international conference and exhibition on biosensors

Biosensors & Bioelectronics-2012May 14-16, 2012 Volume 2 Issue 4 - 46

J Biosens BioelectronISSN:2155-6210 JBSBE an open access journal

International Conference and Exhibition on

Biosensors & BioelectronicsMay 14-16, 2012 Embassy Suites Las Vegas, USA

Novel quantitative FRET assays for protein interaction dissociation constant, Kd, and Protease Kinetics, Kcat/KM determinationsJiayu LiaoDepartment of Bioengineering, Institute for Integrative Genome Biology and Biomedical Science, University of California, USA

Förster resonance energy transfer (FRET) technology has been widely used in biological and biomedical research. We recently developed a novel quantitative FRET analysis method and applied this method to determine protein interaction affinities and

protease kinetics in the SUMOylation cascade. The novel methodology is based on the quantitative analysis of the FRET signal from the total fluorescent signal at acceptor emission wavelength, which consists of three components: donor emission, acceptor emission and FRET signal during the digestion process. SUMOylation is an important protein post-translational modification, which is carried out by multi-step enzymatic cascade reactions for peptide activation and substrate conjugation, and plays critical roles in diverse physiological processes, including transcriptional regulation, signal transduction, cell survival and death and DNA damage response. We developed a new theoretical and experimental procedure for protein interaction Kd determination of SUMO1 and its E2 ligase, Ubc9, and individual interaction in the full SUMOylation cascade by FRET assay. The Kd values are also consistent with those determined with other traditional approaches, such as surface plasmon resonance(SPR) and isothermal titration calorimetry(ITC). We have also developed a novel quantitative FRET-based protease assay for SENP1 kinetics parameter determination. The novel theoretical and experimental procedures to determinethe kinetics parameters,kcat, KM and catalytic efficiency (kcat/KM) of catalytic domain SENP1 towards preSUMO1. These results are superior than those obtained from biochemical assays. These developments represent a novel methodology of biosensor based on FRET, which can be used in general for protein-protein interaction dissociation constant and protease kinetics determinations.

BiographyJiayu Liao has completed his Ph.D in 1999 from the University of California at Los Angeles. He did his post-doc. training with Peter G. Schultz at the Scripps Research Institute. After he finished his post-doc. training, he joined GNF as Principle Investigator and Founding Scientist of GPCR platform. His collaboration work with Prof. Hugh Rosen on the discovery of EDG1-specific agonist as novel immunosuppressant at the Scripps Research Institute led to the award of the Scripps Molecular Screening Center from NIH Roadmap Initiative. His collaborative work with Prof. Mingwei Wang on the discovery of the first non-peptide agonist for Glucagon-like peptide 1 (GLP1) receptor. led to the establishment of Chinese Nation Compound Library in Shanghai. He joined the University of California at Riverside in 1999 as founding faculty of Department of Bioengineering. He has published more than 30 papers in reputed journals with more than 2000 citations and serving in several important review panels in USA and China.

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Jiayu Liao, J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05

mailto:[email protected]

http://dx.doi.org/10.4172/2155-6210.S1.05





Model membrane biosensor for probing the interfacial binding mechanism of phospholipase A2

Joshua A. Jackman1 and Nam-Joon Cho2

1School of Materials Science and Engineering, Nanyang Technological University, Singapore2Department of Medicine, Division of Gastroenterology and Hepatology, Stanford University, USA

Phospholipase A2 (sPLA2) is an important class of interfacial enzyme involved in key biological processes such as signal transduction and lipid metabolism. Pharmacologic modulation of sPLA2 represents an attractive therapeutic strategy for

several medical diseases including cancer, but efforts toward this goal have been stymied by the limited knowledge about the link between how sPLA2 members interact with target lipid membranes and their respective physiological functions. To gain insight into the biophysical mechanism underlying membrane association of sPLA2 enzymes, we developed a model membrane biosensor platform to probe the role of membrane electrostatics in this process. Real-time monitoring of the membrane association step with the quartz crystal microbalance with energy dissipation (QCM-D) technique enabled us to identify a novel bilayer-disrupting behavior that is dependent on membrane electrostatics. The data identified that 1) enzyme adsorption to model membranes is primarily mediated by non-electrostatic interactions; 2) nonhydrolytic bilayer-disruption can follow enzyme adsorption; and 3) this disruptive activity is directly related to electrostatic interactions. Based on these findings, we conclude that interfacial binding of sPLA2 enzymes is a dynamic process, and identify promising opportunities for therapeutic intervention as well as engineering approaches for sPLA2-triggered liposomal drug delivery.

BiographyJackman attended the University of Florida where he earned a bachelor’s degree with highest honors in chemistry, and was a member of Phi Beta Kappa. During his undergraduate studies, he was named a Beckman Scholar and pursued extensive research at Stanford University on the design and application of model membranes for biomedicine. Mr. Jackman then began his doctoral studies in biomedical engineering at Harvard University-Massachusetts Institute of Technology as an NSF Graduate Fellow, before transferring to Nanyang Technological University in Singapore in order to become involved in the emerging Asian biotechnology community, and to reunite with his former mentor from Stanford, Dr. Nam-Joon Cho.

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Joshua A. Jackman et al., J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05



http://dx.doi.org/10.4172/2155-6210.S1.05





Copper( II ) Complex-Based fluorescence sensors for nitric oxide detecting and imagingChunying Duan, Xiaoyue Hu, Xiaolin Zhang, Cheng He, Yongming Bao and Qin TangDalian University of Technology, China

Nitric oxide (NO) has received great attention since the discovery that it is an important signaling agent in biological systems.The method to directly detect NO in real time and in vivo is critical to the investigation of the multiple biological roles

of NO. While a variety of quantification techniques developed, fluorometric techniques become the gold standard for NO sensing, owing to their sensitivity and high spatiotemoral resolution, when combined with microscopy.The first generation of NO fluorescence sensors was based on organic molecules, which has a critical limitation that it cannot provide direct and real-time detection of NO. To resolve this problem, a novel strategy involving the reaction of transition metal and NO was explored recently. It was inspired by the interaction of NO and metal centers of enzyme in organism. Especially, the recent research showed that the sensors based on the Cu(II)-complex were more effective. According to above, we designed and synthesized a series of new Cu(II)-complex luminescence sensors for detection of NO, incorporating rhodamine/naphthalimide moiety as the luminescence active unit and polyamine as the efficient Cu(II) chelator. These sensors could be regarded as an approximate model of the enzyme center, which underwent a redox reaction with NO to restore the emission of fluorophore, giving an enhancement response. The new sensors exhibited high sensitivity (detection limit 1nM) and excellent specificity toward NO, and have been applied to NO imaging successfully.

Scheme 1. Proposed structures of CuQNE and mechanism of the reaction of CuQNE and NO.

BiographyChunying Duan was born in 1967, China. He completed his Ph.D. at Nanjing University in 1992. Then he started his academic career in the Department of Chemistry at Nanjing University. Since 2006, he has worked at Dalian University of Thechnology as the deputy director of State Key Laboratory of Fine Chemicals. His research interests cover aspects of supramolecular chemistry, coordination chemistry, molecular sensors, and chiral materials.

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Chunying Duan et al., J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05

NO

Fluorescence-on

OHN N

N OON OO

N NON NO

NOON

Fluorescence-off

N NO

Cu2+ Cu2+

N O N

X X

=

NHN

O

O

X = solvent


http://dx.doi.org/10.4172/2155-6210.S1.05





Real-time In-situ detection of microbes L. Powers1,2, W. R. Ellis Jr.2 and C. R. Lloyd3 1Electrical and Computer Engineering and 2Biomedical Engineering, The University of Arizona, USA 3MicroBioSystems of Utah, USA

Currently, no methods exist for the real-time detection and quantification of microbes in the environment or the detection and identification of pathogenic organisms in clinical specimens. We have addressed these problems with the development

of technologies which overcome these limitations and provide detection limits as low as a few microbes per L in water, per cm2 on abiotic surfaces, and per mL in body fluids. The detection and quantification of microbes [total microbial load] is based on the intrinsic fluorescence of microbial metabolites and protein cofactors and provides an estimate of the total microbial load as well as the relative distribution of live cells, dead cells, and endospores. Unlike existing methods, no additional reagents or sample contact is needed. This technology has been applied to the in-situ measurements of two sub-glacial microbial communities at sites in the Svalbard Archipelago, Norway, the efficacy of disinfection of contact lenses, determination of water quality from wells in Ifakara, Tanzania, and the in-line monitoring of water quality. In the rapid spread of a life-threatening infection, early diagnosis is of great importance. In such situations, pathogen counts will be very low, which also presents a significant challenge to diagnostic methods. We have developed a point-of care disposable diagnostic based on the en masse capture of bloodborne microbes from 1 mL of fresh whole blood with surface-tethered, small molecule ligands. Quantification is based on the intrinsic fluorescence of captured cells.

BiographyLinda Powers is the Thomas R Brown Chair in Bioengineering and Professor of Electrical and Computer Engineering as well as Biomedical Engineering. After receiving her PhD from Harvard University, she was a member of technical staff at AT&T Bell Laboratories. She has a broad scope of expertise from biochemistry to electrical engineering and has authored more than 125 technical publications in refereed journals. She is a fellow of the American Physical Society and the American Institute of Chemists and her honors include the US Bioenergetics Award of the Biophysical Society. She initiated two start-up companies based on her technology.

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L. Powers et al., J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05


http://dx.doi.org/10.4172/2155-6210.S1.05





Silicon nanowire Bio-FETs for Diabetes diagnosticsWalter Hu1,2, Serena Geene2, Michael Wallace1, Suresh Regonda1, Krutarth Trivedi1, Lisa Spurgin1, Bill Wu2 and Jiahuan Ding2

1Dept. of Electrical Engineering, University of Texas, USA 2Diagtronix Inc., USA

Here we report a nano-enabled electronic biosensor for direct detection of biomarkers e.g. insulin down to 10 femtoMolar levels in human serum and saliva samples. First we present a design and fabrication of silicon nanowire field effect transistor

(SiNW-FET) as biosensor. We use top-down strategy to fabricate silicon nanowire devices by the combination of photolithography and e-beam lithography on silicon on insulator wafers (SOI). The number of nanowires has shown a significant impact on the device performance, with better performance for larger number of nanowires used. We then test this type of sensor for medical diagnostic applications, such as insulin detection. Insulin is a diabetes related hormones and the quick detection of its level in blood is important for diagnosis of diabetes mellitus and also guiding its treatment. The traditional approaches for insulin quantification with mass spectra, ELISA or Western Blot etc are lack of sensitivity, time-consuming and/or requiring expensive equipments. Here we are using the SiNW FET devices to quantify the insulin level in human serum samples. Our preliminary sensing work has demonstrated repeatable detection of insulin directly from diluted patient serum without purification. The sensing experiments have repeatedly shown well correlated sensing of insulin with concentration from 10 femtoMolar (fM) to 1nM in pure PBS buffer and in human serum samples. We will demonstrate detailed sensing results related to diabetes disease in the meeting.

BiographyWalter Hu received the B.S. degree from Peking University, Beijing, China, in 1999, and the Ph.D. degree from the University of Notre Dame, Notre Dame, IN, in 2004. Then, he spent a year as a post-doctoral research fellow at the Department of Electrical Engineering, University of Michigan, Ann Arbor, MI. In September 2005, he joined the University of Texas at Dallas and currently an associate professor of the Department of Electrical Engineering. His research has been focused on lithography, nanofabrication, and applications in semiconductors, medical and energy areas. He has published 37 journal papers and 69 conference abstracts/papers, and 4 patent applications. He has received National Science Foundation’s CAREER award in 2010. He is a member of Sigma Xi, IEEE, AVS, MRS, ACS, and SPIE.

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Walter Hu et al., J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05


http://dx.doi.org/10.4172/2155-6210.S1.05





Magnetic bead-based “sample-to-answer” system for waterborne pathogen detection and enumerationQasem RamadanSwiss Federal Institute of Technology, Switzerland

Continuous surveillance of drinking water is important to provide early warning of contamination and ensure continuous supplies of healthy drinking water. Isolation and detection of a particular type of pathogen with low concentration diluted

in a large water sample and removing associated inhibitors from the concentrated sample present the most important challenges to water quality monitoring laboratories. Despite the advancements in pathogen identification, current diagnostic methods have limitations including laborious sample preparation, bulky instrumentation, and slow data readout. Additionally, field-deployable or “point-of-interest” systems are urgently needed in order to facilitate detection of pathogens even in remote area before spreading these pathogens through the public water systems. Functionalized magnetic particles play significant roles in both the sample preparation and detection processes. The great advances in magnetic particles synthesis, coating, high spatial resolution manipulation accompanied with immune-based or molecular based techniques and advance optics coupled with microfluidic technology offer a great opportunity to realize an integrated system of pathogen detection.This talk highlights our research towards the development of a magneto-fluidic integrated detection system for waterborne pathogen detection, emphasizing the role of functionalized magnetic particles as the gold standard tool in sample preparation and detection processes.

BiographyQasem Ramadan received his PhD in Bio-Microelectromechanical systems at Nanyang Technical University in 2006. From 2004 to 2008 he was a senior research engineer at the A-Star institute of microelectronics/Singapore. Since 2008, he is a research scientist at the Swiss Federal institute of Technology (Lausanne)

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Qasem Ramadan, J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05


http://dx.doi.org/10.4172/2155-6210.S1.05





Improving the PZT piezoelectric resonant sensors through finite-element simulationJang-Zern TsaiNational Central University, Taiwan

Miniaturized piezoelectric resonators of sizes 50 × 50, 100 × 100, 250 × 250, and 400 × 400 µm2 with 600-nm thickness have been made on silicon substrates. These sensors take sandwich structure with gold as the upper layer, platinum/titanium as

the bottom layer, and PZT (lead zirconate titanate) as the middle layer. The sensor has been extensively characterized in terms of surface morphology, atomic composition, crystalline structure, leakage current, ferroelectric tunability, etc. The impedance spectroscopy shows good resonant characteristics. It is found that the size and aspect ratio of the sensor may have significant effects on the resonant property and sensing ability. This research uses the finite-element method (FEM) to direct further improvement of the PZT resonator. Displacement, stress, strain, and oscillation modes of the PZT chip will all be simulated with FEM. The electrical impedance of the sensor simulated with harmonic analysis is compared with that measured by an impedance analyzer. Sinusoidal waves of various frequencies are applied across the upper and bottom layers and the change in the shape of the PZT layer is observed. The frequency at which a PZT has the maximum deformation is recorded as the resonant frequency, which ranged from several MHz to several hundred MHz in our simulation. From the simulation results, the resonant frequency is found to decrease with increased PZT area, which conforms to piezoelectric theory. The improvement in the sensitivity of the resonant chip is aimed for sensitive microbalance measurement of bio-molecules, such as microalbumin from the urine.

BiographyJang-Zern Tsai is an assistant professor of electrical engineering in National Central University, Jung-Li City, Taiwan. He received the BS degree in electrical engineering from National Central University, Taiwan, the MS degree in electrical engineering from National Tsing Hua University, Taiwan, and the PhD degree in electrical engineering from the University of Wisconsin-Madison, Wisconsin, USA. His current research interests include biochip design, biomedical instrumentation, biomedical signal processing, and bio-optics.

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Jang-Zern Tsai, J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05


http://dx.doi.org/10.4172/2155-6210.S1.05





Multifunctional nanoparticles for rapid bacterial capture, detection and decontamination Jin ZhangDepartment of Chemical & Biochemical Engineering, University of Western Ontario, Canada

This research project aims at developing the magnetic fluorescent nanoparticles to quickly capture, detect, and kill bacteria. Engineered nanoparticles (NPs) with multifunctional properties may interact with eukaryotic cells as drug carriers or contrast

agents. The interaction between multifunctional NPs and singled microorganisms, e.g. bacteria, however, remains unclear due to their special structure of cell wall. Here, we demonstrate that gentamicin (Gm)-bioconjugating fluorescent and magnetic NPs (FMNPs) capture gram-negative bacteria, e.g. Escherichia coli, E. coli, (1x107 CFU mL-1 from 10 mL of solution) within 20 min. Our findings indicate: FMNPs (diameter = 65±8 nm) alone inert to E. coli, whereas Gm-FMNPs disrupt the cell wall of E. coli first through the ionic bonding between GM and lipopolysaccharide(LPS) on the outer membrane of E. coli; a low concentration of E. coli, ~1x103 CFU mL-1, dramatically changes the fluorescent signal of Gm-FMNPs; the antimicrobial efficiency of Gm for E. coli is improved by ~20% through the conjugation with FMNPs. It indicates the Gm-conjugating FMNPs are able to be a multifunctional platform for bacterial capture, detection, and decontamination simultaneously.

BiographyJin Zhang received her PhD from the National University of Singapore (NUS) in 2003. Dr. Zhang is now an Assistant Professor of the Department of Chemical and Biochemical Engineering (CBE) at the University of Western Ontario (Western), and an adjunct professor of the Schulich School of Medicine & Dentistry at Western. She has published 25 peer-reviewed journal papers and 14 peer-reviewed proceedings. She has co-authored a book chapter with the publisher Kluwer. As co-inventor, she has two filed patents and one-provisional patent application. Jin was recently recognized as the Grand Challenges Canada-Canadian Rising Stars in Global Health for her research work on “Non-invasive Diagnostic Tool for Diabetes”.

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Jin Zhang, J Biosens Bioelectron, 2:4http://dx.doi.org/10.4172/2155-6210.S1.05


http://dx.doi.org/10.4172/2155-6210.S1.05

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