Il Doh

Deformation measurement of individual cells in large populations using a single-cell microchamber array chip

We analyze the deformability of individual red blood cells (RBCs) using SiCMA technology. Our approach is adequate to quickly measure large numbers of individual cells in heterogeneous populations. Individual cells are trapped in a large-scale array of micro-wells, and dielectrophoretic (DEP) force is applied to deform the cells. The simple structures of micro-wells and DEP electrodes facilitate the analysis of thousands of RBCs in parallel. This unique method allows the correlation of red cell deformation with cell surface and cytosolic characteristics to define the distribution of individual cellular characteristics in heterogeneous populations.

Passive flow-rate regulators using pressure-dependent autonomous deflection of parallel membrane valves

We present passive flow-rate regulators using an autonomous deflection of parallel membrane valves, capable to maintain a constant flow-rate at varying inlet pressure supplied from micropumps. The previous passive flow-rate regulators are difficult to integrate with micropumps, not only because of the complex multi-layer structures, but also because of the high threshold inlet pressure required for flow-rate regulation. In this study, we present passive flow-rate regulators using parallel membrane valves, capable of achieving flow-rate regulation function at the minimum threshold inlet pressure as low as 15 kPa with simple structure formed by a single mask process. The parallel membranes in a flow-rate regulator are designed to deflect and adjust flow resistance autonomously according to the inlet pressure, thus maintaining a constant flow-rate independent of the inlet pressure variation. We designed the four different prototypes of W20, W30, W40, and W50, having parallel membrane widths of 20, 30, 40 and 50 microm, respectively. We estimated the flow-rate based on both analytical and numerical models. In an experimental study, we observed the deformation of parallel membranes and the flow-rate depending on the inlet pressure. The fabricated prototypes achieved the constant flow-rate of 6.09 +/- 0.32 microl s(-1) (W20 fabricated by 10 : 1 PDMS (PolyDiMethylSiloxane)) over an inlet pressure of 20 kPa. We also observed that prototypes fabricated by 20 : 1 PDMS, having lower Youngs modulus than normal 10 : 1 PDMS, showed a lower threshold pressure and higher regulated flow-rate than prototypes fabricated by 10 : 1 PDMS. W40 fabricated by 20 : 1 PDMS showed a constant flow-rate of 14.53 +/- 0.51 microl s(-1) over inlet pressure of 15 kPa. The present passive flow-rate regulators have strong potential for applications in integrated microfluidic systems.

A Continuous Cell Separation Chip Using Hydrodynamic Dielectrophoresis Process

This paper presents a high-throughput continuous cell separation chip using hydrodynamic dielectrophoresis (DEP) process. We design the continuous cell separation chip with three electrodes, where the cells in positive DEP affinity are separated from the central streamline. In the experimental study, we use the mixture of viable (live) and nonviable (dead) yeast cells as sample cells to be separated. We obtain the continuous cell separation conditions in the DEP affinity test: the sinusoidal electric fields of 5 MHz, 8V/sub p-p/ have been applied across the electrode array of 20 /spl mu/m gaps immersed in the medium conductivity of 5 /spl mu/S/cm. Using switched AC signal under these conditions, we continuously separate the yeast cells at the mixture flow rates of 0.1/spl sim/1 /spl mu/l/min. The purity of the separated viable and nonviable yeast cells has been measured in the range of 95.9/spl sim/97.3% and 64.5/spl sim/74.3%, respectively.

On-chip three-dimensional tumor spheroid formation and pump-less perfusion culture using gravity-driven cell aggregation and balanced droplet dispensing.

This paper presents a spheroid chip in which three-dimensional (3D) tumor spheroids are not only formed by gravity-driven cell aggregation but also cultured at the perfusion rates controlled by balanced droplet dispensing without fluidic pumps. The previous spheroid chips require additional off-chip processes of spheroid formation and extraction as well as bulky components of fluidic pumps. However, the present spheroid chip, where autonomous medium droplet dispensers are integrated on a well array, achieves the on-chip 3D tumor spheroid formation and perfusion culture using simple structure without bulky fluidic pumps. In the experimental study, we demonstrated that the spheroid chip successfully forms 3D tumor spheroids in the wide diameter range of 220 μm-3.2 mm (uniformity > 90%) using H358, H23, and A549 non-small cell lung cancer cells. At the pump-less perfusion culture (Q = 0.1-0.3 μl/min) of spheroids, the number of H358 cells in the spheroid increased up to 50% from the static culture (Q = 0 μl/min) and the viability of the cultured cells also increased about 10%. Therefore, we experimentally verified that the perfusion environment created by the spheroid chip offers a favourable condition to the spheroids with high increase rate and viability. The present chip achieves on-chip 3D tumor spheroid formation and pump-less perfusion culture with simple structure, thereby exhibiting potential for use in integrated in-vivo-like cell culture systems.

Viable capture and release of cancer cells in human whole blood

We present viable cancer cell isolation devices utilizing the physical properties of cells. The tapered slit structure is proposed to isolate cancer cells from blood cells and collect them by reversed flow. From the experimental study using the spiked cancer cells in human whole blood, we verified the capability of the present cancer cell isolation chip in terms of capture efficiency, viability, and release rate. The viable cancer cells obtained from the present chip can be used for the further applications of cancer diagnosis, treatment monitoring, and new target drug development for cancer stem cells.

A continuous cell separation chip using hydrodynamic dielectrophoresis process

This paper presents a high-throughput continuous cell separation chip using hydrodynamic dielectrophoresis (DEP) process. We design the continuous cell separation chip with three electrodes, where the cells in positive DEP affinity are separated from the central streamline. In the experimental study, we use the mixture of viable (live) and nonviable (dead) yeast cells as sample cells to be separated. We obtain the continuous cell separation conditions in the DEP affinity test: the sinusoidal electric fields of 5 MHz, 8V/sub p-p/ have been applied across the electrode array of 20 /spl mu/m gaps immersed in the medium conductivity of 5 /spl mu/S/cm. Using switched AC signal under these conditions, we continuously separate the yeast cells at the mixture flow rates of 0.1/spl sim/1 /spl mu/l/min. The purity of the separated viable and nonviable yeast cells has been measured in the range of 95.9/spl sim/97.3% and 64.5/spl sim/74.3%, respectively.

Advanced combinational microfluidic multiplexer using multiple levels of control pressures

We propose an advanced combinational microfluidic multiplexing method capable of increasing the number of addressable fluidic channels dramatically. Using only 4 control lines, the proposed advanced combinational multiplexer, utilizing two different levels of control pressure, could address up to 19 fluidic channels, which is at least two times larger than previous multiplexers. The difference between the maximum addressable channels in the present and previous methods increases dramatically when the control lines and control pressure levels increase. The present multiplexer, with its high control efficiency and simple structure for channel addressing, could be used in the application areas of integrated microfluidic systems such as high-throughput analyzers and dynamic pressure generators.

Label-free Rapid Viable Enrichment of Circulating Tumor Cell by Photosensitive Polymer-based Microfilter Device

We present a clinical device for simple, rapid, and viable isolation of circulating tumor cells (CTCs) from cancer patient bloods. In spite of the clinical importance of CTCs, the lack of easy and non-biased isolation methods is a big hurdle for implementing CTC into clinical use. The present device made of photosensitive polymer was designed to attach to conventional syringe to isolate the CTCs at minimal resources. Its unique tapered-slits on the filter are capable not only to isolate the cell based on their size and deformability, but also to increase sample flow rate, thus achieving label-free rapid viable CTC isolation. We verified our device performance using 9 different types of cancer cells at the cell concentration from 5 to 100cells/ml, showing that the device capture 77.7% of the CTCs while maintaining their viability of 80.6%. We extended our study using the 18 blood samples from lung, colorectal, pancreatic and renal cancer patients and captured 1-172 CTCs or clustered CTCs by immunofluorescent or immunohistochemical staining. The captured CTCs were also molecularly assayed by RT-PCR with three cancer-associated genes (CK19, EpCAM, and MUC1). Those comprehensive studies proved to use our device for cancer study, thereby inaugurating further in-depth CTC-based clinical researches.

Construction of Static 3D Ultrasonography Image by Radiation Beam Tracking Method from 1D Array Probe

Abstract This paper describes the construction of a static 3D ultrasonography image by tracking the radiation beam position during the handy operation of a 1D array probe to enable point-of-care use. The theoretical model of the transformation from the translational and rotational information of the sensor mounted on the probe to the reference Cartesian coordinate system was given. The signal amplification and serial communication interface module was made using a commercially available sensor. A test phantom was also made using silicone putty in a donut shape. During the movement of the hand-held probe, B-mode movie and sensor signals were recorded. B-mode images were periodically selected from the movie, and the gray levels of the pixels for each image were converted to the gray levels of 3D voxels. 3D and 2D images of arbitrary cross-section of the B-mode type were also constructed from the voxel data, and agreed well with the shape of the test phantom. Keywords: Ultrasound, 3D Image, Position Angle Sensor, Beam Tracking Method[Received: January 9, 2015, Revised: February 3, 2015, Accepted: February 3, 2015] *한국표준과학연구원 의료융합측정표준센터, **과학기술연합대학원대학교 의학물리학과, ***㈜메타바이오메드 HCP사업부, ✝Corresponding Author: Center for Medical Metrology, Korea Research Institute of Standards and Science, Daejeon 305-340, Korea (E-mail: il.doh@kriss.re.kr)

Cell-matrix adhesion characterization using multiple shear stress zones in single stepwise microchannel

This paper presents a cell chip capable to characterize cell-matrix adhesion by monitoring cell detachment rate. The proposed cell chip can supply multiple levels of shear stress in single stepwise microchannel. As epithelial-mesenchymal transition (EMT), one of hallmarks of cancer metastasis is closely associated to the interaction with extracelluar matrix (ECM), we took advantage of two lung cancer cell models with different adhesion properties to ECM depending their epithelial or mesenchymal properties, including the pair of lung cancer cells with (A549sh) or without E-cadherin expression (A549sh-Ecad), which would be optimal model to examine the alteration of adhesion properties after EMT induction. The cell-matrix adhesion resisting to shear stress appeared to be remarkably differed between lung cancer cells. The detachment rate of epithelial-like H358 and mesenchymal-like H460 cells was 53%–80% and 25%–66% in the shear stress range of 34–60 dyn/cm2, respectively. A549sh-Ecad cells exhibits lower det...