Biological Cells In Suspension Research Articles

An equivalent electric circuit as a tool for monitoring micron sized-particles in suspension from electrical impedance measurements

In this work, a microfluidic device with a three-electrode system is used to measure the electrical impedance spectrum of micron-sized particles suspended in Phosphate Buffered Saline (PBS). We use polymer particles of 15 μm and 48 μm of diameter, as they can mimic biological cells in suspension. The main purpose is to study the influence of the particle concentration on impedance parameters and to establish the basis for biological cell suspensions assays. An electric equivalent circuit model is used to describe the experimental impedance data. Results show a high dependence of electrical parameters with the mass fraction and particle size. The model considers the resistance of the bulk solution and the electrical double layer, as the two essential electrical elements. Results show a maximum error of 3% by fitting the measurements to the proposed model. This proposal could be of interest for cytotoxicity assays.

Time-Constant-Domain Spectroscopy: An Impedance-Based Method for Sensing Biological Cells in Suspension

Impedance measurement is a common technique to characterize and detect the electrical properties of biological cells. However, to decode the underlying physical processes, it requires complex electrical models alongside prior knowledge of the sample under study. In this work, we introduce an attractive label-free method for sensing biological cells in suspension based on the measurement of electrical impedance and the distribution of relaxation times (DRT) model. The DRT maps impedance data from the frequency-domain to a time-constant-domain spectrum (TCDS) being a useful and robust method for data analysis. We perform impedance measurements in the range from 1 kHz to 1 MHz to obtain the TCDS for sensing mimic samples as well as HeLa cells in suspension. Results show that the TCDS can be seen as an electrical fingerprint for the sample, as it can decode useful information about the composition and structure with high sensitivity and resolution.

Assessing the electro-deformation and electro-poration of biological cells using a three-dimensional finite element model

In this Letter, we explore how cell electro-deformation and electro-poration are connected. We build a time-domain model of layered concentric shells (a model of biological cells) including their dielectric and elastic properties. We simulate delivery of one trapezoidal voltage pulse to either a single spherical cell or an assembly of three neighboring cells in a specific configuration and calculate cell deformation and pore formation. We describe the qualitative features of the electric field, surface charge density, transmembrane voltage, cell elongation, and pore density distribution at specific times i.e., before, during and after the application of the electric pulse and explore the correlations between them. Our results show that (1) the polarization charge redistribution plays a significant role in the spatial distribution of electrical stresses at μs time scales and (2) the cell deformation and pore density can be correlated with regions of high surface charge density. In future work, our model could be used for understanding basic mechanisms of electro-deformation and electro-poration with high-frequency short bipolar pulses of biological cells in suspension or tissues.

Out-of-Plane Rotation Control of Biological Cells With a Robot-Tweezers Manipulation System for Orientation-Based Cell Surgery.

In many cell surgery applications, cell must be oriented properly such that the microsurgery tool can access the target components with minimum damage to the cell. In this paper, a scheme for out of image plane orientation control of suspended biological cells using robotic controlled optical tweezers is presented for orientation-based cell surgery. Based on our previous work on planar cell rotation using optical tweezers, the dynamic model of cell out-of-plane orientation control is formulated by using the T-matrix approach. Vision-based algorithms are developed to extract the cell out of image plane orientation angles, based on 2-D image slices obtained under an optical microscope. A robust feedback controller is then proposed to achieve cell out-of-plane rotation. Experiments of automated out of image plane rotational control for cell nucleus extraction surgery are performed to demonstrate the effectiveness of the proposed approach. This approach advances robot-aided single cell manipulation and produces impactful benefits to cell surgery applications such as nucleus transplantation and organelle biopsy in precision medicine.

Integral refractive index imaging of flowing cell nuclei using quantitative phase microscopy combined with fluorescence microscopy.

We suggest a new multimodal imaging technique for quantitatively measuring the integral (thickness-average) refractive index of the nuclei of live biological cells in suspension. For this aim, we combined quantitative phase microscopy with simultaneous 2-D fluorescence microscopy. We used 2-D fluorescence microscopy to localize the nucleus inside the quantitative phase map of the cell, as well as for measuring the nucleus radii. As verified offline by both 3-D confocal fluorescence microscopy and 2-D fluorescence microscopy while rotating the cells during flow, the nucleus of cells in suspension that are not during division can be assumed to be an ellipsoid. The entire shape of a cell in suspension can be assumed to be a sphere. Then, the cell and nucleus 3-D shapes can be evaluated based on their in-plain radii available from the 2-D phase and fluorescent measurements, respectively. Finally, the nucleus integral refractive index profile is calculated. We demonstrate the new technique on cancer cells, obtaining nucleus refractive index values that are lower than those of the cytoplasm, coinciding with recent findings. We believe that the proposed technique has the potential to be used for flow cytometry, where full 3-D refractive index tomography is too slow to be implemented during flow.

Thermophoretic Tweezers for Low-Power and Versatile Manipulation of Biological Cells.

Optical manipulation of biological cells and nanoparticles is significantly important in life sciences, early disease diagnosis, and nanomanufacturing. However, low-power and versatile all-optical manipulation has remained elusive. Herein, we have achieved light-directed versatile thermophoretic manipulation of biological cells at an optical power 100-1000 times lower than that of optical tweezers. By harnessing the permittivity gradient in the electric double layer of the charged surface of the cell membrane, we succeed at the low-power trapping of suspended biological cells within a light-controlled temperature gradient field. Furthermore, through dynamic control of optothermal potentials using a digital micromirror device, we have achieved arbitrary spatial arrangements of cells at a resolution of ∼100 nm and precise rotation of both single and assemblies of cells. Our thermophoretic tweezers will find applications in cellular biology, nanomedicine, and tissue engineering.

Study on the dielectric properties of biological cells using the electrokinetic method

AbstractDifferent from hard particles with an ordinary surface structure, a biological cell is a soft particle which can be described as a hard particle covered with an ion‐penetrable surface layer of polyelectrolytes. In this paper, according to the electrokinetic theory of suspension of soft particles, the colloid vibration current (CVI) model of suspended biological cells is investigated. A CVI measurement system is set up in which ultrasonic pulses of different intensities are calibrated and used. The experimental results on electrolyte solutions and cell suspensions show that CVI is closely related to the ion and cell concentration of the sample solution. Along with the increased concentration of ions or cells, the CVI signal increased initially and then decreased for electrolyte solutions, but it was the reverse for a chicken blood cell suspension but stayed almost constant for a spawn suspension. All these phenomena were caused by the interactive electrokinetic polarization of ions and cells under the action of ultrasonic pulses. Through the analysis of the electrokinetic course of the cell suspension, it was found that the amplitude and slope of the electrokinetic curve could be used to characterize the dielectric properties of cells. The electrokinetic model and CVI measurement system are rather feasible, as shown by the experimental results. The electrokinetic measurement method shows a new nondestructive way for studying the charging properties of biological cells, and the method has an inherent advantage for the real‐time study of the biological effects of ultrasound on cells. © 2012 Institute of Electrical Engineers of Japan. Published by John Wiley & Sons, Inc.

Hyperpolarized 129Xe NMR signature of living biological cells

We show that the differentiation between internal and external compartments of various biological cells in suspension can be made via simple NMR spectra of hyperpolarized (129) Xe. The spectral separation between the signals of (129) Xe in these two compartments is already known for red blood cells, because of the strong interaction of the noble gas with hemoglobin. The observation of two separate peaks in the 200-ppm region can be seen with both eukaryotic and prokaryotic cells, some of which are not known to contain paramagnetic proteins in large quantities. Using different experiments in which the cells are lysed, swell or are blocked in G2 phase, we demonstrate that the low-field-shifted peak observed corresponds to xenon in the aqueous pool inside the cells and not in the membranes. The presence of this additional peak is a clear indication of cell integrity, and its integration allows the quantification of the total cell volume. The relaxation time of intracellular xenon is sufficiently long to open up promising perspectives for cell characterization. The exchange time between the inner and outer cell compartments (on the order of 30 ms) renders possible the targeting of intracellular receptors, whereas the observation of chemical shift variations represents a method of revealing the presence of toxic species in the cells.

Microfluidic devices for mechanical characterisation of single cells in suspension

This Letter provides an overview of microfluidic technologies for single-cell mechanical characterisation. In particular, the most recent literature is discussed to summarise the working principles and development trend of the state-of-the-art microfluidic devices for mechanical characterisation of biological cells in suspension. The techniques are classified into constriction channel, fluid stress, optical stretcher, electro-deformation, electroporation and microfluidic pipette aspiration, according to the mechanism of mechanical stimuli. The principles are explained along with representative examples demonstrating their applications. The research highlighted in this letter has great potential in realising high-throughput single-cell mechanical characterisation.

Injection molded chips with integrated conducting polymer electrodes for electroporation of cells

We present the design-concept for an all polymer injection molded single use microfluidic device. The fabricated devices comprise integrated conducting polymer electrodes and Luer fitting ports to allow for liquid and electrical access. A case study of low voltage electroporation of biological cells in suspension is presented. The working principle of the electroporation device is based on a focusing of the electric field by means of a constriction in the flow channel for the cells. We demonstrate the use of AC voltage for electroporation by applying a 1 kHz, ±50 V square pulse train to the electrodes and show delivery of polynucleotide fluorescent dye in 46% of human acute monocytic leukemia cells passing the constriction.

Broadband single cell impedance spectroscopy using maximum length sequences: theoretical analysis and practical considerations

Measurements of the dielectric (or impedance) properties of cells can be used as a general characterization and diagnostic tool. In this paper, we describe a novel impedance spectroscopy technique for the analysis of single biological cells in suspension. The technique uses maximum length sequences (MLS) for periodic excitation signal in a microfluidic impedance cytometer. The method allows multi-frequency single cell impedance measurements to be made in a short time period (ms). Spectral information is obtained in the frequency domain by applying a fast M-sequence transform (FMT) and fast Fourier transform (FFT) to the time domain response. Theoretically, the impedance is determined from the transfer function of the system when the MLS is a current excitation. The order of the MLS and sampling rate of A/D conversion are two factors that determine the bandwidth and spectral accuracy of the technique. Experimentally, the applicability of the technique is demonstrated by characterizing the impedance spectrum of red blood cells (RBCs) in a microfluidic cytometer. The impedance is measured within 1 ms at 512 discrete frequencies, evenly distributed in the range from 976.56 Hz to 500 kHz. The measured spectrum shows good agreement with simulations.

Polar nephelometer based on a rotational confocal imaging setup

Rapid measurement of the angular distribution of light scattered by particles, the scattering phase function, is achieved by using a new type of polar nephelometer, a device for measuring the angular scattered-light intensity distribution, with a high angular precision and across many orders of magnitude of intensity. The design offers high-speed measurements and avoids many of the problems often associated with traditional goniometers when they are used for measurements of light scattering from small particles or biological cells in suspension. Our system relies on confocal imaging of the test space with off-axis parabolas, using a rotating mirror to scan the angular field of view at the second focus of a pair of conjugated parabolic mirrors, with the test space located at the first focus. The angular resolution of the system is limited mainly by the data-acquisition sampling frequency. In this proof-of-principle demonstration the system performs multiple scans of a 55 deg field of view in a very short time (<1 s). To significantly increase the signal-to-noise ratio, we averaged the successively acquired scans during this time. Polystyrene spheres dispersed in water at low concentrations were used to test the system. The scattering patterns obtained were found to be in good agreement with Mie theory calculations.

Dielectrophoretic fluidic cell fractionation system

This paper presents the development and experimental verification of a DEP fluidic system capable of fractionation of intact biological cells in suspension into purer subpopulations. This was accomplished by employing a specially shaped nonuniform electric field, synthesized by microfabricated planar microelectrode arrays, housed on an insulating glass substrate. To improve the efficiency of cell sorting, the microelectrodes are individually biased by a variable frequency alternating current (ac) voltage source, which allows us to exploit both positive and negative dielectrophoresis (DEP) to affect cell separation. Furthermore, through suitable establishment of a cell stream supported by sheath flow, such fractionation is achieved in a continuous fashion. The proposed DEP fluidic fractionation may be configured to operate in three (3) different modes. In this work, however, a detailed account is only presented for one mode of operation. The simulation of the electric field and force profiles, together with the experimental results obtained on model cells (plant protoplasts), confirm our theoretical predictions and furthermore demonstrate improvements in both separation efficiency and throughput over a wide range of frequencies (10 Hz to 5 kHz).

Selective detection of viable bacteria using dielectrophoretic impedance measurement method

This paper describes a selective detection technique of viable bacteria based on dielectrophoresis and electrical impedance measurements. The authors have previously proposed a detection technique of biological particles called dielectrophoretic impedance measurement (DEPIM) method using positive dielectrophoretic force to capture biological cells in suspension onto an interdigitated microelectrode array. By combining antigen-antibody reaction with the DEPIM, selective detection of a particular species of bacteria was demonstrated. In this present work, the authors demonstrated another selective DEPIM method utilizing cell viability dependency of dielectrophoretic force without introducing the antigen-antibody reaction. It was found that dielectrophoresis of heat-treated Escherichia coli showed strong dependency on viability when applied field frequency was as high as 1 MHz. As a result, viable bacteria could be exclusively collected by positive dielectrophoresis and selectively detected by the DEPIM technique from a suspension also containing heat-treated nonviable cells. On the other hand, nonviable bacteria obtained by UV irradiation showed little dielectrophoresis dependency on viability. According to a theoretical analysis of the dielectrophoretic force, it is suggested that heat treatment alters the dielectric properties of treated cells. In particular, a decrease in cytoplasmic conductivity, which might be caused by heat-induced perforation of cell membrane, was expected to considerably affect dielectrophoresis characteristics. Proposed selective DEPIM method was also applied to evaluation of heat sterilization effect on a real time basis. It was experimentally proved that DEPIM could evaluate viable cell number variation with heat treatment time in a considerably shorter time than conventional microbiological method based on cell incubation.