Monsur Islam

Enrichment of diluted cell populations from large sample volumes using 3D carbon-electrode dielectrophoresis

Here, we report on an enrichment protocol using carbon electrode dielectrophoresis to isolate and purify a targeted cell population from sample volumes up to 4 ml. We aim at trapping, washing, and recovering an enriched cell fraction that will facilitate downstream analysis. We used an increasingly diluted sample of yeast, 10(6)-10(2) cells/ml, to demonstrate the isolation and enrichment of few cells at increasing flow rates. A maximum average enrichment of 154.2 ± 23.7 times was achieved when the sample flow rate was 10 μl/min and yeast cells were suspended in low electrically conductive media that maximizes dielectrophoresis trapping. A COMSOL Multiphysics model allowed for the comparison between experimental and simulation results. Discussion is conducted on the discrepancies between such results and how the model can be further improved.

Challenges in the Use of Compact Disc-Based Centrifugal Microfluidics for Healthcare Diagnostics at the Extreme Point of Care

Since its inception, Compact Disc (CD)-based centrifugal microfluidic technology has drawn a great deal of interest within research communities due to its potential use in biomedical applications. The technology has been referred to by different names, including compact-disc microfluidics, lab-on-a-disk, lab-on-a-CD and bio-disk. This paper critically reviews the state-of-the-art in CD-based centrifugal microfluidics devices and attempts to identify the challenges that, if solved, would enable their use in the extreme point of care. Sample actuation, manufacturing, reagent storage and implementation, target multiplexing, bio-particle detection, required hardware and system disposal, and sustainability are the topics of focus.

Dielectrophoretic Separation of Live and Dead Monocytes Using 3D Carbon-Electrodes

Blood has been the most reliable body fluid commonly used for the diagnosis of diseases. Although there have been promising investigations for the development of novel lab-on-a-chip devices to utilize other body fluids such as urine and sweat samples in diagnosis, their stability remains a problem that limits the reliability and accuracy of readouts. Hence, accurate and quantitative separation and characterization of blood cells are still crucial. The first step in achieving high-resolution characteristics for specific cell subpopulations from the whole blood is the isolation of pure cell populations from a mixture of cell suspensions. Second, live cells need to be purified from dead cells; otherwise, dead cells might introduce biases in the measurements. In addition, the separation and characterization methods being used must preserve the genetic and phenotypic properties of the cells. Among the characterization and separation approaches, dielectrophoresis (DEP) is one of the oldest and most efficient label-free quantification methods, which directly purifies and characterizes cells using their intrinsic, physical properties. In this study, we present the dielectrophoretic separation and characterization of live and dead monocytes using 3D carbon-electrodes. Our approach successfully removed the dead monocytes while preserving the viability of the live monocytes. Therefore, when blood analyses and disease diagnosis are performed with enriched, live monocyte populations, this approach will reduce the dead-cell contamination risk and achieve more reliable and accurate test results.

Dielectrophoretic characterization and separation of monocytes and macrophages using 3D carbon-electrodes

Monocyte heterogeneity and its prevalence are revealed as indicator of several human diseases ranking from cardiovascular diseases to rheumatoid arthritis, chronic kidney diseases, autoimmune multiple sclerosis, and stroke injuries. When monocytes and macrophages are characterized and isolated with preserved genetic, phenotypic and functional properties, they can be used as label‐free biomarkers for precise diagnostics and treatment of various diseases. Here, the dielectrophoretic responses of the monocytes and macrophages were examined. We present 3D carbon‐electrode dielectrophoresis (carbon‐DEP) as a separation tool for U937 monocytes and U937 monocyte‐differentiated macrophages. The carbon‐electrodes advanced the usability and throughput of DEP separation, presented wider electrochemical stability. Using the 3D carbon‐DEP chip, we first identified the selective positive and negative DEP responses and specific crossover frequencies of monocytes and macrophages as their signatures for separation. The crossover frequency of monocytes and macrophages was 17 and 30 kHz, respectively. Next, we separated monocyte and macrophage subpopulations using their specific dielectrophoretic responses. Afterward, we used a fluorescence‐activated cell sorter to confirm our results. Finally, we enriched 70% of monocyte cells from the mixed cell population, in other words, concentration of monocyte cells to macrophage cells was five times increased, using the 30‐kHz, 10‐Vpp electric field and 1 μL/min flow rate.

Quantitative Investigation for the Dielectrophoretic Effect of Fluorescent Dyes at Single-Cell Resolution

Most of the microscopy-based, quantitative assays rely on fluorescent dyes. In this study, we investigated the impact of fluorescent dyes on the dielectrophoretic response of the mammalian cells. The dielectrophoretic measurements were performed to quantify whether the fluorescent dyes alter the dielectrophoretic properties of the cells at single-cell resolution. Our results present that when 10 Vpp electric field is applied, the fluorescent-labeled cells experienced the crossover frequency at 8–10 kHz, whereas the label-free cells exhibited at 16–18 kHz.

Assessing the importance of the root mean square (RMS) value of different waveforms to determine the strength of a dielectrophoresis trapping force

Different fabrication technologies are now available to implement the electric field gradient required to induce a dielectrophoretic (DEP) force across a sample of interest [1]. However, the optimization of the polarizing waveform is still an understudied topic. Here we present a methodical comparison between the use of sinusoidal, square and triangular signals to polarize a DEP array for particle trapping. Limited work has been done in this area, and always in function of the application being developed [2–4]. It is known that the strength of the DEP force is proportional to the root mean square (RMS) value of the polarizing signal [5]. The RMS amplitude of sinusoidal, square and triangular signals is A √ 2 , A, and A √ 3 respectively, where A is the peak amplitude of the signal. Some authors reported that time-based differences, i.e. shape, amplitude, or frequency, in AC (alternating current) signals contribute to changes in DEP behavior [2–4]. We postulate that these changes are the result of the time-averaged RMS voltage, which normalizes the effect of time on changing AC signals. We show that the trapping of particles using positive DEP (pDEP) is approximately the same regardless of the shape of the polarizing signal; as long as the waveforms feature an equivalent RMS voltage magnitude. We characterized the trapping characteristics of 1.1 ± 0.12 m-diameter polystyrene particles (yellow-green fluorescent, Magsphere Inc.) on an array of 3D carbon post electrodes (100 m height by 50 m diameter). A microchannel made with double-sided pressure-sensitive adhesive contained the electrode array. The reader is referred to our previous publications to consult the fabrication details of these flow-through carbon-electrode DEP devices [6–9]. Particles were suspended in distilled water at a concentration of

3D Carbon-Electrode Dielectrophoresis for Enrichment of a Small Cell Population from a Large Sample Volume

Isolation and enrichment of cells from a diluted sample is necessary for different clinical applications. Here we have demonstrated the use of 3D carbon electrode dielectrophoresis (DEP) to process a diluted yeast sample featuring concentration as low as 102 cells/ml. The yeast cells in the sample were first trapped on carbon electrodes by implementing positive DEP force and then released concentrated in a small volume of clean buffer. The maximum limit of the cell trapping for our device was found to be around 4000 cells. Using 10 μl/min, an enrichment of 154.2 ± 23.7 folds was achieved, where sample of 102 cells/ml concentration was enriched up to 4 X 104 cells/ml. Upon increasing the flow rate up to 30 μl/min, the enrichment dropped down to 18.4 ± 4 folds due to the increase of drag force, though the enriched concentration around 104 cells/ml was still achieved.

Additive Manufacturing of Carbides Using Renewable Resources

Here we report initial results towards additive manufacturing of carbides. We shaped and heat treated biopolymers-metal oxide gel composites in order to obtain 3-D carbide structures. Renewable biopolymers such as iota-carrageenan, chitin and cellulose were used to form the gels. Heat treatment of the gel composites resulted in amorphous porous carbonaceous material with high surface area. The carbonaceous materials preserved the original 3D shape. The ongoing work is on optimization of the conditions for carbide synthesis. We are also studying the rheology of the gel composites to aid to the additive manufacturing.Copyright

Carbon cone electrodes for Selection, Manipulation and Lysis of Single cells

Here we present initial experiments towards an integrated platform for single cell selection, manipulation and lysis. The premise is that an array of polarized conical carbon electrodes can be dipped in a cell culture, trap cells of interest using dielectrophoresis and transport them to specific locations where they can be lyzed electrically. We aim at developing an automated tool to extract intracellular components from targeted particles over specific locations, i.e., a DNA microarray or other functionalized spots. What we contribute in this work is modeling of the electric field and its gradient around carbon cones, as well as initial cone fabrication results. To the best of our knowledge, both the fabrication of conical glassy carbon electrodes and the general concept of the proposed platform are novel. Ongoing work is on demonstrating cell trapping and lysis using these conical electrodes by only varying the magnitude and frequency of their polarizing AC signal.Copyright