove factor IX from the surface of GNP. This study delineates the usage of unmodified GNP for molecular analysis and downstream applications.
Fulltext Thickness Dependent Nanostructural, Morphological, Optical and Impedometric Analyses of Zinc Oxide-Gold Hybrids: Nanoparticle to Thin Film

Perumal V, Hashim U, Gopinath SC, Haarindraprasad R, Liu WW, Poopalan P, et al.

PLoS One, 2015;10(12):e0144964.
PMID: 26694656 DOI: 10.1371/journal.pone.0144964

The creation of an appropriate thin film is important for the development of novel sensing surfaces, which will ultimately enhance the properties and output of high-performance sensors. In this study, we have fabricated and characterized zinc oxide (ZnO) thin films on silicon substrates, which were hybridized with gold nanoparticles (AuNPs) to obtain ZnO-Aux (x = 10, 20, 30, 40 and 50 nm) hybrid structures with different thicknesses. Nanoscale imaging by field emission scanning electron microscopy revealed increasing film uniformity and coverage with the Au deposition thickness. Transmission electron microscopy analysis indicated that the AuNPs exhibit an increasing average diameter (5-10 nm). The face center cubic Au were found to co-exist with wurtzite ZnO nanostructure. Atomic force microscopy observations revealed that as the Au content increased, the overall crystallite size increased, which was supported by X-ray diffraction measurements. The structural characterizations indicated that the Au on the ZnO crystal lattice exists without any impurities in a preferred orientation (002). When the ZnO thickness increased from 10 to 40 nm, transmittance and an optical bandgap value decreased. Interestingly, with 50 nm thickness, the band gap value was increased, which might be due to the Burstein-Moss effect. Photoluminescence studies revealed that the overall structural defect (green emission) improved significantly as the Au deposition increased. The impedance measurements shows a decreasing value of impedance arc with increasing Au thicknesses (0 to 40 nm). In contrast, the 50 nm AuNP impedance arc shows an increased value compared to lower sputtering thicknesses, which indicated the presence of larger sized AuNPs that form a continuous film, and its ohmic characteristics changed to rectifying characteristics. This improved hybrid thin film (ZnO/Au) is suitable for a wide range of sensing applications.
Fulltext Identification of circulating biomarkers in sera of Plasmodium knowlesi-infected malaria patients--comparison against Plasmodium vivax infection

Chen Y, Chan CK, Kerishnan JP, Lau YL, Wong YL, Gopinath SC

BMC Infect Dis, 2015;15:49.
PMID: 25656928 DOI: 10.1186/s12879-015-0786-2

Plasmodium knowlesi was identified as the fifth major malaria parasite in humans. It presents severe clinical symptoms and leads to mortality as a result of hyperparasitemia in a short period of time. This study aimed to improve the current understanding of P. knowlesi and identify potential biomarkers for knowlesi malaria.
Fulltext Identification of host-immune response protein candidates in the sera of human oral squamous cell carcinoma patients

Chen Y, Azman SN, Kerishnan JP, Zain RB, Chen YN, Wong YL, et al.

PLoS One, 2014;9(10):e109012.
PMID: 25272005 DOI: 10.1371/journal.pone.0109012

One of the most common cancers worldwide is oral squamous cell carcinoma (OSCC), which is associated with a significant death rate and has been linked to several risk factors. Notably, failure to detect these neoplasms at an early stage represents a fundamental barrier to improving the survival and quality of life of OSCC patients. In the present study, serum samples from OSCC patients (n = 25) and healthy controls (n = 25) were subjected to two-dimensional gel electrophoresis (2-DE) and silver staining in order to identify biomarkers that might allow early diagnosis. In this regard, 2-DE spots corresponding to various up- and down-regulated proteins were sequenced via high-resolution MALDI-TOF mass spectrometry and analyzed using the MASCOT database. We identified the following differentially expressed host-specific proteins within sera from OSCC patients: leucine-rich α2-glycoprotein (LRG), alpha-1-B-glycoprotein (ABG), clusterin (CLU), PRO2044, haptoglobin (HAP), complement C3c (C3), proapolipoprotein A1 (proapo-A1), and retinol-binding protein 4 precursor (RBP4). Moreover, five non-host factors were detected, including bacterial antigens from Acinetobacter lwoffii, Burkholderia multivorans, Myxococcus xanthus, Laribacter hongkongensis, and Streptococcus salivarius. Subsequently, we analyzed the immunogenicity of these proteins using pooled sera from OSCC patients. In this regard, five of these candidate biomarkers were found to be immunoreactive: CLU, HAP, C3, proapo-A1 and RBP4. Taken together, our immunoproteomics approach has identified various serum biomarkers that could facilitate the development of early diagnostic tools for OSCC.
Sensing strategies for influenza surveillance

Gopinath SC, Tang TH, Chen Y, Citartan M, Tominaga J, Lakshmipriya T

Biosens Bioelectron, 2014 Nov 15;61:357-69.
PMID: 24912036 DOI: 10.1016/j.bios.2014.05.024

Influenza viruses, which are RNA viruses belonging to the family Orthomyxoviridae, cause respiratory diseases in birds and mammals. With seasonal epidemics, influenza spreads all over the world, resulting in pandemics that cause millions of deaths. Emergence of various types and subtypes of influenza, such as H1N1 and H7N9, requires effective surveillance to prevent their spread and to develop appropriate anti-influenza vaccines. Diagnostic probes such as glycans, aptamers, and antibodies now allow discrimination among the influenza strains, including new subtypes. Several sensors have been developed based on these probes, efforts made to augment influenza detection. Herein, we review the currently available sensing strategies to detect influenza viruses.
Bacterial detection: from microscope to smartphone

Gopinath SC, Tang TH, Chen Y, Citartan M, Lakshmipriya T

Biosens Bioelectron, 2014 Oct 15;60:332-42.
PMID: 24836016 DOI: 10.1016/j.bios.2014.04.014

The ubiquitous nature of bacteria enables them to survive in a wide variety of environments. Hence, the rise of various pathogenic species that are harmful to human health raises the need for the development of accurate sensing systems. Sensing systems are necessary for diagnosis and epidemiological control of pathogenic organism, especially in the food-borne pathogen and sanitary water treatment facility' bacterial populations. Bacterial sensing for the purpose of diagnosis can function in three ways: bacterial morphological visualization, specific detection of bacterial component and whole cell detection. This paper provides an overview of the currently available bacterial detection systems that ranges from microscopic observation to state-of-the-art smartphone-based detection.
Use of UV-vis-NIR spectroscopy to monitor label-free interaction between molecular recognition elements and erythropoietin on a gold-coated polycarbonate platform

Citartan M, Gopinath SC, Tominaga J, Chen Y, Tang TH

Talanta, 2014 Aug;126:103-9.
PMID: 24881539 DOI: 10.1016/j.talanta.2014.03.043

Label-free-based detection is pivotal for real-time monitoring of biomolecular interactions and to eliminate the need for labeling with tags that can occupy important binding sites of biomolecules. One simplest form of label-free-based detection is ultraviolet-visible-near-infrared (UV-vis-NIR) spectroscopy, which measure changes in reflectivity as a means to monitor immobilization and interaction of biomolecules with their corresponding partners. In biosensor development, the platform used for the biomolecular interaction should be suitable for different molecular recognition elements. In this study, gold (Au)-coated polycarbonate was used as a platform and as a proof-of-concept, erythropoietin (EPO), a doping substance widely abused by the athletes was used as the target. The interaction of EPO with its corresponding molecular recognition elements (anti-EPO monoclonal antibody and anti-EPO DNA aptamer) is monitored by UV-vis-NIR spectroscopy. Prior to this, to show that UV-vis-NIR spectroscopy is a suitable method for measuring biomolecular interaction, the interaction between biotin and streptavidin was demonstrated via this strategy and reflectivity of this interaction decreased by 25%. Subsequent to this, interaction of the EPO with anti-EPO monoclonal antibody and anti-EPO DNA aptamer resulted in the decrease of reflectivity by 5% and 10%, respectively. The results indicated that Au-coated polycarbonate could be an ideal biosensor platform for monitoring biomolecular interactions using UV-vis-NIR spectroscopy. A smaller version of the Au-coated polycarbonate substrates can be derived from the recent set-up, to be applied towards detecting EPO abuse among atheletes.
Current aspects in immunosensors

Gopinath SC, Tang TH, Citartan M, Chen Y, Lakshmipriya T

Biosens Bioelectron, 2014 Jul 15;57:292-302.
PMID: 24607580 DOI: 10.1016/j.bios.2014.02.029

Sensing applications can be used to report biomolecular interactions in order to elucidate the functions of molecules. The use of an analyte and a ligand is a common set-up in sensor development. For several decades, antibodies have been considered to be potential analytes or ligands for development of so-called "immunosensors." In an immunosensor, formation of the complex between antibody and antigen transduces the signal, which is measurable in various ways (e.g., both labeled and label-free based detection). Success of an immunosensor depends on various factors, including surface functionalization, antibody orientation, density of the antibody on the sensor platform, and configuration of the immunosensor. Careful optimization of these factors can generate clear-cut results for any immunosensor. Herein, current aspects, involved in the generated immunosensors, are discussed.
Single-stranded DNA (ssDNA) production in DNA aptamer generation

Marimuthu C, Tang TH, Tominaga J, Tan SC, Gopinath SC

Analyst, 2012 Mar 21;137(6):1307-15.
PMID: 22314701 DOI: 10.1039/c2an15905h

The discovery that synthetic short chain nucleic acids are capable of selective binding to biological targets has made them to be widely used as molecular recognition elements. These nucleic acids, called aptamers, are comprised of two types, DNA and RNA aptamers, where the DNA aptamer is preferred over the latter due to its stability, making it widely used in a number of applications. However, the success of the DNA selection process through Systematic Evolution of Ligands by Exponential Enrichment (SELEX) experiments is very much dependent on its most critical step, which is the conversion of the dsDNA to ssDNA. There is a plethora of methods available in generating ssDNA from the corresponding dsDNA. These include asymmetric PCR, biotin-streptavidin separation, lambda exonuclease digestion and size separation on denaturing-urea PAGE. Herein, different methods of ssDNA generation following the PCR amplification step in SELEX are reviewed.
Assays for aptamer-based platforms

Citartan M, Gopinath SC, Tominaga J, Tan SC, Tang TH

Biosens Bioelectron, 2012 Apr 15;34(1):1-11.
PMID: 22326894 DOI: 10.1016/j.bios.2012.01.002

Aptamers are single stranded DNA or RNA oligonucleotides that have high affinity and specificity towards a wide range of target molecules. Aptamers have low molecular weight, amenable to chemical modifications and exhibit stability undeterred by repetitive denaturation and renaturation. Owing to these indispensable advantages, aptamers have been implemented as molecular recognition element as alternative to antibodies in various assays for diagnostics. By amalgamating with a number of methods that can provide information on the aptamer-target complex formation, aptamers have become the elemental tool for numerous biosensor developments. In this review, administration of aptamers in applications involving assays of fluorescence, electrochemistry, nano-label and nano-constructs are discussed. Although detection strategies are different for various aptamer-based assays, the core of the design strategies is similar towards reporting the presence of specific target binding to the corresponding aptamers. It is prognosticated that aptamers will find even broader applications with the development of new methods of transducing aptamer target binding.
A new nano-worm structure from gold-nanoparticle mediated random curving of zinc oxide nanorods

Perumal V, Hashim U, Gopinath SC, Haarindraprasad R, Poopalan P, Liu WW, et al.

Biosens Bioelectron, 2016 Apr 15;78:14-22.
PMID: 26584078 DOI: 10.1016/j.bios.2015.10.083

Creating novel nanostructures is a primary step for high-performance analytical sensing. Herein, a new worm like nanostructure with Zinc Oxide-gold (ZnO/Au) hybrid was fabricated through an aqueous hydrothermal method, by doping Au-nanoparticle (AuNP) on the growing ZnO lattice. During ZnO growth, fine tuning the solution temperature expedites random curving of ZnO nanorods and forms nano-worms. The nano-worms which were evidenced by morphological, physical and structural analyses, revealed elongated structures protruding from the surface (length: 1 µm; diameter: ~100 nm). The appropriate peaks for the face centred cubic gold were (111) and (200), as seen from X-ray diffractogram. The strong interrelation between Au and ZnO was manifested by X-ray photoelectron spectroscopy. The combined surface area increment from the nanoparticle radii and ZnO nanorod random curving gives raise an enhancement in detection sensitivity by increasing bio-loading. 'Au-decorated hybrid nano-worm' was immobilized with a probe DNA from Vibrio Cholera and duplexed with a target which was revealed by Fourier Transform Infrared Spectroscopy. Our novel Au-decorated hybrid nano-worm is suitable for high-performance bio-sensing, as evidenced by impedance spectroscopy, having higher-specificity and attained femtomolar (10 fM) sensitivity. Further, higher stability, reproducibility and regeneration on this sensing surface were demonstrated.
Polysilicon nanogap lab-on-chip facilitates multiplex analyses with single analyte

Balakrishnan SR, Hashim U, Gopinath SC, Poopalan P, Ramayya HR, Veeradasan P, et al.

Biosens Bioelectron, 2016 Oct 15;84:44-52.
PMID: 26560969 DOI: 10.1016/j.bios.2015.10.075

Rationally designed biosensing system supports multiplex analyses is warranted for medical diagnosis to determine the level of analyte interaction. The chemically functionalized novel multi-electrode polysilicon nanogap (PSNG) lab-on-chip is designed in this study, facilitates multiplex analyses for a single analyte. On the fabricated 69nm PSNG, biocompatibility and structural characteristics were verified for the efficient binding of Human Chorionic Gonadotropin (hCG). With the assistance of microfluidics, hCG sample was delivered via single-injection to 3-Aminopropyl(triethoxy)silane (APTES) and Glycidoxypropyl(trimethoxy)silane (GPMS) modified PSNG electrodes and the transduced signal was used to investigate the dielectric mechanisms for multiplex analyses. The results from amperometric response and impedance measurement delivered the scale of interaction between anti-hCG antibody and hCG that exhibited 6.5 times higher sensitivity for the chemical linker, APTES than GPMS. Under optimized experimental conditions, APTES and GPMS modified immunosensor has a limit of detection as 0.56mIU/ml and 2.93mIU/ml (at S/N=3), with dissociation constants (Kd) of 5.65±2.5mIU/ml and 7.28±2.6mIU/ml, respectively. These results suggest that multiplex analysis of single target could enhance the accuracy of detection and reliable for real-time comparative analyses. The designed PSNG is simple, feasible, requires low sample consumption and could be applied for any given multiplex analyses.
Fulltext A Point-of-Care Immunosensor for Human Chorionic Gonadotropin in Clinical Urine Samples Using a Cuneated Polysilicon Nanogap Lab-on-Chip

Balakrishnan SR, Hashim U, Gopinath SC, Poopalan P, Ramayya HR, Iqbal Omar M, et al.

PLoS One, 2015;10(9):e0137891.
PMID: 26368287 DOI: 10.1371/journal.pone.0137891

Human chorionic gonadotropin (hCG), a glycoprotein hormone secreted from the placenta, is a key molecule that indicates pregnancy. Here, we have designed a cost-effective, label-free, in situ point-of-care (POC) immunosensor to estimate hCG using a cuneated 25 nm polysilicon nanogap electrode. A tiny chip with the dimensions of 20.5 × 12.5 mm was fabricated using conventional lithography and size expansion techniques. Furthermore, the sensing surface was functionalized by (3-aminopropyl)triethoxysilane and quantitatively measured the variations in hCG levels from clinically obtained human urine samples. The dielectric properties of the present sensor are shown with a capacitance above 40 nF for samples from pregnant women; it was lower with samples from non-pregnant women. Furthermore, it has been proven that our sensor has a wide linear range of detection, as a sensitivity of 835.88 μA mIU(-1) ml(-2) cm(-2) was attained, and the detection limit was 0.28 mIU/ml (27.78 pg/ml). The dissociation constant Kd of the specific antigen binding to the anti-hCG was calculated as 2.23 ± 0.66 mIU, and the maximum number of binding sites per antigen was Bmax = 22.54 ± 1.46 mIU. The sensing system shown here, with a narrow nanogap, is suitable for high-throughput POC diagnosis, and a single injection can obtain triplicate data or parallel analyses of different targets.
Fulltext Biopolymers Regulate Silver Nanoparticle under Microwave Irradiation for Effective Antibacterial and Antibiofilm Activities

Velusamy P, Su CH, Venkat Kumar G, Adhikary S, Pandian K, Gopinath SC, et al.

PLoS One, 2016;11(6):e0157612.
PMID: 27304672 DOI: 10.1371/journal.pone.0157612

In the current study, facile synthesis of carboxymethyl cellulose (CMC) and sodium alginate capped silver nanoparticles (AgNPs) was examined using microwave radiation and aniline as a reducing agent. The biopolymer matrix embedded nanoparticles were synthesized under various experimental conditions using different concentrations of biopolymer (0.5, 1, 1.5, 2%), volumes of reducing agent (50, 100, 150 μL), and duration of heat treatment (30 s to 240 s). The synthesized nanoparticles were analyzed by scanning electron microscopy, UV-Vis spectroscopy, X-ray diffraction, and Fourier transform infrared spectroscopy for identification of AgNPs synthesis, crystal nature, shape, size, and type of capping action. In addition, the significant antibacterial efficacy and antibiofilm activity of biopolymer capped AgNPs were demonstrated against different bacterial strains, Staphylococcus aureus MTCC 740 and Escherichia coli MTCC 9492. These results confirmed the potential for production of biopolymer capped AgNPs grown under microwave irradiation, which can be used for industrial and biomedical applications.
Fulltext Secretion of N- and O-linked Glycoproteins from 4T1 Murine Mammary Carcinoma Cells

Phang WM, Tan AA, Gopinath SC, Hashim OH, Kiew LV, Chen Y

Int J Med Sci, 2016;13(5):330-9.
PMID: 27226773 DOI: 10.7150/ijms.14341

Breast cancer is one of the most common cancers that affect women globally and accounts for ~23% of all cancers diagnosed in women. Breast cancer is also one of the leading causes of death primarily due to late stage diagnoses and a lack of effective treatments. Therefore, discovering protein expression biomarkers is mandatory for early detection and thus, critical for successful therapy. Two-dimensional electrophoresis (2D-E) coupled with lectin-based analysis followed by mass spectrometry were applied to identify potential biomarkers in the secretions of a murine mammary carcinoma cell line. Comparisons of the protein profiles of the murine 4T1 mammary carcinoma cell line and a normal murine MM3MG mammary cell line indicated that cadherin-1 (CDH), collagenase 3 (MMP-13), Viral envelope protein G7e (VEP), Gag protein (GAG) and Hypothetical protein LOC433182 (LOC) were uniquely expressed by the 4T1 cells, and pigment epithelium-derived factor (PEDF) was exclusively secreted by the MM3MG cells. Further analysis by a lectin-based study revealed that aberrant O-glycosylated CDH, N-glycosylated MMP-13 and LOC were present in the 4T1 medium. These differentially expressed N- and O-linked glycoprotein candidates, which were identified by combining lectin-based analysis with 2D-E, could serve as potential diagnostic and prognostic markers for breast cancer.

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