Publicaciones del IMSE

Encontrados resultados para:

Autor: Alberto Olmo Fernández
Año: Desde 2002

Artículos de revistas


The Use of High-Intensity Focused Ultrasound for the Rewarming of Cryopreserved Biological Material
A. Olmo, P. Barroso, F. Barroso and R. Risco
Journal Paper · IEEE Transactions on Ultrasonics Ferroelectrics and Frequency Control, vol. 68, no. 3, pp 599-607, 2021
resumen      doi      

High-intensity focused ultrasound (HIFU) has been used in different medical applications in the last years. In this work, we present for the first time the use of HIFU in the field of cryopreservation, the preservation of biological material at low temperatures. An HIFU system has been designed with the objective of achieving a fast and uniform rewarming in organs, key to overcome the critical problem of devitrification. The finite-element simulations have been carried out using COMSOL Multiphysics software. An array of 26 ultrasonic transducers was simulated, achieving an HIFU focal area in the order of magnitude of a model organ (ovary). A parametric study of the warming rate and temperature gradients, as a function of the frequency and power of ultrasonic waves, was performed. An optimal value for these parameters was found. The results validate the appropriateness of the technique, which is of utmost importance for the future creation of cryopreserved organ banks.

Characterization and Monitoring of Titanium Bone Implants with Impedance Spectroscopy
A. Olmo, M. Hernandez, E. Chicardi and Y. Torres
Journal Paper · Sensors, vol. 20, no. 19, article 4358, 2020
resumen      doi      

Porous titanium is a metallic biomaterial with good properties for the clinical repair of cortical bone tissue, although the presence of pores can compromise its mechanical behavior and clinical use. It is therefore necessary to characterize the implant pore size and distribution in a suitable way. In this work, we explore the new use of electrical impedance spectroscopy for the characterization and monitoring of titanium bone implants. Electrical impedance spectroscopy has been used as a non-invasive route to characterize the volumetric porosity percentage (30%, 40%, 50% and 60%) and the range of pore size (100-200 and 355-500 mm) of porous titanium samples obtained with the space-holder technique. Impedance spectroscopy is proved to be an appropriate technique to characterize the level of porosity of the titanium samples and pore size, in an affordable and non-invasive way. The technique could also be used in smart implants to detect changes in the service life of the material, such as the appearance of fractures, the adhesion of osteoblasts and bacteria, or the formation of bone tissue.

3D-printed sensors and actuators in cell culture and tissue engineering: Framework and research challenges
P. Pérez, J.A. Serrano and A. Olmo
Journal Paper · Sensors, vol. 20, no. 19, article 5617, 2020
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Three-dimensional printing technologies have been recently proposed to monitor cell cultures and implement cell bioreactors for different biological applications. In tissue engineering, the control of tissue formation is crucial to form tissue constructs of clinical relevance, and 3D printing technologies can also play an important role for this purpose. In this work, we study 3D-printed sensors that have been recently used in cell culture and tissue engineering applications in biological laboratories, with a special focus on the technique of electrical impedance spectroscopy. Furthermore, we study new 3D-printed actuators used for the stimulation of stem cells cultures, which is of high importance in the process of tissue formation and regenerative medicine. Key challenges and open issues, such as the use of 3D printing techniques in implantable devices for regenerative medicine, are also discussed.

Use of Impedance Spectroscopy for the Characterization of In-Vitro Osteoblast Cell Response in Porous Titanium Bone Implants
M. Giner, A. Olmo, M. Hernandez, P. Trueba, E. Chicardi, A. Civantos, M.A. Vazquez, M.J. Montoya-Garcia and Y. Torres
Journal Paper · Metals, vol. 10, no. 8, article 1077, 2020
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The use of titanium implants with adequate porosity (content, size and morphology) could solve the stress shielding limitations that occur in conventional titanium implants. Experiments to assess the cellular response (adhesion, proliferation and differentiation of osteoblasts) on implants are expensive, time-consuming and delicate. In this work, we propose the use of impedance spectroscopy to evaluate the growth of osteoblasts on porous titanium implants. Osteoblasts cells were cultured on fully-dense and 40 vol.% porous discs with two ranges of pore size (100-200 mu m and 355-500 mu m) to study cell viability, proliferation, differentiation (Alkaline phosphatase activity) and cell morphology. The porous substrates 40 vol.% (100-200 mu m) showed improved osseointegration response as achieved more than 80% of cell viability and higher levels of Cell Differentiation by Alkaline Phosphatase (ALP) at 21 days. This cell behavior was further evaluated observing an increase in the impedance modulus for all study conditions when cells were attached. However, impedance levels were higher on fully-dense due to its surface properties (flat surface) than porous substrates (flat and pore walls). Surface parameters play an important role on the global measured impedance. Impedance is useful for characterizing cell cultures in different sample types.

Electrical modeling of the growth and differentiation of skeletal myoblasts cell cultures for tissue engineering
A. Olmo, Y. Yuste, J.A. Serrano, A. Maldonado-Jacobi, P. Pérez, G. Huertas, S, Pereira, A. Yufera and F. de la Portilla
Journal Paper · Sensors, vol. 20, no. 11, article 3152, 2020
resumen      doi      pdf

In tissue engineering, of utmost importance is the control of tissue formation, in order to form tissue constructs of clinical relevance. In this work, we present the use of an impedance spectroscopy technique for the real-time measurement of the dielectric properties of skeletal myoblast cell cultures. The processes involved in the growth and differentiation of these cell cultures in skeletal muscle are studied. A circuit based on the oscillation-based test technique was used, avoiding the use of high-performance circuitry or external input signals. The effect of electrical pulse stimulation applied to cell cultures was also studied. The technique proved useful for monitoring in real-time the processes of cell growth and estimating the fill factor of muscular stem cells. Impedance spectroscopy was also useful to study the real-time monitoring of cell differentiation, obtaining different oscillation amplitude levels for differentiated and undifferentiated cell cultures. Finally, an electrical model was implemented to better understand the physical properties of the cell culture and control the tissue formation process.

Evaluation of Implanted Stent Occlusion Status Based on Neointimal Tissue Bioimpedance Simulations
J.M. Portillo-Anaya, P. Perez, A. Olmo, G. Huertas and A. Yufera
Journal Paper · Journal of Sensors, vol. 2019, article 7167186, 2019
resumen      doi      

This paper describes the characterization of the light hole, also known as the lumen, in implanted stents affected by restenosis processes using bioimpedance (BI) as a biomarker. The presented approach will enable real-time monitoring of lumens in implanted stents. The basis of the work hereby reported is the fact that neointimal tissues involved in restenosis can be detected and measured through their impedance properties, namely, conductivity and permittivity. To exploit these properties, a 4-electrode setup for BI measurement is proposed. This setup allows study of the influence of the various tissues involved in restenosis fat, muscle, fibre, and endothelium, together with the blood, on the BI value at several frequencies. In addition, BI simulation tests were performed using the electric physics module available in COMSOL Multiphysics®. Interestingly, fat constitutes the most influential layer on the value of impedance (measured in kΩ/μm-magnitude change per micrometre of lumen occlusion). A case study using a standard stent is also presented. In this study, where the involved tissues and blood were simultaneously considered, we conducted an analysis for stable and vulnerable plaques in restenosis test situations. In this regard, the proposed method is useful to test the stent obstruction and detect potential dangerous cases due to nonstable fat accumulation.

Data-Analytics Modeling of Electrical Impedance Measurements for Cell Culture Monitoring
E. García, P. Pérez, A. Olmo, R. Díaz, G. Huertas and A. Yúfera
Journal Paper · Sensors, vol. 19, no. 21, art. 4639, 2019
resumen      doi      pdf

High-throughput data analysis challenges in laboratory automation and lab-on-a-chip devices´ applications are continuously increasing. In cell culture monitoring, specifically, the electrical cell-substrate impedance sensing technique (ECIS), has been extensively used for a wide variety of applications. One of the main drawbacks of ECIS is the need for implementing complex electrical models to decode the electrical performance of the full system composed by the electrodes, medium, and cells. In this work we present a new approach for the analysis of data and the prediction of a specific biological parameter, the fill-factor of a cell culture, based on a polynomial regression, data-analytic model. The method was successfully applied to a specific ECIS circuit and two different cell cultures, N2A (a mouse neuroblastoma cell line) and myoblasts. The data-analytic modeling approach can be used in the decoding of electrical impedance measurements of different cell lines, provided a representative volume of data from the cell culture growth is available, sorting out the difficulties traditionally found in the implementation of electrical models. This can be of particular importance for the design of control algorithms for cell cultures in tissue engineering protocols, and labs-on-a-chip and wearable devices applications.

Electrical pulse stimulation of skeletal myoblasts cell cultures with simulated action potentials
P. Villanueva, S. Pereira, A. Olmo, P. Pérez, Y. Yuste, A. Yúfera and F. de la Portilla
Journal Paper · Journal of Tissue Engineering and Regenerative Medicine, vol. 13, no. 7, pp 1265-1269, 2019
resumen      doi      

Electrical pulse stimulation has an important effect on skeletal muscle development and maturation. However, the methodology for controlling these stimulation parameters to develop in vitro functional skeletal muscle tissues remains to be established. In this work, we have studied the effect of simulated action potentials on the growth and differentiation of skeletal myoblast cell cultures. A circuit simulating action potentials of 0.15 and 0.3 V/mm, at a frequency of 1 Hz and with a 4-ms pulse width, is proposed. Results show an important improvement of the growth rate and differentiation of myoblasts at a voltage of 0.15 V/mm. Parameters such as electrodes geometry or type of signals must be considered in the development of in vitro skeletal muscle.

Remote Cell Growth Sensing using Self-Sustained Bio-Oscillations
P. Pérez, G. Huertas, A. Olmo, A. Maldonado-Jacobi, J. Serrano, M. Martín, P. Daza and A. Yúfera
Journal Paper · Sensors, vol. 18, no. 8, art. 2550, 2018
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A smart sensor system for cell culture real-time supervision is proposed, allowing for a significant reduction in human effort applied to this type of assay. The approach converts the cell culture under test into a suitable "biological" oscillator. The system enables the remote acquisition and management of the "biological" oscillation signals through a secure web interface. The indirectly observed biological properties are cell growth and cell number, which are straightforwardly related to the measured bio-oscillation signal parameters, i.e., frequency and amplitude. The sensor extracts the information without complex circuitry for acquisition and measurement, taking advantage of the microcontroller features. A discrete prototype for sensing and remote monitoring is presented along with the experimental results obtained from the performed measurements, achieving the expected performance and outcomes.

An empirical-mathematical approach for calibration and fitting cell-electrode electrical models in bioimpedance tests
J.A. Serrano, G. Huertas, A. Maldonado-Jacobi, A. Olmo, P. Pérez, M.E. Martín, P. Daza and A. Yúfera
Journal Paper · Sensors, vol. 18, no. 7, article 2354, 2018
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This paper proposes a new yet efficient method allowing a significant improvement in the on-line analysis of biological cell growing and evolution. The procedure is based on an empirical-mathematical approach for calibration and fitting of any cell-electrode electrical model. It is valid and can be extrapolated for any type of cellular line used in electrical cell-substrate impedance spectroscopy (ECIS) tests. Parameters of the bioimpedance model, acquired from ECIS experiments, vary for each cell line, which makes obtaining results difficult and -to some extent-renders them inaccurate. We propose a fitting method based on the cell line initial characterization, and carry out subsequent experiments with the same line to approach the percentage of well filling and the cell density (or cell number in the well). To perform our calibration technique, the so-called oscillation-based test (OBT) approach is employed for each cell density. Calibration results are validated by performing other experiments with different concentrations on the same cell line with the same measurement technique. Accordingly, a bioimpedance electrical model of each cell line is determined, which is valid for any further experiment and leading to a more precise electrical model of the electrode-cell system. Furthermore, the model parameters calculated can be also used by any other measurement techniques. Promising experimental outcomes for three different cell-lines have been achieved, supporting the usefulness of this technique.

Sensing Cell-Culture Assays with Low-Cost Circuitry
P. Pérez, G. Huertas, A. Maldonado-Jacobi, M. Martín, J.A. Serrano, A. Olmo, P. Daza and A. Yúfera
Journal Paper · Scientific Reports, vol. 8, no. 1, article 8841, 2018
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An alternative approach for cell-culture end-point protocols is proposed herein. This new technique is suitable for real-time remote sensing. It is based on Electrical Cell-substrate Impedance Spectroscopy (ECIS) and employs the Oscillation-Based Test (OBT) method. Simple and straightforward circuit blocks form the basis of the proposed measurement system. Oscillation parameters - frequency and amplitude - constitute the outcome, directly correlated with the culture status. A user can remotely track the evolution of cell cultures in real time over the complete experiment through a web tool continuously displaying the acquired data. Experiments carried out with commercial electrodes and a well-established cell line (AA8) are described, obtaining the cell number in real time from growth assays. The electrodes have been electrically characterized along the design flow in order to predict the system performance and the sensitivity curves. Curves for 1-week cell growth are reported. The obtained experimental results validate the proposed OBT for cell-culture characterization. Furthermore, the proposed electrode model provides a good approximation for the cell number and the time evolution of the studied cultures.

Real-time electrical bioimpedance characterization of neointimal tissue for stent applications
D. Rivas-Marchena, A. Olmo, J.A. Miguel, M. Martinez, G. Huertas and A. Yufera
Journal Paper · Sensors, vol. 17, no. 8, art. 1737, 2017
resumen      doi      pdf

To follow up the restenosis in arteries stented during an angioplasty is an important current clinical problem. A new approach to monitor the growth of neointimal tissue inside the stent is proposed on the basis of electrical impedance spectroscopy (EIS) sensors and the oscillation-based test (OBT) circuit technique. A mathematical model was developed to analytically describe the histological composition of the neointima, employing its conductivity and permittivity data. The bioimpedance model was validated against a finite element analysis (FEA) using COMSOL Multiphysics software. A satisfactory correlation between the analytical model and FEA simulation was achieved in most cases, detecting some deviations introduced by the thin “double layer” that separates the neointima and the blood. It is hereby shown how to apply conformal transformations to obtain bioimpedance electrical models for stack-layered tissues over coplanar electrodes. Particularly, this can be applied to characterize the neointima in real-time. This technique is either suitable as a main mechanism for restenosis follow-up or it can be combined with proposed intelligent stents for blood pressure measurements to auto-calibrate the sensibility loss caused by the adherence of the tissue on the micro-electro-mechanical sensors (MEMSs).

Microcontroller-Based Sinusoidal Voltage Generation for Electrical Bio-Impedance Spectroscopy Applications
J.A. Castro, A. Olmo, P. Pérez and A. Yúfera
Journal Paper · Journal of Computer and Communications, vol. 4, no. 17, pp 51-58, 2016
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A sinusoidal voltage wave generator is proposed based on the use of micro-processor digital signals with programmable duty-cycles, with application to real-time Electrical Cell-substrate Impedance Spectroscopy (ECIS) assays in cell cultures. The working principle relies on the time convolution of the programmed microcontroller (μC) digital signals. The expected frequency is easily tuned on the bio-impedance spectroscopy range [100 Hz, 1 MHz] thanks to the μC clock frequency selection. This system has been simulated and tested on the 8 bits μC ArduinoTM Uno with ATmega328 version. Results obtained prove that only three digital signals are required to fit the general specification in ECIS experiments, below 1% THD accuracy, and show the appropriateness of the system for the real-time monitoring of this type of biological experiments.

Monitoring living cell assays with bio-impedance sensors
P. Daza, A. Olmo, D. Cañete and A. Yúfera
Journal Paper · Sensors and Actuators, B: Chemical, vol. 176, pp 605-610, 2013
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This work proposes a cell¿microelectrode model to be used on cell culture assays as an alternative to end-point protocols employed in cell growth and cell biometry applications. The microelectrode model proposed is based on the area overlap between the microelectrode sensor and the living cells as main parameter. This model can be applied to cell size identification, cell count, and their extension to cell growth, motility and dosimetry protocols. A procedure to fit the proposed model to microelectrode electrical performance is presented, enabling the decoding of empirical measurements and its interpretation in terms of number of cells. This fitting procedure depends on three parameters: microelectrode geometry, gap resistance between substrate attached cells and microelectrode and, mainly, on microelectrode area covered by cells. The model has been implemented employing Analog Hardware Descriptions Language (AHDL) to be incorporated also to mixed-mode simulation processes during circuit design flow.

Cell-culture real time monitoring based on bio-impedance measurements
P. Daza, D. Cañete, A. Olmo, J.A. García and A. Yúfera
Journal Paper · Sensors & Transducers Journal, vol. 14-1, Special Issue, pp 266-275, 2012
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This paper proposes the application of a cell-microelectrode model in cell biometry experiments, using the cell-electrode area overlap as its main parameter. The model can be applied to cell size identification and cell count, and further extended to study cell growth and dosimetry protocols. Experiments have been conducted in AA8 cell line, obtaining promising results.

Use of Electrical Impedance Spectroscopy (EIS) to Monitor Cryoprotectant Concentration in Cellular and Tissue Cryopreservation Protocols
A. Olmo, B. Buzon, A. Yufera and R. Risco
Journal Paper · Cryobiology, vol. 61, no. 3, pp 392, 2010
resumen      doi      

In this paper we theoretically analyse the use of Electrical Impedance Spectroscopy (EIS) to efficiently monitor cryoprotectant concentration in cryopreservation protocols.
In order to correctly determine the perfusion of cryoprotectant inside tissues and organs, with its spatial distribution, it is necessary to previously study the influence that the frequency, temperature and electrode configuration have in bioimpedance measurements.
We have analysed with COMSOL Multiphysics software the frequency response of a 2-electrode system to different concentrations of Me2SO, perfused into 3T3 fibroblasts and monolayers of Mesenchymal Stem Cells (MSCs), fundamental for tissue-based therapeutics.
An electrical model based on a previous work was used (Olmo et al., 2010). This model constitutes a complete electrical description of the electrode-cell system, including the electrode-electrolyte double layer, electrode-cell gap, electrolyte-cell double layer and cellular membrane, besides taking into account electrical properties of intra and extra-cellular medium. The Quasi-statics module of COMSOL was used to perform the finite element simulations, for different frequencies.
Simulations show the system easily detects changes in cryoprotectant concentration, being necessary to optimize both frequency and electrode configuration to efficiently detect cryoprotectant perfusion inside the biological material. Temperature influence on conductivity and permittivity, as well as on electrode interfaces, has also been discussed in detail.
The model and finite element method simulation herewith reported has proved to be a good tool, which can be used to optimize the design of experimental setups. These models can also be extended in the future, to analyse the use of more complex three-dimensional EIS systems, which can monitor cryoprotectant perfusion in the cryopreservation of more complex tissues and organs.

Congresos


Characterization of Implanted Stents through Neointimal Tissue Bioimpedance Simulations
J.M. Portillo-Anaya, P. Pérez, G. Huertas, A. Olmo, J.A. Serrano, A. Maldonado-Jacobi and A. Yúfera
Conference · International Conference of the IEEE Engineering in Medicine and Biology Society EMBC2019
resumen     

This work describes how is possible the definition of the light hole or lumen in implanted stents affected by restenosis processes using the BioImpedance (BI) as biomarker. The main approach is based on the fact that neointimal tissues implied in restenosis can be detected and measured thanks to their respective conductivity and dielectric properties. For this goal, it is proposed a four-electrode setup for bioimpedance measurement. The influence of the several involved tissues in restenosis: fat, muscle, fiber, endothelium and blood, have been studied at several frequencies, validating the setup and illustrating the sensitivity of each one. Finally, a real example using a standard stent, has been analyzed for stable and vulnerable plaques in restenosis test cases, demonstrating that the proposed method is useful for the stent obstruction test. Bioimpedance simulation test has been performed using the electric physics module in COMSOL Multiphysics®.

Practical Characterization of Cell-Electrode Electrical Models in Bio-Impedance Assays
J.A. Serrano, P. Pérez, A. Maldonado, M. Martín, A. Olmo, P. Daza, G. Huertas and A. Yúfera
Conference · International Conference on Biomedical Electronics and Devices BIODEVICES 2018
resumen     

This paper presents the fitting process followed to adjust the parameters of the electrical model associated to a cell-electrode system in Electrical Cell-substrate Impedance Spectroscopy (ECIS) technique, to the experimental results from cell-culture assays. A new parameter matching procedure is proposed, under the basis of both, mismatching between electrodes and time-evolution observed in the system response, as consequence of electrode fabrication processes and electrochemical performance of electrode-solution interface, respectively. The obtained results agree with experimental performance, and enable the evaluation of the cell number in a culture, by using the electrical measurements observed at the oscillation parameters in the test circuits employed.

Monitoring Muscle Stem Cell Cultures with Impedance Spectroscopy
Y. Yuste, J.A. Serrano, A. Olmo, A. Maldonado-Jacobi, P. Pérez, G. Huertas, S. Pereira, F. de la Portilla and A. Yúfera
Conference · International Conference on Biomedical Electronics and Devices BIODEVICES 2018
resumen     

The aim of this work is to present a new circuit for the real-time monitoring the processes of cellular growth and differentiation of skeletal myoblast cell cultures. An impedance spectroscopy Oscillation-Based technique is proposed for the test circuit, converting the biological system into a voltage oscillator, and avoiding the use of very high performance circuitry or equipment. This technique proved to be successful in the monitoring of cell cultures growth levels and could be useful for determining the degree of differentiation achieved, of practical implications in tissue engineering.

Bioimpedance real-time characterization of neointimal tissue inside stents
D. Rivas-Marchena, A. Olmo, G. Huertas, A. Yúfera, J. A. Miguel and M. Martinez
Conference · Conference on Design of Circuits and Integrated Systems DCIS 2017
resumen     

It is hereby presented a new approach to monitor restenosis in arteries fitted with a stent during an angioplasty. The growth of neointimal tissue is followed up by measuring its bioimpedance with Electrical Impedance Spectroscopy (EIS). Besides, a mathematical model is derived to analytically describe the neointima´s histological composition from its bioimpedance. The model is validated by finite-element analysis (FEA) with COMSOL Multiphysics ®. Satisfactory correlation between the analytical model and the FEA simulation is achieved for most of the characterization range, detecting some deviations introduced by the thin "double layer" that separates the neointima and the blood. It is shown how to apply conformal transformations to obtain bioimpedance models for stack-layered tissues over coplanar electrodes. Particularly, this is applied to characterize the neointima in real-time. This technique is either suitable as a main mechanism of restenosis follow-up or it can be combined with proposed blood-pressure-measuring intelligent stents to auto-calibrate the sensibility loss caused by the adherence of the tissue on the micro-electro-mechanical sensors (MEMS).

A Tracking Algorithm For Cell Motility Assays in CMOS Systems
C. Martínez-Gómez, A. Olmo, G. Huertas, P. Pérez, A. Maldonado-Jacobi and A. Yúfera
Conference · International Conference of the IEEE Engineering in Medicine and Biology Society EMBC 2017
resumen     

This work proposes a method for the study and real-time monitorization of a single cell on a 2D electrode matrix, of great interest in cell motility assays and in the characterization of cancer cell metastasis. A CMOS system proposal for cell location based on occupation maps data generated from Electrical Cell-substrate Impedance Spectroscopy (ECIS) has been developed. From experimental assays data, an algorithm based on analysis of the eight nearest neighbours has been implemented to find the cell center of mass. The path followed by a cell, proposing a Brownian route, has been simulated with the proposed algorithm. The presented results give an accuracy over 95% in the determination of the coordinates (x, y) from the expected cell center of mass.

A CMOS Tracking System Approach for Cell Motility Assays
C. Martínez-Gómez, A. Olmo, G. Huertas, P. Pérez, A. Maldonado-Jacobi and A. Yufera
Conference · International Conference on Biomedical Electronics and Devices BIODEVICES 2017
resumen     

This work proposes a method for studying and monitoring in real-time a single cell on a 2D electrode matrix, of great interest in cell motility assays and in the characterization of cancer cell metastasis. A CMOS system proposal for cell location based on occupation maps data generated from Electrical Cell-substrate Impedance Spectroscopy (ECIS) has been developed. From this cell model, obtained from experimental assays data, an algorithm based on analysis of the 8 nearest neighbors has been implemented, allowing the evaluation of the cell center of mass. The path followed by a cell, proposing a Brownian route, has been simulated with the proposed algorithm. The presented results show the success of the approach, with accuracy over 95% in the determination of any coordinate (x, y) from the expected center of mass.

An Impedance-Based Microscopy for Cell-Culture Imaging Using Microelectrode Sensors
A. Olmo, G. Huertas and A. Yúfera
Conference · Conference on Design of Circuits and Integrated Systems DCIS 2012
resumen     

Abstract not available

A Microscopy Technique based on Bio-impedance Sensors
A. Yúfera, G. Huertas and A. Olmo
Conference · European Conference on Solid-State Transducers EUROSENSOR 2012
resumen      pdf

It is proposed a microscopy for cell culture applications based on impedance sensors. The imagined signals are measured with the Electrical Cell-Substrate Spectroscopy (ECIS) technique, by identifying the cell area. The proposed microscopy allows real-time monitoring inside the incubator, reducing the contamination risk by human manipulation. It requires specific circuits for impedance measurements, a two-dimensional sensor array (pixels), and employing electrical models to decode efficiently the measured signals. Analogue Hardware Description Language (AHDL) circuits for cell-microelectrode enables the use of geometrical and technological data into the system design flow. A study case with 8x8 sensor array is reported, illustrating the evolution and power of the proposed image acquisition.

Use of Electrical Impedance Spectroscopy (EIS) to Monitor Cryoprotectant Concentration in Cellular and Tissue Cryopreservation Protocols
A. Olmo, B. Buzon, A. Yufera and R. Risco
Conference · Annual Meeting on the Society for Cryobiology CRYO 2010
resumen     

In this paper we theoretically analyse the use of Electrical Impedance Spectroscopy (EIS) to efficiently monitor cryoprotectant concentration in cryopreservation protocols.
In order to correctly determine the perfusion of cryoprotectant inside tissues and organs, with its spatial distribution, it is necessary to previously study the influence that the frequency, temperature and electrode configuration have in bioimpedance measurements.
We have analysed with COMSOL Multiphysics software the frequency response of a 2-electrode system to different concentrations of Me2SO, perfused into 3T3 fibroblasts and monolayers of Mesenchymal Stem Cells (MSCs), fundamental for tissue-based therapeutics.
An electrical model based on a previous work was used (Olmo et al., 2010). This model constitutes a complete electrical description of the electrode-cell system, including the electrode-electrolyte double layer, electrode-cell gap, electrolyte-cell double layer and cellular membrane, besides taking into account electrical properties of intra and extra-cellular medium. The Quasi-statics module of COMSOL was used to perform the finite element simulations, for different frequencies.
Simulations show the system easily detects changes in cryoprotectant concentration, being necessary to optimize both frequency and electrode configuration to efficiently detect cryoprotectant perfusion inside the biological material. Temperature influence on conductivity and permittivity, as well as on electrode interfaces, has also been discussed in detail.
The model and finite element method simulation herewith reported has proved to be a good tool, which can be used to optimize the design of experimental setups. These models can also be extended in the future, to analyse the use of more complex three-dimensional EIS systems, which can monitor cryoprotectant perfusion in the cryopreservation of more complex tissues and organs.

Bioimpedance monitoring for cryopreservation process control
A. Olmo, B. Buzón, A. Yúfera and R. Risco
Conference · International Conference of IEEE Engineering in Medicine and Biology Society EMBS 2010
resumen      pdf

This paper analyses the use of Electrical Impedance Spectroscopy (EIS) to efficiently monitor cryoprotectant concentrations in cryopreservation protocols. The proposed technique can improve methods such as Liquidus Tracking (LT), allowing vitrification without exposing tissues to damaging concentrations of cryoprotectant at relatively high temperatures, and avoiding rapid temperature changes. This work is focused to continuous monitoring of cryoprotectant concentrations by detecting changes in electrical impedance. These variations, derived from cryoprotectant perfusion inside cells and tissues, can be efficiently measure by using of EIS. Finite element simulation performed with COMSOL Multiphysics software was used to analyse the frequency response of a two-electrode system to several concentrations of Me2SO, perfused into 3T3 fibroblasts and monolayers of Mesenchymal Stem Cells (MSCs), fundamental in tissue-based therapeutics.

Finite element simulation of microelectrodes for bio-impedance sensor applications
A. Olmo and A. Yúfera
Conference · International Conference on Sensor Device Technologies and Applications SENSORDEVICES 2010
resumen      pdf

Electrical models for microelectrode-cell interfaces are essential to match electrical simulations to real bio-systems performance and correctly to decode the results obtained experimentally. The accurate performance simulation of a microelectrode sensor to changes in the cell-electrode system, such as cell growth, enables the optimum microelectrode design process. We report the use of COMSOL quasi-static mode, contrary to other DC modes frequently used, including magnetic fields to calculate the bioimpedance of the system. A fully electrode-cell model has been built, and the effect of fibroblasts of different diameters on the simulated impedance of small microelectrodes (32-um square) has been studied, in order to validate the model and to characterize the microelectrode sensor response to changes in cell size and density. © 2010 IEEE.

Computer simulation of microelectrode based bio-impedance measurements with COMSOL
A. Olmo and A. Yúfera
Conference · International Conference on Biomedical Electronics and Devices BIODEVICES 2010
resumen     

Electrical models for microelectrode-cell interfaces are essential to match electrical simulations to real bio-systems performance and correctly to decode the results obtained experimentally. The accurate response simulation of a microelectrode sensor to changes in the cell-electrode system, such as cell growth, enables the optimum microelectrode design process. We report the use of COMSOL quasi-static mode, contrary to other DC modes frequently used, including magnetic fields to calculate the bioimpedance of the system. A fully electrode-cell model has been built, and the effect of fibroblasts of different diameters on the simulated impedance of small microelectrodes (32-um square) has been studied, in order to validate the model and to characterize the microelectrode sensor response to changes in cell size and density.

Libros


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Capítulos de libros


Remote Sensing of Cell-Culture Assays
P. Pérez, A. Maldonado-Jacobi, A.J. López, C. Martínez, A. Olmo, G. Huertas and A. Yufera
Book Chapter · New Insights into Cell Culture Technology, pp 135-155, 2017
resumen      doi      

This chapter describes a full system developed to perform the remote sensing of cell-culture experiments from any access point with internet connection. The proposed system allows the real-time monitoring of cell assays thanks to bioimpedance measurement circuits developed to count the number of cell present in a culture. Cell-culture characterization is performed through the measurement of the increasing bioimpedance parameter over time. The circuit implementation is based on the oscillation-based test (OBT) methodology. Bioimpedance of cell cultures is measured in terms of the oscillation parameters (frequency, amplitude, phase, etc.) and used as empirical markers to carry out an appropriate interpretation in terms of cell size identification, cell counting, cell growth, growth rhythm, etc. The device is capable of managing the whole sensing task and performs wireless communication through a Bluetooth module. Data are interpreted and displayed on a computer or a mobile phone through a web application. The system has its practical application in drug development processes, offering a label-free, high-throughput, and high-content screening method for cellular research, avoiding the classical end-point techniques and a significant workload and cost material reduction.

Cell biometrics based on bio-impedance measurements
A. Yúfera, A. Olmo, P. Daza, P and D. Cañete
Book Chapter · Advanced Biometric Technologies, pp 343-366, 2011
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Many biological parameters and processes can be sensed and monitored using their impedance as marker (Grimmes, 2008), (Beach. 2005), (Yúfera, 2005), (Radke, 2004), with the advantage that it is a non-invasive, relatively cheap technique. Cell growth, cell activity, changes in cell composition, shapes or cell location are only some examples of processes which can be detected by microelectrode-cell impedance sensors (Huang, 2004) (Borkholder, 1998). The electrical impedance of a biological sample reflects actual physical properties of the tissue. In frequency dependent analyses, the β-dispersion ranging from kilohertzs to hundreds of megahertzs (Schwan, 1957) is mainly affected by the shape of the cells, the structure of the cell membranes, and the amount of intra and extra cellular solution. Electrical bio-impedance can be used to assess the properties of biological materials (Ackmann, 1993) involved in processes such as cancer development (Giaever, 1991), (Blady, 1996), (Aberg, 2004); because the cells of healthy tissues and cancer are different in shape, size and orientation, abnormal cells can be detected using their impedance as a marker.
Among Impedance Spectroscopy (IS) techniques, Electrical Cell-substrate Impedance Spectroscopy (ECIS) (Giaever, 1986), based on two-electrode setups, allows the measurement of cell-culture impedances and makes it possible to determine the biological condition (material, internal activity, motility and size) of a cell type and its relationship with the environment; for example, the transfer flow through the cell membrane (Wang, 2010). One of the main drawbacks of the ECIS technique is the need to use efficient models to decode the electrical results obtained. To efficiently manage bio-impedance data, reliable electrical models of the full system comprising electrodes, medium and cells are required. Several studies have been carried out in this field (Giaever, 1991), (Huang, 2004), (Borkholder, 1998), (Joye, 2008), (Olmo, 2010), some of them employing Finite Element simulation (FEM) for impedance model extraction. These models are the key for matching electrical simulations to real system performances and hence for correctly decoding the results obtained in experiments.

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