Modeling of on-chip (bio)-particle separation and counting using 3D electrode structures

This paper was presented at the 4th Micro and Nano Flows Conference (MNF2014), which was held at University College, London, UK. The conference was organised by Brunel University and supported by the Italian Union of Thermofluiddynamics, IPEM, the Process Intensification Network, the Institution of Mechanical Engineers, the Heat Transfer Society, HEXAG - the Heat Exchange Action Group, and the Energy Institute, ASME Press, LCN London Centre for Nanotechnology, UCL University College London, UCL Engineering, the International NanoScience Community, www.nanopaprika.eu.In lab-on-a-chip applications, manipulation and quantification of (bio)particles is required in a variety of biomedical applications such as drug screening, disease detection and treatment. For manipulation of particles, electrical techniques such as dielectrophoresis (DEP) is very suitable. For the quantification or counting of the bioparticles, flow cytometer, fluorescence-activated cell sorting (FACS) and magneticactivated cell sorting (MACS) are common techniques. In this study, modeling of microfluidic (bio)-particle separation based on dielectrophoresis and counting based on capacitance measurement using COMSOL Multiphysics have been presented. The device performance with planar and 3D electrode structures have been compared. Microfluidic devices with an asymmetric pair (for separation) and a symmetric pair (for counting) of electrodes are considered. The effects of the geometrical parameters, material properties, flow rate, particle size and applied voltage on the device performance have been discussed. The fabrication procedure of 3D electrode structures is also addressed

A Microfluidic Programmable Array for Label-free Detection of Biomolecules

One of the most promising ways to improve clinical diagnostic tools is to use microfluidic Lab-on-a-chip devices. Such devices can provide a dense array of fluidic components and sensors at the micro-scale which drastically reduce the necessary sample volumes and testing time. This dissertation develops a unique electrochemical sensor array in a microfluidic device for high-throughput, label-free detection of both DNA hybridization and protein adsorption experiments. The device consists of a patterned 3 x 3 grid of electrodes which can be individually addressed and microfluidic channels molded using the elastomer PDMS. The channels are bonded over the patterned electrodes on a silicon or glass substrate. The electrodes are designed to provide a row-column addressing format to reduce the number of contact pads required and to drastically reduce the complexity involved in scaling the device to include larger arrays. The device includes straight channels of 100 micron height which can be manually rotated to provide either horizontal or vertical fluid flow over the patterned sensors. To enhance the design of the arrayed device, a series of microvalves were integrated with the platform. This integrated system requires rounded microfluidic channels of 32 micron height and a second layer of channels which act as pneumatic valves to pinch off selected areas of the microfluidic channel. With the valves, the fluid flow direction can be controlled autonomously without moving the bonded PDMS layer. Changes to the mechanism of detection and diffusion properties of the system were examined after the integration of the microvalve network. Protein adhesion studies of three different proteins to three functionalized surfaces were performed. The electrochemical characterization data could be used to help identify adhesion properties for surface coatings used in biomedical devices or for passivating sensor surfaces. DNA hybridization experiments were performed and confirmed both arrayed and sensitive detection. Hybridization experiments performed in the valved device demonstrated an altered diffusion regime which directly affected the detection mechanism. On average, successful hybridization yielded a signal increase 8x higher than two separate control experiments. The detection limit of the sensor was calculated to be 8 nM

A Simple Micromilled Microfluidic Impedance Cytometer with Vertical Parallel Electrodes for Cell Viability Analysis

Microfluidic impedance cytometry has been demonstrated as an effective platform for single cell analysis, taking advantage of microfabricated features and dielectric cell sensing methods. In this study, we present a simple microfluidic device to improve the sensitivity, accuracy, and throughput of single suspension cell viability analysis using vertical sidewall electrodes fabricated by a widely accessible negative manufacturing method. A microchannel milled through a 75 µm platinum wire, which was embedded into poly-methyl-methacrylate (PMMA), created a pair of parallel vertical sidewall platinum electrodes. Jurkat cells were interrogated in a custom low-conductivity buffer (1.2 ± 0.04 mS/cm) to reduce current leakage and increase device sensitivity. Confirmed by live/dead staining and electron microscopy, a single optimum excitation frequency of 2 MHz was identified at which live and dead cells were discriminated based on the disruption in the cell membrane associated with cell death. At this frequency, live cells were found to exhibit changes in the impedance phase with no appreciable change in magnitude, while dead cells displayed the opposite behavior. Correlated with video microscopy, a computational algorithm was created that could identify cell detection events and determine cell viability status by application of a mathematical correlation method

Single-cell microfluidic impedance cytometry: From raw signals to cell phenotypes using data analytics

The biophysical analysis of single-cells by microfluidic impedance cytometry is emerging as a label-free and high-throughput means to stratify the heterogeneity of cellular systems based on their electrophysiology. Emerging applications range from fundamental life-science and drug assessment research to point-of-care diagnostics and precision medicine. Recently, novel chip designs and data analytic strategies are laying the foundation for multiparametric cell characterization and subpopulation distinction, which are essential to understand biological function, follow disease progression and monitor cell behaviour in microsystems. In this tutorial review, we present a comparative survey of the approaches to elucidate cellular and subcellular features from impedance cytometry data, covering the related subjects of device design, data analytics (i.e., signal processing, dielectric modelling, population clustering), and phenotyping applications. We give special emphasis to the exciting recent developments of the technique (timeframe 2017-2020) and provide our perspective on future challenges and directions. Its synergistic application with microfluidic separation, sensor science and machine learning can form an essential tool-kit for label-free quantification and isolation of subpopulations to stratify heterogeneous biosystems

Enhanced Microfluidic Electromagnetic Measurements

Techniques for enhanced microfluidic impedance spectroscopy include causing a core fluid to flow into a channel between two sheath flows of one or more sheath fluids different from the core fluid. Flow in the channel is laminar. A dielectric constant of a fluid constituting either sheath flow is much less than a dielectric constant of the core fluid. Electrical impedance is measured in the channel between at least a first pair of electrodes. In some embodiments, enhanced optical measurements include causing a core fluid to flow into a channel between two sheath flows of one or more sheath fluids different from the core fluid. An optical index of refraction of a fluid constituting either sheath flow is much less than an optical index of refraction of the core fluid. An optical property is measured in the channel

DEVELOPMENT OF HIGH-THROUGHPUT IMPEDANCE SPECTROSCOPY-BASED MICROFLUIDIC PLATFORM FOR DETECTING AND ANALYZING CELLS AND PARTICLES

Impedance spectroscopy based microfluidics have the capability to characterize the dielectric properties of mediums, particles, cellular and sub-cellular contents in response to stimulating voltage signals over a frequency range. This label-free technology has broad ranges of applications in life sciences where there is a need for high-throughput, label-free, non-contact, and low-cost microsystems. To address these limitations, three innovative impedance spectroscopy microfluidic platforms have been developed and presented in this dissertation. The first platform was developed for detecting and characterizing the transverse position of a single cell flowing within a microfluidic channel using a single impedance spectroscopy electrode pair. Regardless of the cell separation methods used, identifying and quantifying the position of cells and particles within a microchannel are important, as these information indicate both the degree of separation as well as how many cells are separated into each position. Using a single pair of non-parallel surface microelectrodes, five different transverse positions of single cells flowing through a microfluidic channel were successfully identified at a throughput of more than 400 particles/s using the detected impedance peak height and width. The second platform utilizes the above technology to count and quantify cells flowing through multiple outlets of microfluidic cell separation systems. A single pair of step-shaped electrodes was developed by integrating five different electrode-to-electrode gaps within a single pair of electrodes. Using this platform, an overall misclassification error rate of only 1.85% was achieved. The result shows the technology’s capability in achieving efficient on-chip cell counting and quantification, regardless of the cell separation methods used, making it a promising on-chip, low-cost and label-free quantification method for cell and particle sorting and separation applications. The third platform was developed for counting cells and particles encapsulated in water-in-oil emulsion droplets using microfluidic based impedance spectroscopy systems. Impedance signal peak height and width were utilized to successfully quantify the number of cells encapsulated within a droplet, and was successfully applied for various cell types and growth media. In addition, the developed platform has been also successfully tested for identifying and discriminating filamentous fungal cell growth, where single fungal spores and filamentous fungi of different lengths could be discriminated inside droplets. Overall in this research, several impedance spectroscopy based microfluidic systems have been successfully developed to solve current limitations in technologies that need high-throughput, low-cost and label-free detection and characterization method for a broad range of cell/particle screening applications

Integrated systems for rapid point of care (PoC) blood cell analysis

Counting the different subpopulations of cells in a fingerprick of human blood is important for a number of clinical point-of-care (PoC) applications. It is a challenge to demonstrate the integration of sample preparation and detection techniques in a single platform. In this paper we demonstrate a generic microfluidic platform that combines sample processing and characterisation and enumeration in a single, integrated system. Results of microfluidic 3-part differential leukocyte (granulocyte, lymphocyte and monocyte) counts, together with erythrocyte and thrombocyte (platelet) counts, in human blood are shown and corroborated with results from hospital clinical laboratory analysis

Three-Dimensional Electrode Integration with Microwave Sensors for Precise Microparticle Detection in Microfluidics

Microwave sensors integrated with microfluidic platforms can provide the size and permittivity of single cells and microparticles. Amongst the microwave sensor topologies, the planar arrangement of electrodes is a popular choice owing to the ease of fabrication. Unfortunately, planar electrodes generate a non-uniform electric field which causes the responsivity of the sensor to depend on the vertical position of a microparticle in the microfluidic channel. To overcome this problem, we fabricated three-dimensional (3D) electrodes at the coplanar sensing region of an underlying microwave resonator. The 3D electrodes are based on SU8 polymer which is then metallized by sputter coating. With this system, we readily characterized a mixture composed of 12 um and 20 um polystyrene particles and demonstrated separation without any position-related calibration. The ratio of the electronic response of the two particle types is approximately equal to the ratio of the particle volumes, which indicates the generation of a uniform electric field at the sensing region. The current work obviates the need for using multiple coplanar electrodes and extensive processing of the data for the calibration of particle height in a microfluidic channel: as such, it enables the fabrication of more sophisticated microwave resonators for environmental and biological applications

Surface Acoustic Wave-Based Microfluidic Device for Microparticles Manipulation: Effects of Microchannel Elasticity on the Device Performance

: Size sorting, line focusing, and isolation of microparticles or cells are fundamental ingredients in the improvement of disease diagnostic tools adopted in biology and biomedicine. Microfluidic devices are exploited as a solution to transport and manipulate (bio)particles via a liquid flow. Use of acoustic waves traveling through the fluid provides non-contact solutions to the handling goal, by exploiting the acoustophoretic phenomenon. In this paper, a finite element model of a microfluidic surface acoustic wave-based device for the manipulation of microparticles is reported. Counter-propagating waves are designed to interfere inside a PDMS microchannel and generate a standing surface acoustic wave which is transmitted to the fluid as a standing pressure field. A model of the cross-section of the device is considered to perform a sensitivity analysis of such a standing pressure field to uncertainties related to the geometry of the microchannel, especially in terms of thickness and width of the fluid domain. To also assess the effects caused by possible secondary waves traveling in the microchannel, the PDMS is modeled as an elastic solid material. Remarkable effects and possible issues in microparticle actuation, as related to the size of the microchannel, are discussed by way of exemplary results