Ebru Özgür

Dielectrophoresis: Applications and future outlook in point of care

Dielectrophoresis (DEP) is a label free, noninvasive, stand alone, rapid, and sensitive particle manipulation and characterization technique. Improvements in micro‐electro‐mechanical systems technology have enabled the biomedical applications of DEP over the past decades. By this way, integration of DEP into lab‐on‐a‐chip systems has become achievable, creating a potential tool for point‐of‐care (POC) systems. DEP can be utilized in many different POC applications including early detection and prognosis of various cancer types, diagnosis of infectious diseases, blood cell analysis, and stem cell therapy. However, there are still some challenges to be resolved to have DEP‐based devices available in POC market. Today, researchers have focused on these challenges to have this powerful theory as a solution for many POC applications. Here, DEP theory, cell modeling, and most common device structures are introduced briefly. Next, POC applications of DEP theory, such as cell (blood, cancer, stem, and fetal) and microorganism separation, manipulation, and enrichment for diagnosis and prognosis, are explained. Integration of DEP with other detection techniques to have more sensitive systems is summarized. Finally, future outlook for DEP‐based systems are discussed with some challenges, which are currently preventing these systems to be a common tool for POC applications, and possible solutions.

Antioxidant Responses of Lentil to Cold and Drought Stress

The effects of cold and drought stress on antioxidant responses and growth parameters in shoots and roots of lentil (Lens culinaris M cv Sultan 1) seedlings were investigated. Ten-day-old hydroponically grown seedlings were subjected to drought and cold (4°C) stress for 5 days. The length and fresh weight of shoots decreased significantly under both stress conditions, contrary to the increase in these growth parameters for roots under the same conditions. The oxidative damage as generation of malondialdehyde and hydrogen peroxide, was markedly higher in shoots under cold. Both stress conditions caused a significant increase in malondialdehyde levels in root tissues. The increase in proline levels was more pronounced under cold stress in shoots and roots. The tested stress conditions had no significant effect on chlorophyll contents. Superoxide dismutase activity was differentially altered in shoot and root tissues under drought and cold stress. The catalase activity was higher in roots under drought stress. On the other hand, ascorbate peroxidase activity increased in root tissues under cold stress. The results indicate that improved tolerance to cold and drought stress in root and shoot tissues of lentil might be correlated to the increased capacity of antioxidative defense system.

Assessment of effects of multi drug resistance on dielectric properties of K562 leukemic cells using electrorotation

In this study, dielectric characterization of multidrug resistant (MDR) K562 human leukemia cells was carried out using a MEMS based electrorotation (ER) device with 3D electrodes. P-glycoprotein (P-gp) dependent MDR causes variation in cell dielectric properties (cell interior conductivity (σi), membrane capacitance (Cm) and total effective membrane conductance (G*m)) due to overexpression of P-gp, which modulates the activity of membrane-bound Cl− channels. Different cell populations resistant to varying levels of doxorubicin (DOX, 0.1–0.5 μM) and imatinib (IMA, 0.2–0.5 μM) were studied to reveal the relationship between cell dielectric properties and the degree of drug resistance. ER characterization results proved considerable changes in cell membrane and interior dielectric properties as the resistance level to chemotherapeutic drugs changes. The membrane dielectric properties of the cells increase significantly at low (0.1–0.2 μM) drug resistance levels (K562/IMA-0.2: Cm = 15.63 ± 3.02 mF m−2 and G*m = 2953 ± 82 S m−2, and K562/DOX-0.1: Cm = 12.29 ± 2.15 mF m−2 and G*m = 1810 ± 14 S m−2), compared to the sensitive ones (Cm = 8.93 ± 1.43 mF m−2 and G*m = 336 ± 73 S m−2). However, they follow a decreasing trend as the drug resistance level increases (0.3–0.5 μM). The membrane capacitance and effective conductance for IMA resistant K562 cells falls to 8.10 ± 1.69 mF m−2 and 113 ± 18 S m−2 in 0.5 μM resistant cells, respectively. Similarly, the membrane capacitance and effective conductance of DOX resistant cells falls to 8.70 ± 1.71 mF m−2 and 1377 ± 22 S m−2 in 0.5 μM resistant cells, respectively. However, no direct relationship could be observed between increased drug resistance and cell interior conductivity, which showed an oscillating behavior. Results prove that the degree of drug resistance significantly affects the dielectric properties of K562 cells, although they possess a similar size and morphology. Variations in cell dielectric properties result in differences in DEP crossover frequencies, which could be utilized in the detection and separation of MDR using dielectrophoretic based devices.

Applications of Photofermentative Hydrogen Production

Scientific and market strategy is essential in developing biological hydrogen production processes. Plans for future research should be based on current knowledge, experience and techniques. This chapter focuses on the applied issues of photofermentative H2 production using purple non sulfur bacteria (PNSB) in combined systems, and in particular, the optimization of the process on real feedstock such as olive mill wastewater and dark fermenter effluents (DFEs) of thick juice, molasses, and potato steam peels. Based on the current state of the knowledge in the field, the future applicability and prospects of these systems are evaluated. Strategies to overcome the problems are outlined.

Identification and characterization of hydrolytic enzymes from the midgut of Sunn Pest of wheat (Eurygaster integriceps)

To help in the development of Sunn Pest-resistant transgenic plants employing protease or α-amylase inhibitors, midgut hydrolytic enzymes of Sunn Pest (Eurygaster integriceps, Put.) (Heteroptera: Scutelleridae) were identified and characterized biochemically. We observed levels of very low proteolytic activity of trypsin (3 nmoles/min/mg), elastase (0.66 nmoles/min/mg) and leucine aminopeptidase-like (14.4 nmoles/min/mg) proteases, but no chymotrypsin and papain-like activity. Proteolytic activities were insensitive to inhibition by soybean trypsin inhibitor (SBTI) and aprotinin, but were inhibited to varying degrees (40–100%) by the synthetic protease inhibitors PMSF, TPCK, CdCl2 and CuCl2. Compared to proteolytic activity, significantly higher amylolytic activity (4.45 mmoles/min/mg) was observed. Αlpha-amylase activity was found to be resistant to inhibition by bean and chickpea α-amylase inhibitors, and only slightly inhibited by wheat (36.9 ± 1.8%) and maize (40.2 ± 2.8) α-amylase inhibitors. Here, we report on the biochemical properties of digestive enzymes from Sunn Pest midgut, and their inhibition patterns by several synthetic and natural hydrolytic enzyme inhibitors.

Label-free detection of multidrug resistance in K562 cells through isolated 3D-electrode dielectrophoresis.

Dielectrophoresis (DEP), a technique used to separate particles based on different sizes and/or dielectric properties under nonuniform electric field, is a promising method to be applied in label‐free, rapid, and effective cell manipulation and separation. In this study, a microelectromechanical systems‐based, isolated 3D‐electrode DEP device has been designed and implemented for the label‐free detection of multidrug resistance in K562 leukemia cells, based on the differences in their cytoplasmic conductivities. Cells were hydrodynamically focused to the 3D‐electrode arrays, placed on the side walls of the microchannel, through V‐shaped parylene‐C obstacles. 3D‐electrodes extruded along the z‐direction provide uniformly distributed DEP force through channel depth. Cell suspension containing resistant and sensitive cancer cells with 1:100 ratio was continuously flown through the channel at a rate of 10 μL/min. Detection was realized at 48.64 MHz, the cross‐over frequency of sensitive K562 cells, at which sensitive cells flow with the fluid, while the resistant ones are trapped by positive DEP force. Device can be operated at considerably low voltages (<9 Vpp). This is achieved by means of a very thin (0.5 μm) parylene coating on electrodes, providing the advantages offered by the isolation of electrodes from the sample, while the working voltage can still be kept low. Results prove that the presented DEP device can provide an efficient platform for the detection of multidrug resistance in leukemia, in a label‐free manner.

Characterization of the distribution of rotational torque on electrorotation chips with 3D electrodes

This is a study of in‐plane and out‐of‐plane distribution of rotational torque (ROT‐T) and effective electric field (EEF) on electrorotation (ER) devices with 3D electrodes using finite element modeling (FEM) and experimental method. The objective of this study is to investigate electrical characteristics of the ER devices with five different electrode geometries and obtain an optimum structure for ER experiments. Further, it provides a comparison between characteristics of the 3D electrodes and traditionally used 2D electrodes. 3D distributions of EEF were studied by the time‐variant FEM. FEM results were verified experimentally by studying the rotation of biological cells. The results show that the variations of ROT‐T and EEF over the measurement area of the devices are considerably large. This can potentially lead to misinterpretation of recorded data. Therefore, it is essential to specify the boundaries of the measurement area with minimum deviation from the central EEF. For this purpose, FE analyses were utilized to specify the optimal region. Thereby, with confining the measurements to these regions, the dependency of ROT‐T on the spatial position of the particles can be eliminated. Comparisons have been made on the sustainability of the EEF and ROT‐T distributions for each device, to find an optimum design. Analyses of the devices prove that utilization of the 3D electrodes eliminate irregularities of EEF and ROT‐T along the z‐axis. The Results show that triangular electrodes provide the highest sustainability for the in‐plane ROT‐T and EEF distribution, while the oblate elliptical and circular electrodes have the lowest variances along the z‐axis.

Label-free detection of leukemia cells with a lab-on-a-chip system integrating dielectrophoresis and cmos imaging

This paper presents a fully-integrated lab-on-a-chip (LOC) system for label-free detection and real-time counting of dielectrophoretically trapped multidrug resistant (MDR) K562 cells. The system integrates a parylene-based microfluidic DEP channel on top of a CMOS image sensor for the first time in the literature. The DEP channel can trap MDR K562 cells with 9 Vpp and 10 μl/min flow rate, and the CMOS image sensor can detect the trapped cells as small as 3 μm in diameter with a noise level of 28.3 e-rms.

Performance Enhancement Of Mems-Based Microbial Fuel Cells (μMFC) For Microscale Power Generation

This paper reports the design, fabrication, and testing of a microliter scale Microbial Fuel Cell (μMFC) based on silicon MEMS fabrication technology. μMFC systems are operated under different loads or open circuit to compare the effect of different acclimatization conditions on start-up time. Shewanella oneidensis MR-1 is preferred to be the biocatalyst. The internal resistance is calculated as 20 kΩ under these conditions. Acclimatization of μMFC under a finite load resulted in shorter start-up time (30 hours) when compared to the open load case. Power and current densities normalized to anode area are 2 μW/cm2 and 12 μA/cm2 respectively. When the load resistance value is closer to the internal resistance of the μMFC, higher power and current densities are achieved as expected, and it resulted in a shorter start-up time. Further studies focusing on the different acclimatization techniques for μMFC could pave the way to use μMFCs as fast and efficient portable power sources.

Transcriptional Profiling of Hydrogen Production Metabolism of Rhodobacter capsulatus under Temperature Stress by Microarray Analysis

Biohydrogen is a clean and renewable form of hydrogen, which can be produced by photosynthetic bacteria in outdoor large-scale photobioreactors using sunlight. In this study, the transcriptional response of Rhodobacter capsulatus to cold (4 °C) and heat (42 °C) stress was studied using microarrays. Bacteria were grown in 30/2 acetate/glutamate medium at 30 °C for 48 h under continuous illumination. Then, cold and heat stresses were applied for two and six hours. Growth and hydrogen production were impaired under both stress conditions. Microarray chips for R. capsulatus were custom designed by Affymetrix (GeneChip®. TR_RCH2a520699F). The numbers of significantly changed genes were 328 and 293 out of 3685 genes under cold and heat stress, respectively. Our results indicate that temperature stress greatly affects the hydrogen production metabolisms of R. capsulatus. Specifically, the expression of genes that participate in nitrogen metabolism, photosynthesis and the electron transport system were induced by cold stress, while decreased by heat stress. Heat stress also resulted in down regulation of genes related to cell envelope, transporter and binding proteins. Transcriptome analysis and physiological results were consistent with each other. The results presented here may aid clarification of the genetic mechanisms for hydrogen production in purple non-sulfur (PNS) bacteria under temperature stress.