The problem of rapidly detecting pathogens directly from clinical samples poses significant analy... more The problem of rapidly detecting pathogens directly from clinical samples poses significant analytical challenges. Addressing this issue in relation to urinary tract infections, we propose an effective protocol and related immunomagnetic test kits enabling versatile screening for the presence of pathogenic bacteria in unprocessed urine samples. To achieve this, the components of a typical immunomagnetic separation protocol were optimized towards the sensitive assessment of the aggregates formed out of immunomagnetically tagged target pathogens collected from clinical samples. Specifically, a dedicated immunomagnetic material was developed via the functionalization of standardized, micron-sized magnetic beads with generic antibodies against gram-specific bacterial constituents with mannan binding lectin. As such, we demonstrate efficient procedures for achieving the enhanced, specific, and pathogen-mediated cluster formation of these tailored affinity-coated magnetic beads in complex...
Cross linked gold-dynamic constitutional frameworks (DCFs) are functional materials of potential ... more Cross linked gold-dynamic constitutional frameworks (DCFs) are functional materials of potential relevance for biosensing applications, given their adaptivity and high responsivity against various external stimuli (such as pH, temperature) or specific interactions with biomolecules (enzymes or DNA) via internal constitutional dynamics. However, characterization and assessment of their dynamic conformational changes in response to external stimuli has never been reported. This study proves the capability of Surface Plasmon Resonance (SPR) assays to analyse the adaptive structural modulation of a functional matrix encompassing 3D gold-dynamic constitutional frameworks (Au-DCFs) when exposed to pH variations, as external stimuli. We analyse Au-DCFs formed from Au nanoparticles, (AuNP) connected through constitutionally dynamic polymers, dynamers, with multiple functionalities. For increased generality of this proof-of-concept assay, Au-DCFs, involving DCFs designed from 1,3,5-benzene-t...
Ovidius la Tomis: satul global și vocile exegezei actuale Ovidius at Tomis: The Global Village an... more Ovidius la Tomis: satul global și vocile exegezei actuale Ovidius at Tomis: The Global Village and the Voices of the Contemporary Exegesis Sabina ...
If the analyte does not only change the electrochemical but also the optical properties of the el... more If the analyte does not only change the electrochemical but also the optical properties of the electrode/solution interface, the spatial resolution of an electrochemical sensor can be substantially enhanced by combining the electrochemical sensor with optical microscopy. In order to demonstrate this, electrochemical biosensors for the detection of hydrogen peroxide and glucose were developed by drop casting enzyme and redox polymer mixtures onto planar, optically transparent electrodes. These biosensors generate current signals proportional to the analyte concentration via a reaction sequence which ultimately changes the oxidation state of the redox polymer. Images of the interface of these biosensors were acquired using bright field reflected light microscopy (BFRLM). Analysis showed that the intensity of these images is higher when the redox polymer is oxidized than when it is reduced. It also revealed that the time needed for the redox polymer to change oxidation state can be ass...
2011 7TH INTERNATIONAL SYMPOSIUM ON ADVANCED TOPICS IN ELECTRICAL ENGINEERING (ATEE), 2011
Development of a novel magneto-optic surface-plasmon-resonance (MOSPR) platform and its use for t... more Development of a novel magneto-optic surface-plasmon-resonance (MOSPR) platform and its use for the detection of biomolecules are presented. The physical transduction principle is based on the combination of the magneto-optic activity of magnetic materials and a surface-plasmon resonance of metallic layers. The structure of layers has been optimized using a Transfer Matrix approach. Hints on deposition of thin layers of Cr, Au and Co will be outlined. Such combination can produce a significant enhancement of the magneto-optic effects that strongly depends on the optical properties of the surrounding medium, allowing its use for biosensing applications. Besides the magento-plasmonic sensor, the platform comprises the surface plasmon resonance module, the electromagnet with oscillating magnetic field (with controlled field strength and frequency) for MOSPR and the flow cell with integrated microfluidics. Calibration curves based on solutions with different refractive indices show a st...
L'invention concerne un procede et un systeme servant a detecter la presence et/ou a determin... more L'invention concerne un procede et un systeme servant a detecter la presence et/ou a determiner la concentration de certaines substances a analyser. Le procede selon l'invention consiste a recevoir dans une chambre de mesure un echantillon contenant une substance a analyser cible, la chambre de mesure comprenant une paire d'electrodes (42a-b) dont au moins une (42a) est compatible avec des mesures optiques, agissant comme capteur photosensible connecte a un analyseur optique (32); a appliquer un champ d'activation oscillant (par ex. magnetique, electromagnetique, mecanique) a la chambre de mesure (12), le champ applique oscillant a une premiere frequence; a mesurer une impedance electrique cible de la paire d'electrodes (42a-b), l'impedance indiquant un changement de position de la substance a analyser cible (60) en reponse a l'application du champ d'activation oscillant, la mesure de l'impedance electrique cible consistant a appliquer un signal e...
Cell culture assays have become a routine testing method in the medical and pharmaceutical resear... more Cell culture assays have become a routine testing method in the medical and pharmaceutical research. Real-time monitoring of cells on a substrate using electrical impedance is a convenient and non-invasive method of analysis. Cell proliferation and apoptosis can be quantified and characterized by electrical impedance in a time-dependent manner. Unfortunately monitoring several cell cultures at the same time is challenging because of the prohibitive costs of impedance analysis equipment or rather slow response if multiplexing the front-end of an impedance analyzer. We propose a low cost high performance impedance analyzer based on a two channel delta-sigma 16-bit ADC. The device can handle frequencies between 30 Hz and 10 kHz with input impedances ranging: 100mOhms - 10Mohms. The Automatic Level Control (ALC) algorithm allows for steady non-invasive voltage levels on the measured impedance independent of measuring frequency and impedance value. The device prototype performance is com...
ABSTRACT We present an advanced system for monitoring the behavior of a fish population in an exp... more ABSTRACT We present an advanced system for monitoring the behavior of a fish population in an experimental tank and derive a set of characteristic features pertaining to their dynamics. Using a 3-D array of ultrasound transducers, the system provides time series of fish positions, allowing for assessment of the dynamics of both the geometric centre (centroid) and the mean spatial extension of the fish population. We reveal that evolution of the mean spatial extension of fish population and the related standard deviation represents a sensitive indicator of fish behavior in respect to the quality of the aquatic environment. Emphasis has been given to reversible effects i.e. whenever water parameters returned to normal (physiological) levels, fish behavior exhibits similar dynamical features. Whereas fish behavior was previously assessed by several monitoring techniques (e.g. electrical impedance, video recording, hydro-acoustics etc.), this study reveals the advantages of using ultrasounds to continuously monitor the three-dimensional position of fish, having also the quality of being independent on water turbidity or light intensity.
The present work compares the responses of a model bioaffinity sensor based on a dielectric funct... more The present work compares the responses of a model bioaffinity sensor based on a dielectric functionalization layer, in terms of specific and nonspecific binding, when interrogated simultaneously by Surface Plasmon Resonance (SPR), non-Faradaic Electrochemical Impedance Spectroscopy (EIS), and Plasmonic based-EIS (P-EIS). While biorecognition events triggered a sensitive SPR signal, the related EIS response was rather negligible. Contrarily, even a limited nonspecific adsorption onto the surface of the metallic electrode, allowed by the intrinsic imperfect compactness of the functionalization layers, was signaled by EIS and not by SPR. The source of this finding has been addressed from both theoretical and experimental perspectives, demonstrating that EIS signals are mainly sensitive to adsorptions that alter the current pathway through defects of the functionalization layer exposing the electrode. These observations are of importance for those developing biosensors analyzed by SPR,...
We present novel solutions to surpass current analytic limitations of Magneto-Optical Surface Pla... more We present novel solutions to surpass current analytic limitations of Magneto-Optical Surface Plasmon Resonance (MOSPR) assays, concerning both the chip structure and the method for data analysis. The structure of the chip is modified to contain a thin layer of Co-Au alloy instead of successive layers of homogeneous metals, as currently used. This alloy presents improved plasmonic and magnetic properties, yet a structural stability similar to Au-SPR chips, allowing for bioaffinity assays in saline solutions. Analyzing the whole reflectivity curve at multiple angles of incidence instead of the reflectivity value at a single incidence angle provides a high signal-to-noise ratio suitable for detection of minute analyte concentrations. Based on assessment of solutions with known refractive indices as well as of a model biomolecular interaction (i.e. IgG-AntiIgG) we demonstrate that the proposed structure of the MOSPR sensing chip and the procedure of data analysis allows for long-time a...
ABSTRACT The capabilities of differential impedance spectroscopy to assess bioaffinity sensors (w... more ABSTRACT The capabilities of differential impedance spectroscopy to assess bioaffinity sensors (with the surface modified with high specificity compounds) for both direct and competitive determinations were investigated using a novel Dual Surface Plasmon Resonance (SPR) - Differential Electrical Impedance (DEI) measurement set-up, aiming for development of an effective assay for detection of target analytes. The surface changes due to the specific recognition events are simultaneously monitored, on the surface of the same chip, using this custom designed system that comprises: a 3 channel SPR module, a multi-channel, differential impedance analyzer, a fully automated injection system (with capabilities for manual injecting and programmed protocols) and a 3 micro channels flow through measurement chamber. In this format, the variations of impedance of the sensor/specimen interface in response to fast specific recognition events (Ag-Ab) are revealed, while eliminating the nonspecific influences; simultaneous SPR & DEI measurements provide inner validation and expand the analyte detection range.
Although modeling and experimental approaches to probe antimicrobial peptides-lipid membranes int... more Although modeling and experimental approaches to probe antimicrobial peptides-lipid membranes interaction have already been reported, quantitative evaluation of the whole process, including full dissolution of the lipid, is still missing. We report on the real-time assessment of the entire set of stages of melittin-membrane interaction, based on surface plasmon resonance (SPR) measurements, using supported lipid matrices on L1 sensors and long peptide injections. We advance a mathematical model which comprises a set of coupled kinetic equations and relates via the transfer matrix the evolution of lipid and peptide concentrations with the SPR sensorgram. Upon fitting the sensorgrams of melittin injections on POPC lipid matrices, in agreement with literature data, the model provides: association and dissociation rates, concentration thresholds, and evolution within each interacting layer of lipid and peptide concentrations as well as of peptide to lipid ratios. The proposed model combined with appropriate experimental protocols adds new depths to SPR investigation of peptide-lipid interaction offering a quantitative platform for research and controlled design of improved antimicrobial peptides. A wider applicability for quantitative assessment of other pore forming compounds on different lipid matrices is suggested.
Surface quality of the Surface Plasmon Resonance (SPR) chips is a major limiting issue in most SP... more Surface quality of the Surface Plasmon Resonance (SPR) chips is a major limiting issue in most SPR analyses, even more for supported lipid membranes experiments, where both the organization of the lipid matrix and the subsequent incorporation of the target molecule depend on the surface quality. A novel quantitative method to characterize the quality of SPR sensors chips is described for L1 chips subject to formation of lipid films, injection of membrane disrupting compounds, followed by appropriate regeneration procedures. The method consists in analysis of the SPR reflectivity curves for several standard solutions (e.g. PBS, HEPES or deionized water). This analysis reveals the decline of sensor surface as a function of the number of experimental cycles (consisting in biosensing assay and regeneration step) and enables active control of surface regeneration for enhanced reproducibility. We demonstrate that quantitative evaluation of the changes in reflectivity curves (shape of the SPR dip) and of the slope of the calibration curve provides a rapid and effective procedure for surface quality assessment. Whereas the method was tested on L1 SPR sensors chips, we stress on its amenability to assess the quality of other types of SPR chips, as well.
The problem of rapidly detecting pathogens directly from clinical samples poses significant analy... more The problem of rapidly detecting pathogens directly from clinical samples poses significant analytical challenges. Addressing this issue in relation to urinary tract infections, we propose an effective protocol and related immunomagnetic test kits enabling versatile screening for the presence of pathogenic bacteria in unprocessed urine samples. To achieve this, the components of a typical immunomagnetic separation protocol were optimized towards the sensitive assessment of the aggregates formed out of immunomagnetically tagged target pathogens collected from clinical samples. Specifically, a dedicated immunomagnetic material was developed via the functionalization of standardized, micron-sized magnetic beads with generic antibodies against gram-specific bacterial constituents with mannan binding lectin. As such, we demonstrate efficient procedures for achieving the enhanced, specific, and pathogen-mediated cluster formation of these tailored affinity-coated magnetic beads in complex...
Cross linked gold-dynamic constitutional frameworks (DCFs) are functional materials of potential ... more Cross linked gold-dynamic constitutional frameworks (DCFs) are functional materials of potential relevance for biosensing applications, given their adaptivity and high responsivity against various external stimuli (such as pH, temperature) or specific interactions with biomolecules (enzymes or DNA) via internal constitutional dynamics. However, characterization and assessment of their dynamic conformational changes in response to external stimuli has never been reported. This study proves the capability of Surface Plasmon Resonance (SPR) assays to analyse the adaptive structural modulation of a functional matrix encompassing 3D gold-dynamic constitutional frameworks (Au-DCFs) when exposed to pH variations, as external stimuli. We analyse Au-DCFs formed from Au nanoparticles, (AuNP) connected through constitutionally dynamic polymers, dynamers, with multiple functionalities. For increased generality of this proof-of-concept assay, Au-DCFs, involving DCFs designed from 1,3,5-benzene-t...
Ovidius la Tomis: satul global și vocile exegezei actuale Ovidius at Tomis: The Global Village an... more Ovidius la Tomis: satul global și vocile exegezei actuale Ovidius at Tomis: The Global Village and the Voices of the Contemporary Exegesis Sabina ...
If the analyte does not only change the electrochemical but also the optical properties of the el... more If the analyte does not only change the electrochemical but also the optical properties of the electrode/solution interface, the spatial resolution of an electrochemical sensor can be substantially enhanced by combining the electrochemical sensor with optical microscopy. In order to demonstrate this, electrochemical biosensors for the detection of hydrogen peroxide and glucose were developed by drop casting enzyme and redox polymer mixtures onto planar, optically transparent electrodes. These biosensors generate current signals proportional to the analyte concentration via a reaction sequence which ultimately changes the oxidation state of the redox polymer. Images of the interface of these biosensors were acquired using bright field reflected light microscopy (BFRLM). Analysis showed that the intensity of these images is higher when the redox polymer is oxidized than when it is reduced. It also revealed that the time needed for the redox polymer to change oxidation state can be ass...
2011 7TH INTERNATIONAL SYMPOSIUM ON ADVANCED TOPICS IN ELECTRICAL ENGINEERING (ATEE), 2011
Development of a novel magneto-optic surface-plasmon-resonance (MOSPR) platform and its use for t... more Development of a novel magneto-optic surface-plasmon-resonance (MOSPR) platform and its use for the detection of biomolecules are presented. The physical transduction principle is based on the combination of the magneto-optic activity of magnetic materials and a surface-plasmon resonance of metallic layers. The structure of layers has been optimized using a Transfer Matrix approach. Hints on deposition of thin layers of Cr, Au and Co will be outlined. Such combination can produce a significant enhancement of the magneto-optic effects that strongly depends on the optical properties of the surrounding medium, allowing its use for biosensing applications. Besides the magento-plasmonic sensor, the platform comprises the surface plasmon resonance module, the electromagnet with oscillating magnetic field (with controlled field strength and frequency) for MOSPR and the flow cell with integrated microfluidics. Calibration curves based on solutions with different refractive indices show a st...
L'invention concerne un procede et un systeme servant a detecter la presence et/ou a determin... more L'invention concerne un procede et un systeme servant a detecter la presence et/ou a determiner la concentration de certaines substances a analyser. Le procede selon l'invention consiste a recevoir dans une chambre de mesure un echantillon contenant une substance a analyser cible, la chambre de mesure comprenant une paire d'electrodes (42a-b) dont au moins une (42a) est compatible avec des mesures optiques, agissant comme capteur photosensible connecte a un analyseur optique (32); a appliquer un champ d'activation oscillant (par ex. magnetique, electromagnetique, mecanique) a la chambre de mesure (12), le champ applique oscillant a une premiere frequence; a mesurer une impedance electrique cible de la paire d'electrodes (42a-b), l'impedance indiquant un changement de position de la substance a analyser cible (60) en reponse a l'application du champ d'activation oscillant, la mesure de l'impedance electrique cible consistant a appliquer un signal e...
Cell culture assays have become a routine testing method in the medical and pharmaceutical resear... more Cell culture assays have become a routine testing method in the medical and pharmaceutical research. Real-time monitoring of cells on a substrate using electrical impedance is a convenient and non-invasive method of analysis. Cell proliferation and apoptosis can be quantified and characterized by electrical impedance in a time-dependent manner. Unfortunately monitoring several cell cultures at the same time is challenging because of the prohibitive costs of impedance analysis equipment or rather slow response if multiplexing the front-end of an impedance analyzer. We propose a low cost high performance impedance analyzer based on a two channel delta-sigma 16-bit ADC. The device can handle frequencies between 30 Hz and 10 kHz with input impedances ranging: 100mOhms - 10Mohms. The Automatic Level Control (ALC) algorithm allows for steady non-invasive voltage levels on the measured impedance independent of measuring frequency and impedance value. The device prototype performance is com...
ABSTRACT We present an advanced system for monitoring the behavior of a fish population in an exp... more ABSTRACT We present an advanced system for monitoring the behavior of a fish population in an experimental tank and derive a set of characteristic features pertaining to their dynamics. Using a 3-D array of ultrasound transducers, the system provides time series of fish positions, allowing for assessment of the dynamics of both the geometric centre (centroid) and the mean spatial extension of the fish population. We reveal that evolution of the mean spatial extension of fish population and the related standard deviation represents a sensitive indicator of fish behavior in respect to the quality of the aquatic environment. Emphasis has been given to reversible effects i.e. whenever water parameters returned to normal (physiological) levels, fish behavior exhibits similar dynamical features. Whereas fish behavior was previously assessed by several monitoring techniques (e.g. electrical impedance, video recording, hydro-acoustics etc.), this study reveals the advantages of using ultrasounds to continuously monitor the three-dimensional position of fish, having also the quality of being independent on water turbidity or light intensity.
The present work compares the responses of a model bioaffinity sensor based on a dielectric funct... more The present work compares the responses of a model bioaffinity sensor based on a dielectric functionalization layer, in terms of specific and nonspecific binding, when interrogated simultaneously by Surface Plasmon Resonance (SPR), non-Faradaic Electrochemical Impedance Spectroscopy (EIS), and Plasmonic based-EIS (P-EIS). While biorecognition events triggered a sensitive SPR signal, the related EIS response was rather negligible. Contrarily, even a limited nonspecific adsorption onto the surface of the metallic electrode, allowed by the intrinsic imperfect compactness of the functionalization layers, was signaled by EIS and not by SPR. The source of this finding has been addressed from both theoretical and experimental perspectives, demonstrating that EIS signals are mainly sensitive to adsorptions that alter the current pathway through defects of the functionalization layer exposing the electrode. These observations are of importance for those developing biosensors analyzed by SPR,...
We present novel solutions to surpass current analytic limitations of Magneto-Optical Surface Pla... more We present novel solutions to surpass current analytic limitations of Magneto-Optical Surface Plasmon Resonance (MOSPR) assays, concerning both the chip structure and the method for data analysis. The structure of the chip is modified to contain a thin layer of Co-Au alloy instead of successive layers of homogeneous metals, as currently used. This alloy presents improved plasmonic and magnetic properties, yet a structural stability similar to Au-SPR chips, allowing for bioaffinity assays in saline solutions. Analyzing the whole reflectivity curve at multiple angles of incidence instead of the reflectivity value at a single incidence angle provides a high signal-to-noise ratio suitable for detection of minute analyte concentrations. Based on assessment of solutions with known refractive indices as well as of a model biomolecular interaction (i.e. IgG-AntiIgG) we demonstrate that the proposed structure of the MOSPR sensing chip and the procedure of data analysis allows for long-time a...
ABSTRACT The capabilities of differential impedance spectroscopy to assess bioaffinity sensors (w... more ABSTRACT The capabilities of differential impedance spectroscopy to assess bioaffinity sensors (with the surface modified with high specificity compounds) for both direct and competitive determinations were investigated using a novel Dual Surface Plasmon Resonance (SPR) - Differential Electrical Impedance (DEI) measurement set-up, aiming for development of an effective assay for detection of target analytes. The surface changes due to the specific recognition events are simultaneously monitored, on the surface of the same chip, using this custom designed system that comprises: a 3 channel SPR module, a multi-channel, differential impedance analyzer, a fully automated injection system (with capabilities for manual injecting and programmed protocols) and a 3 micro channels flow through measurement chamber. In this format, the variations of impedance of the sensor/specimen interface in response to fast specific recognition events (Ag-Ab) are revealed, while eliminating the nonspecific influences; simultaneous SPR & DEI measurements provide inner validation and expand the analyte detection range.
Although modeling and experimental approaches to probe antimicrobial peptides-lipid membranes int... more Although modeling and experimental approaches to probe antimicrobial peptides-lipid membranes interaction have already been reported, quantitative evaluation of the whole process, including full dissolution of the lipid, is still missing. We report on the real-time assessment of the entire set of stages of melittin-membrane interaction, based on surface plasmon resonance (SPR) measurements, using supported lipid matrices on L1 sensors and long peptide injections. We advance a mathematical model which comprises a set of coupled kinetic equations and relates via the transfer matrix the evolution of lipid and peptide concentrations with the SPR sensorgram. Upon fitting the sensorgrams of melittin injections on POPC lipid matrices, in agreement with literature data, the model provides: association and dissociation rates, concentration thresholds, and evolution within each interacting layer of lipid and peptide concentrations as well as of peptide to lipid ratios. The proposed model combined with appropriate experimental protocols adds new depths to SPR investigation of peptide-lipid interaction offering a quantitative platform for research and controlled design of improved antimicrobial peptides. A wider applicability for quantitative assessment of other pore forming compounds on different lipid matrices is suggested.
Surface quality of the Surface Plasmon Resonance (SPR) chips is a major limiting issue in most SP... more Surface quality of the Surface Plasmon Resonance (SPR) chips is a major limiting issue in most SPR analyses, even more for supported lipid membranes experiments, where both the organization of the lipid matrix and the subsequent incorporation of the target molecule depend on the surface quality. A novel quantitative method to characterize the quality of SPR sensors chips is described for L1 chips subject to formation of lipid films, injection of membrane disrupting compounds, followed by appropriate regeneration procedures. The method consists in analysis of the SPR reflectivity curves for several standard solutions (e.g. PBS, HEPES or deionized water). This analysis reveals the decline of sensor surface as a function of the number of experimental cycles (consisting in biosensing assay and regeneration step) and enables active control of surface regeneration for enhanced reproducibility. We demonstrate that quantitative evaluation of the changes in reflectivity curves (shape of the SPR dip) and of the slope of the calibration curve provides a rapid and effective procedure for surface quality assessment. Whereas the method was tested on L1 SPR sensors chips, we stress on its amenability to assess the quality of other types of SPR chips, as well.
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Papers by Eugen Gheorghiu