Orthogonal Multitone Electrical Impedance Spectroscopy-OMEIS-for the Study of Fibrosis Induced by Active Cardiac Implants

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L’archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d’enseignement et de recherche français ou étrangers, des laboratoires publics ou privés. Orthogonal Multitone Electrical Impedance Spectroscopy OMEIS for the Study of Fibrosis Induced by Active Cardiac Implants Edwin de Roux, Amélie Degache, Mehdi Terosiet, Florian Kolbl, Michel Boissière, Emmanuel Pauthe, Aymeric Histace, Olivier Bernus, Noëlle Lewis, Olivier Romain


Introduction
These systems use the latest micro-nano-electronics technologies, with electrodes that stimulate and sense the surrounding biological environment.Such implanted devices induce an immediate and sustained inflammatory response of the body.This chronic and unresolved inflammation induces fibrosis, which is a complex biological process involving multiscale phenomena.At the cellular scale, fibroblasts are activated and differentiate to myofibroblasts; at the tissular scale, excessive secretion of extracellular matrix components, like collagen, finally produces a dense fibrous capsule around the implants, especially the electrodes [1].
In case of PCM, fibrosis reduces both the functionality and the efficacy of the implant to target the desired tissue, diverting even the stimulating current to unforeseen regions and altering the impedance of the tissue around electrodes.Furthermore, it could change the shape and magnitude of the electric field generated [2] and forces the increase of the PCM stimulation threshold and the reduction of battery lifespan [3].
The fibrosis can be treated medically to reduce its consequences, but the effectiveness of the treatment depends on the accurate diagnosis.The standard method for determining the degree of tissue reaction surrounding implanted electrodes is histology.Immunohistochemical methods enable the visualization of specific markers like collagen, fibronectin or smooth muscle actin [4].The disadvantage of these methods is an inability to follow the tissue reaction in real time in vivo [5].Optical methods such as Late Gadolinium enhanced Cardiac Magnetic Resonance (CMR) are used to detect fibrosis in cardiac tissues even if it is scattered and in low concentration [6].However, their effectiveness decreases when the patient carries a PCM because it could alter the quality of the image.In addition, due to risk factors, it is contraindicated to apply CMR in patients with PCM, and even if this is done, it is recommended to carry it out weeks after post-implant surgery, which allows a considerable accumulation of fibrous tissue on the electrodes [7].Chronic monitoring of tissue alteration around implanted electrodes could be a first step to understand this long-term biological process.This

Orthogonal Multitone Electrical Impedance Spectroscopy -OMEISfor the Study of Fibrosis Induced by Active Cardiac Implants
Over the last fifty years, electrotherapy has shown a very rapid development with many innovators contributing to a whole series of devices.Electrotherapy uses an external source of electricity to stimulate human tissue in ways that produce a beneficial therapeutic effect.The best known electrotherapy devices are the active implantable medical devices, among them cardiac defibrillators (1949), Peacemakers (PCM) (1957), cochlear (1971) and deep brain (2000) stimulators.
advance could be used to ascertain the treatment effectiveness or to test new biocompatibility strategies of materials.
Electrical Impedance S known technique for characterizing living tis Preliminary studies conclude between the measured resistance and the morphology of the tissue next to the electrode [ show that in or epicardium allowed discriminating healthy and infarcted tissue.These results suggest that the tissue remodeling occurring in fibrosis has an EIS signature.propose to apply local EIS, taking benefit of th and the electronic circuitry already exis continuous and 'low fibrosis.This system should be capable of performing the measurements under the severe condition imposed by the dynamics of the heart, such as the heartbeat movement.Here the impedance must be sensed over the desired frequency range during the short slot time between two consecutive muscle contractions, in order to avoid distorting the measurements, as shown in Figure 1.The method must meet the following requirements: high measurement speed, flexibility in spectrum manipulation (bandwidth and frequency resolution) and feasibility of the digital implementation.
Based on this strategy we have devised a measurement approach, fast and flexible, easy to synchronize with the PCM pulses and not affected by the heart dynamics.The innovation is an original application of the Orthogonal Frequency Division Multiplexing (OFDM) technique, well digital communication.This technique, here adapted to embedded EIS, exhibits competitive performances, compared to traditional EIS methods, and meet all the previous requirements.This new appro the Orthogonal Multitone Electrical Impedance Spectroscopy This article is structured as following: firstly, the EIS measurement principle and its traditional implementations are presented.Then, the described in the secon of an OMEIS prototype, its validation and properties.Subsequently, preliminary experiments are conducted in both in vitro cardiac tissues, respectively.Finally the results the impedance measurements are discussed.advance could be used to ascertain the treatment effectiveness or to test new biocompatibility strategies of Electrical Impedance S known technique for characterizing living tis Preliminary studies conclude between the measured resistance and the morphology of the tissue next to the electrode [ in situ impedance measurements in myocardium or epicardium allowed discriminating healthy and infarcted tissue.These results suggest that the tissue remodeling ing in fibrosis has an EIS signature.propose to apply local EIS, taking benefit of th and the electronic circuitry already exis and 'low-cost' monitoring of electrode fibrosis.This system should be capable of performing the measurements under the severe condition imposed by the f the heart, such as the heartbeat movement.Here the impedance must be sensed over the desired frequency range during the short slot time between two consecutive muscle contractions, in order to avoid distorting the ements, as shown in Figure 1.The method must meet the following requirements: high measurement speed, flexibility in spectrum manipulation (bandwidth and frequency resolution) and feasibility of the digital implementation.
Based on this strategy we have devised a measurement approach, fast and flexible, easy to synchronize with the PCM pulses and not affected by the heart dynamics.The innovation is an original application of the Orthogonal Frequency Division Multiplexing (OFDM) technique, well-known and s digital communication.This technique, here adapted to embedded EIS, exhibits competitive performances, compared to traditional EIS methods, and meet all the previous requirements.This new appro Orthogonal Multitone Electrical Impedance Spectroscopy -OMEIS-.
This article is structured as following: firstly, the EIS measurement principle and its traditional implementations are presented.Then, the in the second section of an OMEIS prototype, its validation and properties.Subsequently, preliminary experiments are conducted in in vitro and ex vivo cardiac tissues, respectively.Finally the results the impedance measurements are discussed.
Strategy for measuring cardiac tissue PCM structure.The ECG signal advance could be used to ascertain the treatment effectiveness or to test new biocompatibility strategies of Electrical Impedance Spectroscopy (EIS) is a well known technique for characterizing living tis Preliminary studies conclude that there is a correlation between the measured resistance and the morphology of the tissue next to the electrode [8].Amoros impedance measurements in myocardium or epicardium allowed discriminating healthy and infarcted tissue.These results suggest that the tissue remodeling ing in fibrosis has an EIS signature.propose to apply local EIS, taking benefit of th and the electronic circuitry already existing in PCM, for the cost' monitoring of electrode fibrosis.This system should be capable of performing the measurements under the severe condition imposed by the f the heart, such as the heartbeat movement.Here the impedance must be sensed over the desired frequency range during the short slot time between two consecutive muscle contractions, in order to avoid distorting the ements, as shown in Figure 1.The method must meet the following requirements: high measurement speed, flexibility in spectrum manipulation (bandwidth and frequency resolution) and feasibility of the Based on this strategy we have devised a measurement approach, fast and flexible, easy to synchronize with the PCM pulses and not affected by the heart dynamics.The innovation is an original application of the Orthogonal Frequency Division Multiplexing (OFDM) known and successfully used in the field of digital communication.This technique, here adapted to embedded EIS, exhibits competitive performances, compared to traditional EIS methods, and meet all the previous requirements.This new approach is called hereafter Orthogonal Multitone Electrical Impedance This article is structured as following: firstly, the EIS measurement principle and its traditional implementations are presented.Then, the original OMEIS method d section.Next, we develop the design of an OMEIS prototype, its validation and properties.Subsequently, preliminary experiments are conducted in ex vivo samples, with living cells and cardiac tissues, respectively.Finally the results the impedance measurements are discussed.gy for measuring cardiac tissue impedance while using the PCM structure.The ECG signal is in black together with the PCM pulse.
advance could be used to ascertain the treatment effectiveness or to test new biocompatibility strategies of pectroscopy (EIS) is a well known technique for characterizing living tis that there is a correlation between the measured resistance and the morphology of the ].Amoros-Figueras, et al. impedance measurements in myocardium or epicardium allowed discriminating healthy and infarcted tissue.These results suggest that the tissue remodeling ing in fibrosis has an EIS signature.Hence propose to apply local EIS, taking benefit of the electrodes ting in PCM, for the cost' monitoring of electrode-induced fibrosis.This system should be capable of performing the measurements under the severe condition imposed by the f the heart, such as the heartbeat movement.Here the impedance must be sensed over the desired frequency range during the short slot time between two consecutive muscle contractions, in order to avoid distorting the ements, as shown in Figure 1.Therefore, the selected method must meet the following requirements: high measurement speed, flexibility in spectrum manipulation (bandwidth and frequency resolution) and feasibility of the Based on this strategy we have devised a new EIS measurement approach, fast and flexible, easy to synchronize with the PCM pulses and not affected by the heart dynamics.The innovation is an original application of the Orthogonal Frequency Division Multiplexing (OFDM) uccessfully used in the field of digital communication.This technique, here adapted to embedded EIS, exhibits competitive performances, compared to traditional EIS methods, and meet all the ach is called hereafter Orthogonal Multitone Electrical Impedance This article is structured as following: firstly, the EIS measurement principle and its traditional implementations original OMEIS method .Next, we develop the design of an OMEIS prototype, its validation and properties.Subsequently, preliminary experiments are conducted in samples, with living cells and cardiac tissues, respectively.Finally the results obtained in the impedance measurements are discussed.
impedance while using the in black together with the PCM pulse.
advance could be used to ascertain the treatment effectiveness or to test new biocompatibility strategies of pectroscopy (EIS) is a wellknown technique for characterizing living tissues.
that there is a correlation between the measured resistance and the morphology of the et al. [9], impedance measurements in myocardium or epicardium allowed discriminating healthy and infarcted tissue.These results suggest that the tissue remodeling Hence, we e electrodes ting in PCM, for the induced fibrosis.This system should be capable of performing the measurements under the severe condition imposed by the f the heart, such as the heartbeat movement.Here the impedance must be sensed over the desired frequency range during the short slot time between two consecutive muscle contractions, in order to avoid distorting the , the selected method must meet the following requirements: high measurement speed, flexibility in spectrum manipulation (bandwidth and frequency resolution) and feasibility of the new EIS measurement approach, fast and flexible, easy to synchronize with the PCM pulses and not affected by the heart dynamics.The innovation is an original application of the Orthogonal Frequency Division Multiplexing (OFDM) uccessfully used in the field of digital communication.This technique, here adapted to embedded EIS, exhibits competitive performances, compared to traditional EIS methods, and meet all the ach is called hereafter Orthogonal Multitone Electrical Impedance This article is structured as following: firstly, the EIS measurement principle and its traditional implementations original OMEIS method is .Next, we develop the design of an OMEIS prototype, its validation and properties.Subsequently, preliminary experiments are conducted in samples, with living cells and obtained in

2.
The EIS applied to the analysis consists of tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation electrodes.methods for EIS used for this purpose, with the and disadvantages.

2.1.
found in almost all commercial available EIS instruments, due to its sinusoidal signal.Here the measurements are carried out at a specific frequency, as is the case for these full designs [ found in the commercia which measures the impedance at three discrete frequencies.
spectroscopy could be performed within a larger set of determined frequencies.The impedance analysis frequency range provides more insights regarding the tissue features.

EIS Overview
The EIS applied to the analysis consists of injecting an alternating current or voltage into the tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation electrodes.There are several alternating methods for EIS used for this purpose, with the and disadvantages.

Classic M
found in almost all commercial available EIS instruments, due to its implementation simplicity, is the fixed frequency sinusoidal signal.Here the measurements are carried out at a specific frequency, as is the case for these full designs [10,11], or in a small set of frequencies such as it is found in the commercia which measures the impedance at three discrete frequencies.
When the frequency sweep technique is used, the spectroscopy could be performed within a larger set of determined frequencies.The impedance analysis frequency range provides more insights regarding the tissue features.Gabriel et al. measured the impedance of various organs of the human body in different conditions.The obtained frequency responses are specific signatures for each tissue [13].This technique presents some limitations in the estimation of time impedance estimation of multiple electrodes or samples needed in a short time frame by the same device.these restrictions, this method does not requirements of the proposed strategy.

Broadband broadband signals have been limitation of classic approaches
back to 1975 where pseudo used for the measurement of an electrode impedance in a wide frequency band [ correlating the Sample U pseudo-random stimulus signal, which results in the impulse response, following by a Fourier transform.Maximum Length Sequence (MLS) than correlation has been proposed in [ Hadamard transform range of frequencies and for t impedance spectrum measurement.random signals or MLS are preferred instead of the Dirac pulse, whose high amplitude peak is not desirable for the stimulation of biological samples [ drawback of this method is the signal amplitude variation at each frequency.The spectrum of a MLS signal is also random and it is possible that the energy at a desired frequency could be too low or equal in amplitude to the noise, which would induce errors in the 2.3.Multisine spectrum measurement of biological samples, reported for instance in [18,19] and [20

Overview
The EIS applied to the analysis injecting an alternating current or voltage into the tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation There are several alternating methods for EIS used for this purpose, with the and disadvantages.These will be discussed Methods for EIS found in almost all commercial available EIS instruments, implementation simplicity, is the fixed frequency sinusoidal signal.Here the measurements are carried out at a specific frequency, as is the case for these full ], or in a small set of frequencies such as it is found in the commercial instrument xCELLigence [ which measures the impedance at three discrete frequencies.
When the frequency sweep technique is used, the spectroscopy could be performed within a larger set of determined frequencies.The impedance analysis frequency range provides more insights regarding the tissue features.Gabriel et al. measured the impedance of various organs of the human body in different conditions.The obtained frequency responses are specific signatures for ].This technique presents some limitations in the estimation of time-variable systems or when the impedance estimation of multiple electrodes or samples needed in a short time frame by the same device.these restrictions, this method does not requirements of the proposed strategy.
Broadband EIS.Different methods for the generation of broadband signals have been classic approaches back to 1975 where pseudo used for the measurement of an electrode impedance in a wide frequency band [14].The impedance is estim correlating the Sample Under random stimulus signal, which results in the impulse e, following by a Fourier transform.Maximum Length Sequence (MLS) than correlation has been proposed in [ Hadamard transform.This kind of signals contains a large range of frequencies and for t impedance spectrum measurement.random signals or MLS are preferred instead of the Dirac pulse, whose high amplitude peak is not desirable for the ulation of biological samples [ of this method is the signal amplitude variation at each frequency.The spectrum of a MLS signal is also random and it is possible that the energy at a desired frequency could be too low or equal in amplitude to the noise, which would induce errors in the Approach.Multisine signals for impedance spectrum measurement of biological samples, reported for 18, 19] and [20], provides also a fast estimation The EIS applied to the analysis of a biological material injecting an alternating current or voltage into the tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation There are several alternating methods for EIS used for this purpose, with the will be discussed ethods for EIS.The most common signal found in almost all commercial available EIS instruments, implementation simplicity, is the fixed frequency sinusoidal signal.Here the measurements are carried out at a specific frequency, as is the case for these full ], or in a small set of frequencies such as it is l instrument xCELLigence [ which measures the impedance at three discrete frequencies.
When the frequency sweep technique is used, the spectroscopy could be performed within a larger set of determined frequencies.The impedance analysis frequency range provides more insights regarding the tissue features.Gabriel et al. measured the impedance of various organs of the human body in different conditions.The obtained frequency responses are specific signatures for ].This technique presents some limitations in variable systems or when the impedance estimation of multiple electrodes or samples needed in a short time frame by the same device.these restrictions, this method does not requirements of the proposed strategy.

Different methods for the generation of broadband signals have been investigated
classic approaches.The first approach is back to 1975 where pseudo-random binary signals were used for the measurement of an electrode impedance in a ].The impedance is estim nder Test (SUT) response with the random stimulus signal, which results in the impulse e, following by a Fourier transform.Maximum Length Sequence (MLS) a more efficient method than correlation has been proposed in [15] based on t This kind of signals contains a large range of frequencies and for this reason allows a rapidly impedance spectrum measurement.In addition, pseudo random signals or MLS are preferred instead of the Dirac pulse, whose high amplitude peak is not desirable for the ulation of biological samples [16, of this method is the signal amplitude variation at each frequency.The spectrum of a MLS signal is also random and it is possible that the energy at a desired frequency could be too low or equal in amplitude to the noise, which would induce errors in the measurement.

Multisine signals for impedance spectrum measurement of biological samples, reported for
], provides also a fast estimation biological material injecting an alternating current or voltage into the tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation signal generation methods for EIS used for this purpose, with their advantages will be discussed next.
The most common signal found in almost all commercial available EIS instruments, implementation simplicity, is the fixed frequency sinusoidal signal.Here the measurements are carried out at a specific frequency, as is the case for these full-custom ], or in a small set of frequencies such as it is l instrument xCELLigence [ which measures the impedance at three discrete frequencies.
When the frequency sweep technique is used, the spectroscopy could be performed within a larger set of determined frequencies.The impedance analysis on a large frequency range provides more insights regarding the tissue features.Gabriel et al. measured the impedance of various organs of the human body in different conditions.The obtained frequency responses are specific signatures for ].This technique presents some limitations in variable systems or when the impedance estimation of multiple electrodes or samples needed in a short time frame by the same device.Due to these restrictions, this method does not meet the Different methods for the generation of investigated to overcome the The first approach is dated inary signals were used for the measurement of an electrode impedance in a ].The impedance is estimated by est (SUT) response with the random stimulus signal, which results in the impulse e, following by a Fourier transform.In the case of a more efficient method ] based on the Fast This kind of signals contains a large his reason allows a rapidly In addition, pseudo random signals or MLS are preferred instead of the Dirac pulse, whose high amplitude peak is not desirable for the , 17].The of this method is the signal amplitude variation at each frequency.The spectrum of a MLS signal is also random and it is possible that the energy at a desired frequency could be too low or equal in amplitude to the measurement.
Multisine signals for impedance spectrum measurement of biological samples, reported for ], provides also a fast estimation biological material injecting an alternating current or voltage into the tissue under study, and then measuring the resultant (voltage or current, respectively) that appears through the stimulation signal generation ir advantages The most common signal found in almost all commercial available EIS instruments, implementation simplicity, is the fixed frequency sinusoidal signal.Here the measurements are carried out at a custom ], or in a small set of frequencies such as it is l instrument xCELLigence [12] which measures the impedance at three discrete frequencies.
When the frequency sweep technique is used, the spectroscopy could be performed within a larger set of on a large frequency range provides more insights regarding the tissue features.Gabriel  ].The main of this method is the signal amplitude variation at each frequency.The spectrum of a MLS signal is also random and it is possible that the energy at a desired frequency could be too low or equal in amplitude to the Multisine signals for impedance spectrum measurement of biological samples, reported for ], provides also a fast estimation with the advantage that the Signal to Noise Ratio (SNR) can be improved when using random phases and also that the frequencies can be selected as required, i.e. linear or logarithmic [21].Such approach is simple however does not scale easily with higher numbers of frequencies.The memory required for the generation of the multisine increases with the number of tones, as it is shown in Table 1 and also found in [22].Furthermore, the detection and impedance estimation at the receiver side could impede the implementation of this method.It could be verified, in the mentioned references, that the stimulation part could be implemented in digital form by storing in memory the externally created multisine signal.However, the receiving part requires a more complex hardware for the demodulation of the signal, usually implemented by the use of a data acquisition board or system, such as an oscilloscope, and a Personal Computer (PC) for the impedance computation.
2.4.OFDM for our Application.Finally, orthogonal multitone signals also offer a wide spectrum for a rapid impedance estimation in the frequency bandwidth of interest.The generation of this kind of signals could be efficiently implemented by using the OFDM method [23].This OFDM technique is successfully used in the field of digital communication, however the application for impedance measurement is a novelty and requires of modifications, such as those presented in Section 4. The OFDM method allows to control the spectrum of the multitone signal with great flexibility by defining the values of the OFDM symbols, the use of an appropriate modulation scheme and / or the manipulation of the system parameters, such as the sampling frequency.
Compared to the multisine technique mentioned above, the OFDM method requires less memory for the implementation of the signal generator, as depicted by Table 1.Here the memory required for the multisine and the OFDM approaches for the generation of a broadband signal of length N.M is calculated, where N/2 is the quantity of frequencies of the broadband signal and M is the number of multisine periods or OFDM symbols needed to reduce noise by averaging.
As it can be noticed, the pre-calculated (stored in memory) multisine requires N•M memory words because it is advisable to apply different random phases on each period of the multisine signal for the improvement of the SNR.However, the normal OFDM approach requires only 2N memory words because during each cycle only one symbol of N samples is generated and sequentially exit to the Digital to Analog Converter (DAC).This continues, synchronously, until reaching M symbols.For these reasons, the OFDM approach is finally the solution that we have retained.

OFDM Considerations
The OFDM model used here is composed of M blocks of N symbols, ( , ), X k m 0,1,..., , with each symbol modulating one of a set of N subcarriers for the generation of the multitone signal.The subcarriers are orthogonal and a cost effective approach is achieved when the Inverse Fast Fourier Transformation (IFFT) algorithm is used, giving the following general equation: In our application, a low Crest Factor (CF) of the multitone signal is desirable to avoid intermodulation due non-linearities of the analog parts (such as the saturation of the operational amplifiers).We use a simple technique to reduce the CF that consist of generating multiple set of N random complex values by controlling the seed and the output delay parameters of a random bit generator, then applying each of them to the OFDM emitter and calculating the CF.The seed and delay yielding the best crest factor is retained.The crest factor is calculated as shown in equation ( 2): The output of the IFFT is a sequence of complex values as a function of time.For the sake of simplicity, real-valued signals are assumed hereafter which converts equation (1) into the following equation: where 0,..., -1 n N = is the discrete time index and 1,..., m M = is the symbol index, being N ∈Z Z Z Z and M ∈ Z Z Z Z the IFFT size and the quantity of symbols blocks, respectively and θ X is the random phase.
One step found in a telecommunication OFDM model is the addition of a Cyclic Prefix (CP) to reduce intersymbol interference caused by a multipath fading channel.This CP consist of taking a copy of N cp elements from the end of the symbol block and concatenating them in front of it.However, EIS measurements do not suffer from Multipath propagation, therefore, instead of a CP, a small Guard Interval (GI) with zeros at the extremes of the frequency band (at 0 and Fs/2 Hz) will be used to reduce the energy applied on these not useful frequencies.

OMEIS System Design, Implementation and Validation
The proposed OMEIS technique is based on the OFDM model explained above, however, some additional modifications are necessary for the implementation of the EIS measuring system as following: the shape and quantity of symbols should be generated taking into consideration the properties that a stimulation signal should have for fast EIS: low voltage, short duration and with a desired spectrum.It should assure that the energy is propagated at the frequency tones under test.Furthermore, a perfect synchronization is required in the implementation, therefore a pilot signal is used.Both additions will be explained in Section 4.1.
In the following, the system will be analyzed into two parts, first the emitting and finally the receiving part.

The OMEIS Emitter.
The emitter function is to generate the stimulation signal with the desired spectrum.The structure of the emitter is shown in Figure 2.a.It consists mainly of symbol generation, synchronization, inverse Fourier transformation and digital to analog conversion stages.
In the symbol generator, the code that gives shape to the multitone signal spectrum is created in a synchronized manner.There are several codes that offer a spectrum with specific differences.For example, a code with constant amplitude generates a spectrum with tones of similar values at the IFFT subcarrier frequencies, that is in Fp(n)=n•Fs/N, where Fs is the sampling frequency, N is the size of IFFT and n = 0, 1, ...,(N/2)-1.The SUT output to a signal with this spectrum could have an impulse responses that may require rapid sampling to capture the transients.Another example is a sinusoidal code, with constant frequency Fd, that when applied it is sparsed throughout the whole OFDM bandwidth, locating Fd above and below each subcarrier frequency, resulting in tones with amplitudes at frequencies Fp(n)-Fd and Fp(n)+Fd, being this signal a type of multisine.However, although this code does not inject much energy into the IFFT subcarrier frequencies, it could be used in case of nonlinearity analysis.Finally, a random code generates a random spectrum within the frequency bandwidth between 0 and Fs/2-Fs/N.However, as mentioned above, there is the possibility of a low amplitude in the frequencies of interest resulting in them being corrupted by noise.
Therefore, one solution is to combine a random code with a constant offset, other than zero, large enough to ensure good amplitude at the carrier frequencies.The latter is provided by a random-bit generator whose output values are mapped with a QPSK modulator and then amplified and shifted with a DC offset.The amplification (G) is selected based on the maximum signal amplitude required and the complex QPSK mapping is used because it introduces a random phase information to the IFFT subcarriers, as shown in eq. ( 3), that better improves the CF compared to real values mapping such as BPSK [24].The mathematical backgrounds concerning the performances (BER vs. SNR) of digital modulations show that the OFDM-BPSK and OFDM-QPSK are very similar, but for the CF, QPSK is better [25].The symbols are generated at a rate of one sample every 1/Fs seconds, with a total of N samples per measurement cycle.
Following the symbol generator is the synchronization section.This is a critical step during the calibration stage

S2P
x x x x( ( ( because it measures the system delay.This information is used for the correct synchronization between the transmitter and the receiver.Perfect synchronization between both systems is essential to ensure that the selected N input samples of the FFT corresponds to the IFFT transmitted symbol.
The method for synchronization is the following: before calibration, a pilot signal is used which generates voltage peaks at known intervals.The generated peaks can be detected using an appropriate threshold.This method allows the recording of, in one hand, the delay of the transmitter output signal, and on the other, the joint delay of the analogdigital-analog conversions and the AFE stages.
Next is the modulation stage.Here, as explained above, a GI is used before the IFFT.The size of the IFFT (N) gives the quantity of frequency points for the impedance spectrum.The minimum frequency is at Fs/N and the maximum at Fs/2-Fs/N with a frequency separation of Fs/N.Serial to Parallel (S2P) and Parallel to Serial (P2S) blocks are required before and after the IFFT.
Finally, the real output of the IFFT is selected and converted into an analog signal by the DAC.

The OMEIS Receiver.
Once the signal coming from the AFE is encoded by the ADC, the Control Unit multiplexes it, depending on whether it is the pilot signal, which is sent during the synchronization, or the multitone one; sending it to the Threshold detector (TH) or the OFDM demodulation, respectively.At the end of the synchronization, when performing calibration or measurement, the response of the SUT is first demodulated using the FFT, then accumulated with values of previous measurement cycles and finally stored in the corresponding memories: Ycr, Yci in calibration and Yr, Yi in measurement, where r stands for real and i for imaginary.
As it is shown in Figure 2.b, the demodulation produces complex values which are averaged, stored and transmitted independently.The accumulation can increase the number of bits of the samples but reduces the amount of data to be transmitted.The impedance is then estimated with the following equation: where H CAL is the known calibration SUT which may vary in frequency, but nevertheless, it is assumed constant in each cycle of multitone generation; Y is the measurement data and Y c the calibration data received, k = 1,2, ..., N.
At the end of the measurement, when all the computation cycles were performed, the resulting calibration and measurement data are sent to a PC for the final treatment and display.

The OMEIS Implementation.
The modified OFDM structure, detailed in the preceding sections, is suitable for embedded systems.The implementation takes advantage of the computing power and the parallel features of FPGAs, performing both emitter and receiver in a Cyclone IV FPGA device programmed in VHDL language.
Figure 3, shows the OMEIS system implementation.The emitter part of the OMEIS system is implemented by using the IFFT and the PLL Clock generator Megacores IP.The remaining block, for instance the Cell Automata (CA), the Fs Manager, the Pilot, and the QPSK blocks, were coded in VHDL.
The values of the emitter's parameters, such as IFFT size or sampling frequency can be adjusted as required.The default value used for the IFFT size is 1024.The Fs Manager allows to change the sampling frequency Fs of the system to 1 MHz, 500 kHz and 250 kHz.
The Symbol Generator is the combination of the CA and the QPSK blocks.First the CA uses a Cellular Automata algorithm for the creation of two (2) random bits that are next mapped in the following way: for bits "11" the QPSK output is (G+jG)+DC, "10" gives (G-jG)+DC, a "01" is (-G+jG)+DC and finally "00" results in (-G-jG)+DC, where G is 1023 and DC equal to 80 in our implementation, both values were selected taking into consideration the output voltage amplitude and frequency spectrum shape.
A synchronous state machine, which operates as a Control Unit (CU), manages the overall behavior of the system.Both, the CU and the DAC/ADC manager are coded in VHDL as well.
An Analog custom board has been developed in order to perform EIS in biological samples.It has a parallel input 12  At the receiver using the FFT Megacores of Altera the same as that of SUT data, coming from the 256 Mb SDRAM SPI to a PC (HMI), coded in Matlab, for the control of the system as well as for the analysis of the data received running equation (4) for the final impedance estimati block, coded in VHDL, employs a threshold algorithm to detect the pilot signals and compute the transmitted signal output delay and the system response delay to send them to the CU.

FPGA OMEIS System
Figure 5 shows the OMEIS emitter as a of resources number of multipliers is 24 for all double the quantity 4.4.The Orthogonal Multitone Spectrum.multitone signal was analyzed by using the OMEIS implementation configured with a sampling frequency of Fs = 1 MHz, 32 symbols and an IFFT size of a multitone subcarrier frequency separation of 976.5 kHz.Also, a guard interval of 10 frequencies points at the bits DAC converter (DAC7821), a parallel outpu ADC (ADS850Y) and an A depicted in Figure 4.This AFE board has stimulation voltage amplitude of 200 mV, low dance and a very low WE) and E required for biological mea filters non desired DC and 50/60 multitone signal.The stimulation signal is voltage and the resulted current is capture Transimpedance Amplifier ( feedback resistor is set during calibration. At the receiver side, the demodulator is implemented the FFT Megacores of Altera that of the IFFT.Both the calibration and the oming from the 256 Mb SDRAM (W9825G6EH) PC.The PC runs a , coded in Matlab, for the control of the system as for the analysis of the data received running equation r the final impedance estimati , coded in VHDL, employs a threshold algorithm to detect the pilot signals and compute the transmitted signal output delay and the system response delay to send them to shows the cost in OMEIS emitter as a function of the IFFT size for N = 1024 are depicted on the plots.number of multipliers is 24 for all quantity of frequencies of the multit The Orthogonal Multitone Spectrum.multitone signal was analyzed by using the OMEIS implementation configured with a sampling frequency of = 1 MHz, 32 symbols and an IFFT size of giving a multitone subcarrier frequency separation of 976.5 a guard interval of 10 frequencies points at the The Orthogonal Multitone Spectrum.multitone signal was analyzed by using the OMEIS implementation configured with a sampling frequency of = 1 MHz, 32 symbols and an IFFT size of giving a multitone subcarrier frequency separation of 976.5 a guard interval of 10 frequencies points at the The Orthogonal Multitone Spectrum.The orthogonal multitone signal was analyzed by using the OMEIS implementation configured with a sampling frequency of = 1 MHz, 32 symbols and an IFFT size of N = 1024 giving a multitone subcarrier frequency separation of 976.5 a guard interval of 10 frequencies points at the The orthogonal multitone signal was analyzed by using the OMEIS implementation configured with a sampling frequency of = 1024 giving a multitone subcarrier frequency separation of 976.5 a guard interval of 10 frequencies points at the end of the spectrum is applied giving a maximum frequency of 488.28 kHz.Furthermore, to test the frequency selection capabilities of the system, the subcarriers were also eliminated in the range from corresponds to a bandwidt  end of the spectrum is applied giving a maximum frequency of 488.28 kHz.Furthermore, to test the frequency selection capabilities of the system, the subcarriers were also eliminated in the range from corresponds to a bandwidt F(n)=n•Fs/N).A digital oscilloscope (Tektronic DPO500B), with FFT function capabilities, measure the spectrum of the signal during this tes Figure 6 shows the spectrum of the stimulation multitone voltage signal (above) and the voltage signal spectrum of a resistance Rn = 671.8series with the test impedance Rp = 4.62 kΩ intentionally displaced 20 dB below, visibility.
As shown in Figure 6 above, the spectrum of the multitone signal has a flat shape between 0 Hz to 488.281 kHz, with the exception of the removed bandwidth measured between 97.65 kHz and 146.5 kHz.
When the voltage spectrum of the resistance measured (Figure 6 end of the spectrum is applied giving a maximum frequency of 488.28 kHz.Furthermore, to test the frequency selection capabilities of the system, the subcarriers were also eliminated in the range from corresponds to a bandwidth of .A digital oscilloscope (Tektronic with FFT function capabilities, measure the spectrum of the signal during this tes Figure 6 shows the spectrum of the stimulation multitone (above) and the voltage signal spectrum of a = 671.8Ω series with the test impedance Ω, C = 1.1 nF).The second signal was intentionally displaced 20 dB below, As shown in Figure 6 above, the spectrum of the multitone signal has a flat shape between 0 Hz to 488.281 kHz, with the exception of the removed bandwidth measured between 97.65 kHz and 146.5 kHz.
When the voltage spectrum of the resistance measured (Figure 6, below), the amplitude of the spectrum changes as expected: At the frequency of 488.28  end of the spectrum is applied giving a maximum frequency of 488.28 kHz.Furthermore, to test the frequency selection capabilities of the system, the subcarriers were also eliminated in the range from n = 100 to h of 97.65 kHz to 146.48 kHz .A digital oscilloscope (Tektronic with FFT function capabilities, measure the spectrum of the signal during this tes Figure 6 shows the spectrum of the stimulation multitone (above) and the voltage signal spectrum of a (below), that is connected in series with the test impedance (Rs + Rp || = 1.1 nF).The second signal was intentionally displaced 20 dB below, in the Figure, for better As shown in Figure 6 above, the spectrum of the multitone signal has a flat shape between 0 Hz to 488.281 kHz, with the exception of the removed bandwidth measured between 97.65 kHz and 146.5 kHz.
When the voltage spectrum of the resistance measured (Figure 6, below), the amplitude of the spectrum changes as expected: At the frequency of 488.28  end of the spectrum is applied giving a maximum frequency of 488.28 kHz.Furthermore, to test the frequency selection capabilities of the system, the subcarriers were also = 100 to n = 150, which 97.65 kHz to 146.48 kHz .A digital oscilloscope (Tektronic with FFT function capabilities, is used to measure the spectrum of the signal during this test. Figure 6 shows the spectrum of the stimulation multitone (above) and the voltage signal spectrum of a (below), that is connected in || C, Rs = 265.4= 1.1 nF).The second signal was in the Figure, for better As shown in Figure 6 above, the spectrum of the multitone signal has a flat shape between 0 Hz to 488.281 kHz, with the exception of the removed bandwidth measured between 97.65 kHz and 146.5 kHz.
When the voltage spectrum of the resistance Rn measured (Figure 6, below), the amplitude of the spectrum changes as expected: At the frequency of 488.28 kHz, where Ω, the amplitude is less than the multitone 19.37 dB.This very closely corresponds to the theoretical difference of 4.13 dB.At low frequency ) the delta marker shows -16.2 dB that also is very similar to the theoretical   OMEIS System Performance the OMEIS system, the size of the IFFT = 1 MHz.Therefore, there are 51 frequency points for the impedance estimation with a spectrum resolution and first frequency point of 976.5 Hz (DC frequency is not calculated).The stimulation time depends Fs, the size of the IFFT / FFT ( symbols quantity (M = also corresponds to the number of measurement cycles used for averaging.
test circuit (Rs=750 Ω , Rp=30 k Figure 7.The result shows a good accuracy impedance estimation with a 1.39% mean error. Spectrum of the multitone signal.Below: Spectrum of a voltage signal form a voltage divider circuit between the test impedance Z.The value of the test impedance is shown.The ∆V) show the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.
Impedance comparison between model (red) and the estimation from the OMEIS system (blue).
. Spectrum of a resistance signal (spectrum above).The value frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.0 Performance.For the validation of ize of the IFFT Therefore, there are 51 frequency points for the impedance estimation with a spectrum resolution and first frequency point of 976.5 Hz (DC frequency is not calculated).The stimulation time depends on Fs, the size of the IFFT / FFT ( = 32 in this case), also corresponds to the number of measurement cycles used for averaging.This test was performed by using Ω , Rp=30 kΩ, Cp=2 nF), as shown in Figure 7.The result shows a good accuracy impedance estimation with a 1.39% mean error.For the validation of ize of the IFFT and the FFT is 1024 Therefore, there are 511 available frequency points for the impedance estimation with a spectrum resolution and first frequency point of 976.5 Hz (DC frequency is not calculated).The stimulation time depends on Fs, the size of the IFFT / FFT (N = 1024) and 32 in this case), resulting in 32.76 also corresponds to the number of measurement This test was performed by using Ω, Cp=2 nF), as shown in Figure 7.The result shows a good accuracy impedance estimation with a 1.39% mean error.For the validation of and the FFT is 1024 available frequency points for the impedance estimation with a spectrum resolution and first frequency point of 976.5 Hz (DC frequency is not calculated).The stimulation time 1024) and resulting in 32.76 also corresponds to the number of measurement This test was performed by using , Cp=2 nF), as shown in Figure 7.The result shows a good accuracy in the

5.
After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of preliminary results of performed under conditions related describ correlation between the impedance and observed.resolution flexibility is tested as well.impeda perfused porc commercial human PCM probes, for a more realistic condition.

5.1.
flexibility of the OMEIS system and the estimation during biological sample measurements.  .

Experiments
After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of preliminary results of performed under conditions related describe experiments with cell cultures, in which correlation between the impedance and observed.OMEIS's spectrum management and frequency resolution flexibility is tested as well.impedance measurements conducted on inhibited and perfused porc commercial human PCM probes, for a more realistic condition.

In Vitro Measurements.
flexibility of the OMEIS system and the estimation during biological sample measurements.

Experiments
After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of preliminary results of in vitro performed under conditions related e experiments with cell cultures, in which correlation between the impedance and OMEIS's spectrum management and frequency resolution flexibility is tested as well.
nce measurements conducted on inhibited and perfused porcine heart tissue are described, commercial human PCM probes, for a more realistic In Vitro Measurements.flexibility of the OMEIS system and the estimation during biological sample measurements.

Materials.
A commercially available cultureware of Applied Biophysic Inc. [26 during the experimentation.The cultureware array of 8 wells with 10 electrically interconnected electrodes (250µm diameter central larger electrode common to all wells.The electrodes are delineated with an insulating film.The cultureware a two electrodes setup for EIS measurement: the Working Electrode (WE) and the Counter Electrode together with the Reference Electrode ( The OMEIS system was calibrated with a IFFT/FFT size , a symbols quantity of 32 1 MHz, 500 kHz and 250 kHz. the first and the last frequencies ( minimum frequency (Fm), the frequency separatio voltage signal (below) form a voltage divider circuit between Rn and the test impedance V) show the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.

400
After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of in vitro and ex vivo performed under conditions related to fibrosis e experiments with cell cultures, in which correlation between the impedance and the cell population is OMEIS's spectrum management and frequency resolution flexibility is tested as well.
nce measurements conducted on inhibited and ine heart tissue are described, commercial human PCM probes, for a more realistic In Vitro Measurements.These tests focus on the flexibility of the OMEIS system and the estimation during biological sample measurements.
A commercially available cultureware of 6], shown in Figure during the experimentation.The cultureware array of 8 wells with 10 electrically interconnected µm diameter) on each well and one central larger electrode common to all wells.The electrodes are delineated with an insulating film.The cultureware a two electrodes setup for EIS measurement: the ) and the Counter Electrode together with the Reference Electrode (CE and RE).
The OMEIS system was calibrated with a IFFT/FFT size , a symbols quantity of 32 and with 3 values 1 MHz, 500 kHz and 250 kHz.There is a the first and the last frequencies (Fs/N (Fm), the frequency separatio and the test impedance powered by the multitone V) show the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of ex vivo experiments to fibrosis.First, we e experiments with cell cultures, in which the cell population is OMEIS's spectrum management and frequency Finally, ex vivo nce measurements conducted on inhibited and ine heart tissue are described, using commercial human PCM probes, for a more realistic These tests focus on the flexibility of the OMEIS system and the estimation of errors A commercially available cultureware of ], shown in Figure 8, is use during the experimentation.The cultureware consists of an array of 8 wells with 10 electrically interconnected circular on each well and one central larger electrode common to all wells.The electrodes are delineated with an insulating film.The cultureware a two electrodes setup for EIS measurement: the ) and the Counter Electrode together ).The OMEIS system was calibrated with a IFFT/FFT size and with 3 values for There is a GI at DC and at Fs/N and Fs/2).(Fm), the frequency separation (∆F powered by the multitone V) show the amplitude difference, in dB, between both spectrum at the

[KHz]
After having characterized the performances of our system in terms of hardware resources, signal spectrum and impedance estimation, we proceed to the presentation of experiments .First, we e experiments with cell cultures, in which the the cell population is OMEIS's spectrum management and frequency ex vivo nce measurements conducted on inhibited and using commercial human PCM probes, for a more realistic

These tests focus on the of errors
A commercially available cultureware of , is used consists of an circular on each well and one central larger electrode common to all wells.The electrodes are delineated with an insulating film.The cultureware a two electrodes setup for EIS measurement: the ) and the Counter Electrode together The OMEIS system was calibrated with a IFFT/FFT size for Fs GI at DC and at ).The ∆F) powered by the multitone V) show the amplitude difference, in dB, between both spectrum at the frequency up to the 20 kHz, when it begins to decrease.and the stimulation time (Ts) for each configuration are shown in Table 2 5.1.2.Protocol performed on immortalized mouse myoblast cell line (C2C12 cells.)Under that produce proteins related to fibrosis [2 for in vitro cell following: at the beginning of the experiment, one well of the cultureware was filled with 7,000 C2C12 m a 600 µL of Dulbecco's Modified Eagle's Medium (DMEM) containing 10% Fetal Bovine Serum (FBS), sodium bicarbonate (3.7 filled with only 600 well.Cells were cultured i CO 2 and the medium was changed every 48 hours.Five measurements were taken during the 95 hours of incubation, enough time for a development of a large cell population on the electrode.Visual inspections were performed with a standard inverted microscope.

Results.
at five measurement times when 47h, 71h and 95h (h stand for hours after the beginning of the experimentation).In the frequency domain the spectrum shows a negative constant slope, whose magnitude differs depending on the measurement time, from the first frequency up to the 20 kHz, when it begins to decrease.
The evolution of the impedance in time could be better evaluated by using the   2.
Protocol.The in vitro experimentations are performed on immortalized mouse myoblast cell line (C2C12 cells.)Under appropriate conditions, they are cells that produce proteins related to fibrosis [2 for in vitro cell-substrate EIS measurements is the following: at the beginning of the experiment, one well of the cultureware was filled with 7,000 C2C12 m µL of Dulbecco's Modified Eagle's Medium (DMEM) containing 10% Fetal Bovine Serum (FBS), sodium bicarbonate (3.7 g/L) and 1% antibiotics.Other well was filled with only 600 µL of medium and is used as control well.Cells were cultured i and the medium was changed every 48 hours.Five measurements were taken during the 95 hours of incubation, enough time for a development of a large cell population on the electrode.Visual inspections were performed with a standard inverted microscope.
Results. Figure 9.a shows the impedance spectrum at five measurement times when 47h, 71h and 95h (h stand for hours after the beginning of the experimentation).In the frequency domain the spectrum shows a negative constant slope, whose magnitude differs depending on the measurement time, from the first frequency up to the 20 kHz, when it begins to decrease.
The evolution of the impedance in time could be better evaluated by using the The in vitro experimentations are performed on immortalized mouse myoblast cell line appropriate conditions, they are cells that produce proteins related to fibrosis [2 substrate EIS measurements is the following: at the beginning of the experiment, one well of the cultureware was filled with 7,000 C2C12 m L of Dulbecco's Modified Eagle's Medium (DMEM) containing 10% Fetal Bovine Serum (FBS), sodium g/L) and 1% antibiotics.Other well was µL of medium and is used as control well.Cells were cultured in an incubator at 37°C and 5% and the medium was changed every 48 hours.Five measurements were taken during the 95 hours of incubation, enough time for a development of a large cell population on the electrode.Visual inspections were performed with a standard inverted microscope.
.a shows the impedance spectrum at five measurement times when Fs = 1 MHz: times 0h, 23h, 47h, 71h and 95h (h stand for hours after the beginning of the experimentation).In the frequency domain the spectrum shows a negative constant slope, whose magnitude differs depending on the measurement time, from the first frequency up to the 20 kHz, when it begins to decrease.
The evolution of the impedance in time could be better evaluated by using the The in vitro experimentations are performed on immortalized mouse myoblast cell line appropriate conditions, they are cells that produce proteins related to fibrosis [27].The protocol substrate EIS measurements is the following: at the beginning of the experiment, one well of the cultureware was filled with 7,000 C2C12 myoblast plus L of Dulbecco's Modified Eagle's Medium (DMEM) containing 10% Fetal Bovine Serum (FBS), sodium g/L) and 1% antibiotics.Other well was L of medium and is used as control n an incubator at 37°C and 5% and the medium was changed every 48 hours.Five measurements were taken during the 95 hours of incubation, enough time for a development of a large cell population on the electrode.Visual inspections were performed with a .ashows the impedance spectrum = 1 MHz: times 0h, 23h, 47h, 71h and 95h (h stand for hours after the beginning of the experimentation).In the frequency domain the spectrum shows a negative constant slope, whose magnitude differs depending on the measurement time, from the first frequency up to the 20 kHz, when it begins to decrease.
The evolution of the impedance in time could be better .b,where the normalized impedance is depicted, taking time 0h as the reference Measured impedance at point k divided by the Impedance As shown, the impedance at time 23h and frequency 55 almost 3.5, at time 95h and frequency 95 kHz, OMEIS default parameters substrate impedance sensing (ECIS) protocol and the OMEIS system.Also, the picture and the scheme of the ECIS cultureware board (8W10E) with eight mini wells (∼ Ts [ms] 65.5 131 262 and the stimulation time (Ts) for each configuration are The in vitro experimentations are performed on immortalized mouse myoblast cell line appropriate conditions, they are cells The protocol substrate EIS measurements is the following: at the beginning of the experiment, one well of yoblast plus L of Dulbecco's Modified Eagle's Medium (DMEM) containing 10% Fetal Bovine Serum (FBS), sodium g/L) and 1% antibiotics.Other well was L of medium and is used as control n an incubator at 37°C and 5% and the medium was changed every 48 hours.Five measurements were taken during the 95 hours of incubation, enough time for a development of a large cell population on the electrode.Visual inspections were performed with a .ashows the impedance spectrum = 1 MHz: times 0h, 23h, 47h, 71h and 95h (h stand for hours after the beginning of the experimentation).In the frequency domain the spectrum shows a negative constant slope, whose magnitude differs depending on the measurement time, from the first frequency up to the 20 kHz, when it begins to decrease.
The evolution of the impedance in time could be better .b,where the normalized impedance is depicted, taking time 0h as the reference mpedance As shown, the impedance at time 23h and frequency 55 in correspondence with the increase of the cell population in the electrode.This is validated using microscopy photos to visually correlate the state of the culture with the measurements impedances at 3 sampling frequencies are evaluated together.
1 MHz, 500 kHz and 250 kHz that give the frequency resolution of 976 Hz, 488 Hz and 244 Hz, respectively.expected, the impedance at the same substrate impedance sensing (ECIS) protocol and the OMEIS system.Also, the picture and the scheme ∼0.6 mL) correspondence with the increase of the cell population in the electrode.This is validated using microscopy photos to visually correlate the state of the culture with the measurements, Figure 9.cFor the study of the flexibility of the system, the mpedances at 3 sampling frequencies are evaluated together.Here we are using the sampling frequencies of 1 MHz, 500 kHz and 250 kHz that give the frequency resolution of 976 Hz, 488 Hz and 244 Hz, respectively.expected, the impedance at the same correspondence with the increase of the cell population in the electrode.This is validated using microscopy photos to visually correlate the state of the culture with the For the study of the flexibility of the system, the mpedances at 3 sampling frequencies are evaluated Here we are using the sampling frequencies of 1 MHz, 500 kHz and 250 kHz that give the frequency resolution of 976 Hz, 488 Hz and 244 Hz, respectively.expected, the impedance at the same measurement time but correspondence with the increase of the cell population in the electrode.This is validated using microscopy photos to visually correlate the state of the culture with the For the study of the flexibility of the system, the mpedances at 3 sampling frequencies are evaluated Here we are using the sampling frequencies of 1 MHz, 500 kHz and 250 kHz that give the frequency resolution of 976 Hz, 488 Hz and 244 Hz, respectively.As measurement time but  10.This flexibility allows the addition of more frequency points for a better evaluation of the regions of interest.It should be noted that the sampling frequency ca be changed by software, either manually or automatically without the need to reconfigure the system.
The results obtained in these experiments show that the frequency band from 30 kHz to 200 kHz is optimal for the observation of the impedance signat growth of the cell population.at different sampling frequencies overlaps, as shown in the Figure 10.This flexibility allows the addition of more frequency points for a better evaluation of the regions of interest.It should be noted that the sampling frequency ca be changed by software, either manually or automatically without the need to reconfigure the system.The results obtained in these experiments show that the frequency band from 30 kHz to 200 kHz is optimal for the observation of the impedance signat growth of the cell population.at different sampling frequencies overlaps, as shown in the Figure 10.This flexibility allows the addition of more frequency points for a better evaluation of the regions of interest.It should be noted that the sampling frequency ca be changed by software, either manually or automatically without the need to reconfigure the system.The results obtained in these experiments show that the frequency band from 30 kHz to 200 kHz is optimal for the observation of the impedance signature corresponding to the growth of the cell population.

In vitro Experiments in
Experiments Data Modeling in Figure 9.a were used in a impedance model identification algorithm.Due to the bandwidth of the measurements (2 kHz to 500 kHz) and the limited double layer capacitance associated with the micro-electrodes used, th is associated with the cell membrane and the int and extracellular medium [28].The electrode impedance is fitted in our case with the following equation: ce of the C2C12 cells at times 23h, 47 cell culture time in hours), when using 3 different sampling at different sampling frequencies overlaps, as shown in the Figure 10.This flexibility allows the addition of more frequency points for a better evaluation of the regions of interest.It should be noted that the sampling frequency ca be changed by software, either manually or automatically without the need to reconfigure the system.
The results obtained in these experiments show that the frequency band from 30 kHz to 200 kHz is optimal for the ure corresponding to the odeling.Data presented in Figure 9.a were used in a impedance model identification algorithm.Due to the bandwidth of the measurements (2 kHz to 500 kHz) and the limited double layer capacitance electrodes used, the observable is associated with the cell membrane and the int ].The electrode impedance is fitted in our case with the following equation:

reliability during in S
OMEIS ability for experiments are conducted on the cardiac tissues, freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of a solution that inhibits the cardiac contractions and second on perfused tissue that mimic in vivo conditions.

5.2.1.
the OMEIS system and the Solartron 1260 (Ametek, USA), whic spectrometer commonly measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation electrodes, but for this experimentati termination was used.

5.2.2.
accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the local ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 The swine was pre acepromazine (Calmivet, 1mL/50 induced with intravenous inject (10 O 2 (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold cardioplegic 1.2; KCl, 16; MgC 4°C.

5.2.3.
wall was dissected and placed in the cardioplegic solution (in mM: 110 NaCl, 1,2 CaCl NaHCO measurements.ce of the C2C12 cells at times 23h, 47h , when using 3 different sampling In high frequency, the resistance increase over time. of the Constant Phase Element 1, corresponding to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the measurements were conclusion on the cor and the biological evolution of the cells can be drawn.However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data can fit standard electrode impedance model conventional optimization algorithms.

Ex Vivo Measurements
reliability during in Section 5.1.The objective now is the evaluation of OMEIS ability for experiments are conducted on the cardiac tissues, freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of a swine heart: first on a tissue immersed in a car solution that inhibits the cardiac contractions and second on perfused tissue that mimic in vivo conditions.

Materials
the OMEIS system and the Solartron 1260 (Ametek, USA), whic spectrometer commonly measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation electrodes, but for this experimentati termination was used.

Animal Model.
accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the cal ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 The swine was pre acepromazine (Calmivet, 1mL/50 induced with intravenous inject (10 mg/kg) and maintained under isofluorane, 2%, in 100% 2 .The swine was euthanized by sodium pentobarbital (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold cardioplegic solution, containing (mM): NaCl, 110; CaC 1.2; KCl, 16; MgC 4°C.
In high frequency, the resistance over time.Nevertheless Constant Phase Element 1, corresponding to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the s were performed over a single well, no conclusion on the correlation between the model parameters iological evolution of the cells can be drawn.However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data can fit standard electrode impedance model conventional optimization algorithms.

Ex Vivo Measurements reliability during impedance measurement have been
.
The objective now is the evaluation of OMEIS ability for in vivo experiments are conducted on the cardiac tissues, freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of swine heart: first on a tissue immersed in a car solution that inhibits the cardiac contractions and second on perfused tissue that mimic in vivo conditions.
Materials.The measurements were performed with the OMEIS system and the Solartron 1260 (Ametek, USA), whic spectrometer commonly measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation electrodes, but for this experimentati termination was used.
Animal Model.This study was carried out in accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the cal ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 The swine was pre-medicated with ketamine (20 mg/kg acepromazine (Calmivet, 1mL/50 induced with intravenous inject mg/kg) and maintained under isofluorane, 2%, in 100% .The swine was euthanized by sodium pentobarbital (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold solution, containing (mM): NaCl, 110; CaC 1.2; KCl, 16; MgCl 2 , 16; NaHCO Protocol in Cardioplegia wall was dissected and placed in the cardioplegic solution (in mM: 110 NaCl, 1,2 CaCl , 10 glucose) and then In high frequency, the resistance R Nevertheless, there is a clear evolution Constant Phase Element (CPE) order from values near 1, corresponding to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the performed over a single well, no relation between the model parameters iological evolution of the cells can be drawn.However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data can fit standard electrode impedance model conventional optimization algorithms.
Ex Vivo Measurements.The OMEIS flexibility and impedance measurement have been .The objective now is the evaluation of in vivo conditions.The following experiments are conducted on the cardiac tissues, freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of swine heart: first on a tissue immersed in a car solution that inhibits the cardiac contractions and second on perfused tissue that mimic in vivo conditions.
The measurements were performed with the OMEIS system and the commercial Solartron 1260 (Ametek, USA), which is used for bioimpedance measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation electrodes, but for this experimentation only the pacing This study was carried out in accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the cal ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 medicated with ketamine (20 mg/kg acepromazine (Calmivet, 1mL/50kg).Anesthesia was induced with intravenous injection of sod mg/kg) and maintained under isofluorane, 2%, in 100% .The swine was euthanized by sodium pentobarbital (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold solution, containing (mM): NaCl, 110; CaC , 16; NaHCO 3 , 10; and glucose, 9.01 at in Cardioplegia.The right ventricle (RV) wall was dissected and placed in the cardioplegic solution (in mM: 110 NaCl, 1,2 CaCl 2 , 16 KCl, and then chilled wi R shows a slight , there is a clear evolution order from values near 1, corresponding to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the performed over a single well, no relation between the model parameters iological evolution of the cells can be drawn.However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data can fit standard electrode impedance model using The OMEIS flexibility and impedance measurement have been proven .The objective now is the evaluation of conditions.The following experiments are conducted on the cardiac tissues, ex vivo freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of swine heart: first on a tissue immersed in a cardioplegic solution that inhibits the cardiac contractions and second on perfused tissue that mimic in vivo conditions. The measurements were performed with commercial lab instrument h is an impedance used for bioimpedance measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation on only the pacing This study was carried out in accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the cal ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 medicated with ketamine (20 mg/kg kg).Anesthesia was ion of sodium pentobarbital mg/kg) and maintained under isofluorane, 2%, in 100% .The swine was euthanized by sodium pentobarbital (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold solution, containing (mM): NaCl, 110; CaC , 10; and glucose, 9.01 at The right ventricle (RV) wall was dissected and placed in the cardioplegic solution , 16 KCl, 16 MgCl 2 chilled with ice during the shows a slight , there is a clear evolution order from values near 1, corresponding to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the performed over a single well, no relation between the model parameters iological evolution of the cells can be drawn.However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data using The OMEIS flexibility and proven .The objective now is the evaluation of conditions.The following ex vivo, freshly explanted, using a commercial pacemaker electrode.Two types of characterization are performed on ventricles of dioplegic solution that inhibits the cardiac contractions and second on The measurements were performed with lab instrument an impedance used for bioimpedance measurements.A human PCM cardiac lead, Sprint Quattro Secure 6947M (Medtronic, USA) is used as sensing electrode.This lead has both pacing and defibrillation on only the pacing This study was carried out in accordance with the recommendations of the Directive 2010/63/EU of the European Parliament on the protection of animals used for scientific purposes and approved by the cal ethical committee of Bordeaux CEEA50.The heart was obtained from a young swine (Large White, 40±5 kg).medicated with ketamine (20 mg/kg) and kg).Anesthesia was ium pentobarbital mg/kg) and maintained under isofluorane, 2%, in 100% .The swine was euthanized by sodium pentobarbital (40 mL, from 50 mg/mL of stock) and the heart rapidly excised, cannulated by the aorta, and rinsed with cold solution, containing (mM): NaCl, 110; CaCl 2 , , 10; and glucose, 9.01 at The right ventricle (RV) wall was dissected and placed in the cardioplegic solution The OMEIS system was configured of size N = 1024 Solartron 1260 peak voltage level (simil logarithmic frequency sweep from 1 MHz to 0.1 Hz, with 10 points per decade.For OMEIS, each measure was carried out only once, for the Solartron they were repeated five times and the mean of these five The ex vivo different endocardial regions of the ventricle: lateral wall and a more 'collagen' rich region near the basal area and auriculo endocardium tissue will be termed 'Muscle' and will be compared to the 'Collagen' region the lead is placed in Figure 1 region just above the tri 11.)

Protocol
Langendorff system used for the perfusion protocol in this section: The left ventricle (LV) was dissected and cannulated by the left anterior descending artery.The LV was mounted on to a frame perfused (20 mL/min) with a warm (37°C) saline solution containing (mM): NaCl, 130; NaHCO MgCl 2 , 1; glucose, 5.6; KCl, 4; CaCl O 2 /5% CO 2 at 37°C (pH 7.4).A v was measured in the bath to monitor the electrical activity of the tissue preparation.
The ventricle was stimulated from the epicardium at 1 Hz using bipolar tungsten electrodes coupled to a pulse generator and constant current sti Digitimer, UK).twice the excitation threshold for a duration of 2ms.OMEIS system was configured 1024, 32 symbols Solartron 1260, the measurements were done at a 100 mV voltage level (similar to the OMEIS system), with a logarithmic frequency sweep from 1 MHz to 0.1 Hz, with 10 points per decade.For OMEIS, each measure was carried out only once, for the Solartron they were repeated five times and the mean of these five vivo measurements were performed in two different endocardial regions of the ventricle: lateral wall and a more 'collagen' rich region near the basal and auriculo-ventricular ring.From now on, the normal endocardium tissue will be termed 'Muscle' and will be compared to the 'Collagen' region the lead is placed in Figure 1 region just above the tricuspid valve (white tissue in Protocol for Perfused Tissue.Langendorff system used for the perfusion protocol in this section: The left ventricle (LV) was dissected and cannulated by the left anterior descending artery.The LV was mounted on to a frame perfused (20 mL/min) with a warm (37°C) saline solution containing (mM): NaCl, 130; NaHCO , 1; glucose, 5.6; KCl, 4; CaCl at 37°C (pH 7.4).A v was measured in the bath to monitor the electrical activity of the tissue preparation.

Ring electrode
The ventricle was stimulated from the epicardium at 1 Hz using bipolar tungsten electrodes coupled to a pulse generator and constant current sti Digitimer, UK).The stimulation currents were applied at twice the excitation threshold for a duration of 2ms.

Ring electrode Helix electrode
Ex vivo experimentation on a swine heart.placed in cardioplegic solution.'Muscle' region.
OMEIS system was configured , 32 symbols and a Fs the measurements were done at a 100 mV ar to the OMEIS system), with a logarithmic frequency sweep from 1 MHz to 0.1 Hz, with 10 points per decade.For OMEIS, each measure was carried out only once, for the Solartron they were repeated five times and the mean of these five measurements is calculated.
measurements were performed in two different endocardial regions of the ventricle: lateral wall and a more 'collagen' rich region near the basal ventricular ring.From now on, the normal endocardium tissue will be termed 'Muscle' and will be compared to the 'Collagen' region.The first region is wher the lead is placed in Figure 11, and the second one is the cuspid valve (white tissue in for Perfused Tissue.Figure 1 Langendorff system used for the perfusion protocol in this section: The left ventricle (LV) was dissected and cannulated by the left anterior descending artery.The LV was mounted on to a frame where it is perfused (20 mL/min) with a warm (37°C) saline solution containing (mM): NaCl, 130; NaHCO 3 , 1; glucose, 5.6; KCl, 4; CaCl 2 , 1.8; gassed with 95% at 37°C (pH 7.4).A volume was measured in the bath to monitor the electrical activity of The ventricle was stimulated from the epicardium at 1 Hz using bipolar tungsten electrodes coupled to a pulse generator and constant current stimulator (DG2A, DS3, The stimulation currents were applied at twice the excitation threshold for a duration of 2ms.

Muscle
experimentation on a swine heart.placed in cardioplegic solution.Pacemaker lead in the endocardial OMEIS system was configured with an IFFT/FFT Fs = 1 MHz.For the the measurements were done at a 100 mV ar to the OMEIS system), with a logarithmic frequency sweep from 1 MHz to 0.1 Hz, with 10 per decade.For OMEIS, each measure was carried out only once, for the Solartron they were repeated five measurements is calculated.measurements were performed in two different endocardial regions of the ventricle: the healthy lateral wall and a more 'collagen' rich region near the basal ventricular ring.From now on, the normal endocardium tissue will be termed 'Muscle' and will be .The first region is wher , and the second one is the cuspid valve (white tissue in Figure 14 shows the Langendorff system used for the perfusion protocol in this section: The left ventricle (LV) was dissected and cannulated by the left anterior descending artery.The where it is submerg perfused (20 mL/min) with a warm (37°C) saline solution 3 , 24; NH 2 PO , 1.8; gassed with 95% olume-conducted ECG was measured in the bath to monitor the electrical activity of The ventricle was stimulated from the epicardium at 1 Hz using bipolar tungsten electrodes coupled to a pulse mulator (DG2A, DS3, The stimulation currents were applied at twice the excitation threshold for a duration of 2ms.

Collagen
experimentation on a swine heart.Right V Pacemaker lead in the endocardial IFFT/FFT 1 MHz.For the the measurements were done at a 100 mV ar to the OMEIS system), with a logarithmic frequency sweep from 1 MHz to 0.1 Hz, with 10 points per decade.For OMEIS, each measure was carried out only once, for the Solartron they were repeated five measurements is calculated.measurements were performed in two the healthy lateral wall and a more 'collagen' rich region near the basal ventricular ring.From now on, the normal endocardium tissue will be termed 'Muscle' and will be .The first region is where , and the second one is the cuspid valve (white tissue in Figure shows the Langendorff system used for the perfusion protocol in this section: The left ventricle (LV) was dissected and cannulated by the left anterior descending artery.Then, the erged and perfused (20 mL/min) with a warm (37°C) saline solution PO 4 , 1.2; , 1.8; gassed with 95% conducted ECG was measured in the bath to monitor the electrical activity of The ventricle was stimulated from the epicardium at 1 Hz using bipolar tungsten electrodes coupled to a pulse mulator (DG2A, DS3, The stimulation currents were applied at 5.2.5. the following, measurement first performed wi the different h solution (CPG) the followed by the 'collagen' region and lastly the CPG solution that has a much low fact that a reduced number of symbols order to increase measureme averaged version fo reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation between the impedance of the tissues.

5.2.6.
During the experimentation with the heart in state, the OMEIS system delivered valid results the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electro physiological and mechani behave by an instru OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' tissue has a greater impedance t both lower than the values described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here compl submerged in the saline solution.lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring)

Results of Inhibited Cardiac Tissue Experiments. the following, measurement first performed wi
Figure 12 shows the impedance spectrum measured in the different h solution (CPG) the 'Muscle' has followed by the 'collagen' region and lastly the CPG solution that has a much low The values of the OMEIS system were noisy due to the fact that a reduced number of symbols order to increase measureme averaged version fo reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation between the impedance of the tissues.

Results of Cardiac Tissue Perfusion Experiments
During the experimentation with the heart in state, the OMEIS system delivered valid results the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electro physiological and mechani behave like a time invariant system that could by an instrument that uses the frequency sweep EIS method.
Figure 13 shows the impedance values received from the OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' tissue has a greater impedance t both lower than the values described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here compl submerged in the saline solution.
Furthermore, the Sprint Quattro Secure 6947M pacing lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring)

Results of Inhibited Cardiac Tissue Experiments.
the following, measurements first performed with Solartron then with OMEIS.
shows the impedance spectrum measured in the different heart regions as well in the C solution (CPG) by both instruments has the greatest magnitude of impedance, followed by the 'collagen' region and lastly the CPG solution that has a much lower impedance of around 190 The values of the OMEIS system were noisy due to the fact that a reduced number of symbols order to increase measureme averaged version follows very closely the values reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation between the impedance of the tissues.

Results of Cardiac Tissue Perfusion Experiments
During the experimentation with the heart in state, the OMEIS system delivered valid results the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electro physiological and mechanical activities of the heart.The a time invariant system that could ment that uses the frequency sweep EIS method.
shows the impedance values received from the OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' tissue has a greater impedance t both lower than the values obtained in the first experience described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here compl submerged in the saline solution.
Furthermore, the Sprint Quattro Secure 6947M pacing lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring) Magnitude of impedance measurement for 3 conditions: e' stands for normal endocardial region and 'CPG' for Cardioplegia.OFDM is performed at Fs = 1MHz.Colored curves represent OMEIS measurements, grey curves represent n measurements.
Results of Inhibited Cardiac Tissue Experiments.s of impedance spectrum w th Solartron then with OMEIS.shows the impedance spectrum measured in eart regions as well in the C by both instruments.It can be noticed t the greatest magnitude of impedance, followed by the 'collagen' region and lastly the CPG er impedance of around 190 The values of the OMEIS system were noisy due to the fact that a reduced number of symbols (M = 32 order to increase measurement speed.However, the llows very closely the values reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation between the impedance of the tissues.

Results of Cardiac Tissue Perfusion Experiments
During the experimentation with the heart in state, the OMEIS system delivered valid results the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electro cal activities of the heart.The a time invariant system that could ment that uses the frequency sweep EIS method.
shows the impedance values received from the OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' tissue has a greater impedance than the 'Collagen' although, obtained in the first experience described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here compl submerged in the saline solution.
Furthermore, the Sprint Quattro Secure 6947M pacing lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring) of impedance measurement for 3 conditions: e' stands for normal endocardial region, 'Collagen' for collagen region and 'CPG' for Cardioplegia.OFDM is performed at Fs = 1MHz.Colored curves represent OMEIS measurements, grey curves represent

Results of Inhibited Cardiac Tissue Experiments.
of impedance spectrum w th Solartron then with OMEIS.shows the impedance spectrum measured in eart regions as well in the Cardioplegi .It can be noticed t the greatest magnitude of impedance, followed by the 'collagen' region and lastly the CPG er impedance of around 190 The values of the OMEIS system were noisy due to the M = 32) was used in nt speed.However, the llows very closely the values of the reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation

Results of Cardiac Tissue Perfusion Experiments
During the experimentation with the heart in the perfusion state, the OMEIS system delivered valid results, however, the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electro cal activities of the heart.The a time invariant system that could not be sensed ment that uses the frequency sweep EIS method.
shows the impedance values received from the OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' han the 'Collagen' although, obtained in the first experience described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here compl Furthermore, the Sprint Quattro Secure 6947M pacing lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring) of impedance measurement for 3 conditions: region, 'Collagen' for collagen region and 'CPG' for Cardioplegia.OFDM is performed at Fs = 1MHz.

Colored curves represent OMEIS measurements, grey curves represent
Results of Inhibited Cardiac Tissue Experiments.In of impedance spectrum were shows the impedance spectrum measured in ardioplegic .It can be noticed that the greatest magnitude of impedance, followed by the 'collagen' region and lastly the CPG er impedance of around 190 Ω.The values of the OMEIS system were noisy due to the was used in nt speed.However, the of the reference instrument with a mean error of about 8% and a standard deviation of 5.8%.This allow a differentiation

Results of Cardiac Tissue Perfusion Experiments.
perfusion however, the Solartron yielded meaningless values.We attribute this anomaly, in the reference instrument, to the electrocal activities of the heart.These not be sensed ment that uses the frequency sweep EIS method.
shows the impedance values received from the OMEIS in the perfusion condition, for the 'Muscle' and 'Collagen' regions.It can be noticed that again the 'Muscle' han the 'Collagen' although, obtained in the first experience described above.Experimental conditions differ indeed largely, especially in temperature, and also in the fact that both the heart and the electrode were here completely Furthermore, the Sprint Quattro Secure 6947M pacing lead has two poles, the proximal electrode (helix electrode) is inserted in the tissue and the distal electrode (larger ring) of impedance measurement for 3 conditions: region, 'Collagen' for collagen region and 'CPG' for Cardioplegia.OFDM is performed at Fs = 1MHz.Colored curves represent OMEIS measurements, grey curves represent is used as the returning cardiac tissue is immobilized and the ring electrode relies simply on the tissue, as can be seen in Figure 11.In the second experiment, the perfused cardiac tissue is immersed and exhibits contractions; the ring electrode more in contact with the tissue but moves in the liquid.However, with the impedance measurement the differentiation between both tissues is possible, even with the dynamic of the heart imposed by the perfusion system.

Conclusion
In this article we propose a detection of fibrosis induced by the electrodes of cardiac implants.This strategy Electrical Impedance S sensing fibrosis at short is used as the returning curren cardiac tissue is immobilized and the ring electrode relies simply on the tissue, as can be seen in Figure 11.In the second experiment, the perfused cardiac tissue is immersed and exhibits contractions; the ring electrode more in contact with the tissue but moves in the liquid.However, with the impedance measurement the differentiation between both tissues is possible, even with the dynamic of the heart imposed by the perfusion system.

Conclusion and Perspectives
In this article we propose a detection of fibrosis induced by the electrodes of cardiac implants.This strategy Electrical Impedance Spectroscopy (EIS) as a techniq sensing fibrosis at short intervals between the Magnitude of impedance regions with the heart in perfusion, measured by Figure 14.current path.In the first experiment, cardiac tissue is immobilized and the ring electrode relies simply on the tissue, as can be seen in Figure 11.In the second experiment, the perfused cardiac tissue is immersed and exhibits contractions; the ring electrode more in contact with the tissue but moves in the liquid.However, with the impedance measurement the differentiation between both tissues is possible, even with the dynamic of the heart imposed by the perfusion system.

and Perspectives
In this article we propose a novel strategy for the chronic detection of fibrosis induced by the electrodes of cardiac implants.This strategy consists of the application of pectroscopy (EIS) as a techniq intervals between the Magnitude of impedance of the Muscle and the Collagen regions with the heart in perfusion, measured by Figure 14.Langendorff system maintaining the ventricle in the in vi t path.In the first experiment, cardiac tissue is immobilized and the ring electrode relies simply on the tissue, as can be seen in Figure 11.In the second experiment, the perfused cardiac tissue is immersed and exhibits contractions; the ring electrode is certainly no more in contact with the tissue but moves in the liquid.However, with the impedance measurement the differentiation between both tissues is possible, even with the dynamic of the heart imposed by the perfusion system.strategy for the chronic detection of fibrosis induced by the electrodes of cardiac consists of the application of pectroscopy (EIS) as a techniq intervals between the peacemake of the Muscle and the Collagen regions with the heart in perfusion, measured by the OMEIS system at Langendorff system maintaining the ventricle in the in vi the perfusion chamber is shown at right.t path.In the first experiment, cardiac tissue is immobilized and the ring electrode relies simply on the tissue, as can be seen in Figure 11.In the second experiment, the perfused cardiac tissue is immersed is certainly no more in contact with the tissue but moves in the liquid.However, with the impedance measurement the differentiation between both tissues is possible, even with the dynamic of the heart imposed by the perfusion system.strategy for the chronic detection of fibrosis induced by the electrodes of cardiac consists of the application of pectroscopy (EIS) as a technique for peacemaker pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the constraint of we have devised, orthogonal multitone signals c implementation method Multitone EIS based platform frequency and and offers a maximum OFDM multitone signal bandwidth of 499 MHz with 511 subcarriers separated system circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount of 32 OFDM symbol capacity OMEIS system were stu living cell in between the magnitude of the impedance and the cell population at 250 kHz, when measuring in different times C2C12 cel cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
validated by inhibited and in perfusion states.In the first case, the OMEIS system showed averaged, were similar to that of the with an 8% average error and 5% standard deviation.I second case, given the dynamics of the perfusion system, only the OMEIS per of the Muscle and the Collagen the OMEIS system at Langendorff system maintaining the ventricle in the in vi the perfusion chamber is shown at right.
pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the constraint of low we have devised, orthogonal multitone signals c implementation method Multitone EIS -This proposed based platform frequency and IFFT/FFT size, with a default value of and 1024, respectively offers a maximum OFDM multitone signal bandwidth of 499 MHz with 511 subcarriers separated system is validated circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount of 32 OFDM symbol capacity of biological samples and the flexibility of the OMEIS system were stu living cell in in vitro between the magnitude of the impedance and the cell population at sampling frequencies of 1 MHz 250 kHz, when measuring in different times C2C12 cel cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
Finally, the strategy mentioned at the beginning has been validated by ex vivo inhibited and in perfusion states.In the first case, the OMEIS system showed averaged, were similar to that of the with an 8% average error and 5% standard deviation.I second case, given the dynamics of the perfusion system, only the OMEIS per Langendorff system maintaining the ventricle in the in vivo conditions the perfusion chamber is shown at right.
pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the low hardware cost that this st we have devised, as a innovative orthogonal multitone signals c implementation method, resulting in the Orthogonal -OMEIS-approach.This proposed OMEIS system based platform that provides an adjustable IFFT/FFT size, with a default value of , respectively.With the default offers a maximum OFDM multitone signal bandwidth of 499 MHz with 511 subcarriers separated validated through experiments with circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount of 32 OFDM symbols.In addition, the of biological samples and the flexibility of the OMEIS system were studied by in vitro cultures.It showed between the magnitude of the impedance and the cell sampling frequencies of 1 MHz 250 kHz, when measuring in different times C2C12 cel cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
Finally, the strategy mentioned at the beginning has been ex vivo measurements of cardiac tissue in inhibited and in perfusion states.In the first case, the OMEIS system showed impedance spectrum averaged, were similar to that of the with an 8% average error and 5% standard deviation.I second case, given the dynamics of the perfusion system, only the OMEIS performed measurements as expected.The vo conditions.A picture of the heart inside pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the cost that this st innovative solution orthogonal multitone signals combined with the OFDM , resulting in the Orthogonal approach.system is prototyped on a FPGA that provides an adjustable IFFT/FFT size, with a default value of .With the default offers a maximum OFDM multitone signal bandwidth of 499 MHz with 511 subcarriers separated through experiments with circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount s.In addition, the of biological samples and the flexibility of the died by mean of cultures.It showed a good correlation between the magnitude of the impedance and the cell sampling frequencies of 1 MHz 250 kHz, when measuring in different times C2C12 cel cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
Finally, the strategy mentioned at the beginning has been measurements of cardiac tissue in inhibited and in perfusion states.In the first case, the impedance spectrum averaged, were similar to that of the reference instrument, with an 8% average error and 5% standard deviation.I second case, given the dynamics of the perfusion system, formed measurements as expected.The . A picture of the heart inside pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the cost that this strategy imposes, solution, the use of ombined with the OFDM , resulting in the Orthogonal otyped on a FPGA that provides an adjustable sampling IFFT/FFT size, with a default value of 1 MHz setup, the system offers a maximum OFDM multitone signal bandwidth of 499 MHz with 511 subcarriers separated 976.5 Hz.
through experiments with electrical circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount s.In addition, the measurement of biological samples and the flexibility of the mean of measurements of a good correlation between the magnitude of the impedance and the cell sampling frequencies of 1 MHz, 500 kHz and 250 kHz, when measuring in different times C2C12 cel cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
Finally, the strategy mentioned at the beginning has been measurements of cardiac tissue in inhibited and in perfusion states.In the first case, the impedance spectrum that, reference instrument, with an 8% average error and 5% standard deviation.I second case, given the dynamics of the perfusion system, formed measurements as expected.The pulses and the heartbeat.Given the speed and flexibility requirements in the measurement, in addition to the rategy imposes, the use of ombined with the OFDM , resulting in the Orthogonal otyped on a FPGA sampling 1 MHz setup, the system offers a maximum OFDM multitone signal bandwidth of This electrical circuits resulting in an impedance spectrum with an average error of 1.39% when using 511 frequencies and an amount measurement of biological samples and the flexibility of the measurements of a good correlation between the magnitude of the impedance and the cell , 500 kHz and 250 kHz, when measuring in different times C2C12 cells cultivated during a period of 95 hours.The adjustment of the sampling frequency can be done manually or automatically, on the flight, without the need to reconfigure the system.
Finally, the strategy mentioned at the beginning has been measurements of cardiac tissue in inhibited and in perfusion states.In the first case, the once reference instrument, with an 8% average error and 5% standard deviation.In the second case, given the dynamics of the perfusion system, formed measurements as expected.The acquired values from OMEIS allow us to distinguish between 'Muscular' tissue from the 'Collagen' tissue.Since the cells used for the in vitro experimentation produce collagen in the extracellular matrix, and due the fact that fibrosis tissue is composed mainly by collagen (with few cells), we have performed both experimentation (in vitro and ex vivo) in order to study the OMEIS system in both scenarios.As shown in the in vitro results, when the cells reproduce the impedance also increases, therefore as it is shown in the ex vivo experimentation, the "collagen" region has a lower impedance than that of the "muscle" (full of cells) tissue.
The performance in the measurement speed is inside the requirements, giving 510 frequency points in a stimulation time of about 64 ms with the OMEIS parameters: N = 1024, M = 32 and Fs = 1 MHz.
The tradeoff between the noise reduction and the symbol quantity is a critical step in the calibration of the system.We were using 32 symbols that showed good results during electrical circuits and in vitro experimentations, but that was in the limit for the ex vivo experimentation.The increase of the symbols quantity should solve the problem with the cost of reducing the measurement speed, however this solution should be carefully analyzed in order to remain in the time frame required for the proposed strategy.Another solution could be the use of another OFDM code, that with the appropriated modulation scheme, could improve better the CF.These are topics for future research in our group to further improve the OMEIS system since it is the tool that will allow us to establish an electrical signature of a fibrotic tissue.

Fs
bits DAC converter (DAC7821), a parallel outpu ADC (ADS850Y) and an A depicted in Figure stimulation voltage amplitude of 200 mV, low impedance and a very low Electrode (WE required for biological mea filters non desired DC and 50/60 multitone signal.The stimulation signal is voltage and the resulted current is capture Transimpedance A feedback resistor is set during calibration.

Figure 4 .
Figure 4.The OMEIS System board.bits DAC converter (DAC7821), a parallel outpu ADC (ADS850Y) and an Analog Front E .This AFE board has stimulation voltage amplitude of 200 mV, low dance a very low voltage offset between Working Counter Electrode ( required for biological measurements.Additionally, t filters non desired DC and 50/60 Hz components from the multitone signal.The stimulation signal is voltage and the resulted current is capture and converted into voltage via a mplifier (TIA) in which feedback resistor is set during calibration.the demodulator is implemented the FFT Megacores of Altera.The size of the FFT is the IFFT.Both the calibration and the oming from the average block are (W9825G6EH).Then they The PC runs a Human , coded in Matlab, for the control of the system as for the analysis of the data received running equation r the final impedance estimation.The OFDM TH , coded in VHDL, employs a threshold algorithm to detect the pilot signals and compute the transmitted signal output delay and the system response delay to send them to cost in hardware resource function of the IFFT size 1024 are depicted on the plots.number of multipliers is 24 for all N of frequencies of the multit

Figure 4 .
Figure 4.The OMEIS System board.bits DAC converter (DAC7821), a parallel output 14 bits og Front End (AFE) board, as .This AFE board has maximum stimulation voltage amplitude of 200 mV, low voltage offset between Working lectrode (CE), which are Additionally, the AFE Hz components from the multitone signal.The stimulation signal is voltage and the and converted into voltage via a ) in which the values of feedback resistor is set during calibration.the demodulator is implemented .The size of the FFT is the IFFT.Both the calibration and the average block are stored in a .Then they are sent via Human-Machine Interface , coded in Matlab, for the control of the system as for the analysis of the data received running equation on.The OFDM TH , coded in VHDL, employs a threshold algorithm to detect the pilot signals and compute the transmitted signal output delay and the system response delay to send them to hardware resources function of the IFFT size N.The values 1024 are depicted on the plots.N. Notices that of frequencies of the multitone signal.

Figure 4 .
Figure 4.The OMEIS System board.t14 bits nd (AFE) board, as maximum stimulation voltage amplitude of 200 mV, low output voltage offset between Working ), which are he AFE Hz components from the multitone signal.The stimulation signal is voltage and the and converted into voltage via a lues of a the demodulator is implemented .The size of the FFT is the IFFT.Both the calibration and the stored in a are sent via nterface , coded in Matlab, for the control of the system as for the analysis of the data received running equation on.The OFDM TH , coded in VHDL, employs a threshold algorithm to detect the pilot signals and compute the transmitted signal output delay and the system response delay to send them to of the .The values 1024 are depicted on the plots.The Notices that N is one signal.

Figure 5 .
Figure 5. implemented in a Cyclone IV FPGA.
Figure 5. Hardware resource utilization of the OMEIS emitter implemented in a Cyclone IV FPGA.

Figure 6
Figure 6 shows the spectrum of the stimulation multitone (above) and the voltage signal spectrum of a (below), that is connected in = 265.4Ω, = 1.1 nF).The second signal was in the Figure, for betterAs shown in Figure6above, the spectrum of the multitone signal has a flat shape between 0 Hz to 488.281 kHz, with the exception of the removed bandwidth Rn is measured (Figure6, below), the amplitude of the spectrum changes as expected: At the frequency of 488.28 kHz, where , the amplitude is less than the multitone 19.37 dB.This very closely corresponds to the theoretical difference of 4.13 dB.At low frequency 16.2 dB the theoretical

Figure 6 .
Figure 6.Above: Spectrum of the multitone signal.Below: Spectrum of a resistance Rn voltage signal form a voltage divider circuit between the test impedance markers (∆V) s spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.

Figure 7 .
Figure 7. Impedance comparison between model (red) and the estimation from the Above: Spectrum of the multitone signal.Below: Spectrum of a voltage signal form a voltage divider circuit between .The value of the test impedance is shown.The how the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.Impedance comparison between model (red) and the system (blue).. Spectrum of a resistance Rn voltage signal (below) form a voltage divider circuit between signal (spectrum above).The values of the test impedance is shown.The markers ( frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.
Above: Spectrum of the multitone signal.Below: Spectrum of a voltage signal form a voltage divider circuit between .The value of the test impedance is shown.The how the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.Impedance comparison between model (red) and the voltage signal (below) form a voltage divider circuit between impedance is shown.The markers ( frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.

5. 1 . 1 .
Applied Biophysic Inc. [ during the experimentation.The cultureware array of 8 wells with 10 electrically interconnected gold central larger electrode common to all wells.The electrodes are delineated with an insulating film.The cultureware offers Working Electrode ( with the Reference Electrode ( N=1024 of the first and the last frequencies ( minimum frequency Above: Spectrum of the multitone signal.Below: Spectrum of a voltage signal form a voltage divider circuit between Rn and .The value of the test impedance is shown.The how the amplitude difference, in dB, between both spectrum at the frequency of 19.53 kHz and 488.28 kHz.The second Impedance comparison between model (red) and the voltage signal (below) form a voltage divider circuit between impedance is shown.The markers ( frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.

Figure 8 .
Figure 8.The circuit of the electric cell (ECIS) protocol and the OMEIS system.Also, the picture and the scheme of the ECIS cultureware board (8W10E) with eight are depicted.
Figure impedance is depicted, taking time 0h as the reference [Measured impedance at point k divided by the I of time 0h at the same point k].module from 1.5 kHz to almost 3.5, at time 95h and frequency 95 kHz,

Figure 8 .Fs
Figure 8.The circuit of the electric cell (ECIS) protocol and the OMEIS system.Also, the picture and the scheme of the ECIS cultureware board (8W10E) with eight are depicted.

Figure 8 .Fs
Figure 8.The circuit of the electric cell (ECIS) protocol and the OMEIS system.Also, the picture and the scheme of the ECIS cultureware board (8W10E) with eight

Figure 9 .Figure 8 .
Figure 8.The circuit of the electric cell-substrate impedance sensing (ECIS) protocol and the OMEIS system.Also, the picture and the scheme of the ECIS cultureware board (8W10E) with eight

Figure 9 .
Figure 9. Impedance spectrum performed on C2C12 cells cultured Five measures were taken at 0, 23, 47, 71 and 95 hours.a) Impedance Module.b) Normalized impedance.c) Microscopy photos at 0h, 23h and 71h.The scale bar represents Impedance spectrum performed on C2C12 cells cultured Five measures were taken at 0, 23, 47, 71 and 95 hours.a) Impedance Module.b) Normalized impedance.c) Microscopy photos at 0h, 23h and µm.0h 71 correspondence with the increase of the cell population in the electrode.This is validated using microscopy photos to visually correlate the state of the culture with the For the study of the flexibility of the system, the mpedances at 3 sampling frequencies are evaluated Here we are using the sampling frequencies of 1 MHz, 500 kHz and 250 kHz that give the frequency resolution of 976 Hz, 488 Hz and 244 Hz, respectively.measurement time but Impedance spectrum performed on C2C12 cells cultured in vitro Five measures were taken at 0, 23, 47, 71 and 95 hours.a) Impedance Module.b) Normalized impedance.c) Microscopy photos at 0h, in vitro.Five measures were taken at 0, 23, 47, 71 and 95 hours.a) Impedance Module.b) Normalized impedance.c) Microscopy photos at 0h, 23h and h proliferation Few adhered and spread cells at different sampling frequencies overlaps, as shown in the Figure

5. 1 . 4 .
In vitro in Figure9.a were used in a impedance model identification algorithm.Due to the bandwidth of the measurements (2 kHz to 500 kHz) and the limited double layer capacitance associated with the micro behavior is associated with the cell membrane and the int and extracellular medium [2 fitted in our case with the following equation:where R is fractional order of the constant phase element and transition frequency.Data were fit by minimizing the residual function of the distance between the measurement and the modeled using a Powell method [2 following table:

Figure 10 .
Figure 10.Normalized impedan and 71h (h = cell culture time in hours frequencies.

Figure 9 .
a were used in a impedance model identification algorithm.Due to the bandwidth of the measurements (2 kHz to 500 kHz) and the limited double layer capacitance associated with the micro is associated with the cell membrane and the int and extracellular medium [2 fitted in our case with the following equation: 0 1213 (4 the resistance at high frequency, fractional order of the constant phase element and transition frequency.Data were fit by minimizing the residual function of the distance between the measurement and the modeled using a Powell method [2 following table:

TABLE 3 :
Data fit parameters resultsNormalized impedance of the C2C12 cells at times 23h, 47 cell culture time in hours high frequency, fractional order of the constant phase element and Data were fit by minimizing the residual function of the distance between the measurement and the modeled using a Powell method[29].Results are given in the the resistance at high frequency, γ is the fractional order of the constant phase element and Data were fit by minimizing the residual function of the distance between the measurement and the modeled ]. Results are given in the ce of the C2C12 cells at times 23h, 47 , when using 3 different sampling at different sampling frequencies overlaps, as shown in the Figure10.This flexibility allows the addition of more frequency points for a better evaluation of the regions of interest.It should be noted that the sampling frequency can be changed by software, either manually or automaticallyThe results obtained in these experiments show that the frequency band from 30 kHz to 200 kHz is optimal for the ure corresponding to the Data presented in Figure9.a were used in a impedance model identification algorithm.Due to the bandwidth of the measurements (2 kHz to 500 kHz) and the limited double layer capacitance e observable is associated with the cell membrane and the intra ].The electrode impedance is , (4) γ is the fractional order of the constant phase element and f u the Data were fit by minimizing the residual function of the distance between the measurement and the modeled data ].Results are given in the to a pure integrator of capacitor, to values around 0.5, corresponding to a Warburg CPE.As the measurement conclusion on the cor and the b However, the measurement method clearly enables to observe changes in the electrical impedance over time for the targeted application.Moreover, the measurement data can fit standard electrode impedance model conventional optimization algorithms.

Figure 13 .
Figure 13.Magnitude of impedance regions with the heart in perfusion, measured by Fs = 1 MHz.

TABLE 1
Hardware resource utilization of the OMEIS emitter implemented in a Cyclone IV FPGA.
the frequency of 19.53 kHz and 488.28 kHz.The second signal was intentionally shifted 20 dB down for better visibility.
voltage signal (below) form a voltage divider circuit between impedance is shown.The markers (∆V) show the amplitude difference, in dB, between both spectrum at