Alberto Olmo

Cell Biometrics Based on Bio-Impedance Measurements

Many biological parameters and processes can be sensed and monitored using their impedance as marker (Grimmes, 2008), (Beach. 2005), (Yufera, 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 -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.

Assessment of the cryoprotectant concentration inside a bulky organ for cryopreservation using X-ray computed tomography

Cryoprotection of bulky organs is crucial for their storage and for subsequent transplantation. In this work we demonstrate the capability of the X-ray computed tomography (CT) as a non-invasive method to measure the cryoprotectant (cpa) concentration inside a tissue or an organ, specifically for the case of dymethil sulfoxide (Me2SO). It is remarkable that the use of Me2SO has been leader in techniques of cells and tissues cryopreservation. Although CT technologies are mainly based in density differences, and many cpas are alcohols with densities similar to water, the use of very low energies as acceleration voltage (∼70 kV) and the sulfur atom in the molecule of Me2SO makes possible the visualization of this cpa inside tissues. As result we obtain a CT signal proportional to the Me2SO concentration with a spatial resolution up to 50 μm in the case of our device.

Sensing Cell-Culture Assays with Low-Cost Circuitry

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

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).

Integration of multimedia contents in the teaching of electronics: A practical test case in the teaching of digital circuits at the University of Seville

In this paper we present the integration of multimedia contents in the teaching of Digital Electronic Circuits and Computer Structure, of the first course of Computer Engineering at the University of Seville. Different tools for screenshot and video recording have been used for the preparation of audiovisual material, integrated in the learning platform currently used at the University of Seville. Feedback on the prepared material was collected in a survey, showing the interest and utility found by students in the preparation of theoretical and experimental classes with the videos. Successful results have been obtained in the evaluation of students. Suggestions of improvement and further work to be carried out are also described in the paper.

Bioimpedance monitoring for cryopreservation process control

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

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-µm square) has been studied, in order to validate the model and to characterize the microelectrode sensor response to changes in cell size and density.

Practical Characterization of Cell-Electrode Electrical Models in Bio-Impedance Assays

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

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.

An Empirical-Mathematical Approach for Calibration and Fitting Cell-Electrode Electrical Models in Bioimpedance Tests

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.