Won Gu Lee

A novel electroporation method using a capillary and wire-type electrode

Electroporation is widely used to achieve gene transfection. A common problem in electroporation is that it has a lower viability than any other transfection method. In this study, we developed a novel electroporation device using a capillary tip and a pipette that was effective on a wide range of mammalian cells, including cell lines, primary cells, and stem cells. The capillary electroporation system considerably reduced cell death during electroporation because of its wire-type electrode, which has a small surface area. The experimental results also indicated that the cell viability was dependent on the change in pH induced by electrolysis during electroporation. Additionally, the use of a long and narrow capillary tube combined with simple pipetting shortened the overall time of the electroporation process by up to 15 min, even under different conditions with 24 samples. These results were supported by comparison with a conventional electroporation system. The transfection rate and the cell viability were enhanced by the use of the capillary system, which had a high transfection rate of more than 80% in general cell lines such as HeLa and COS-7, and more than 50% in hard-to-transfect cells such as stem or primary cells. The viability was approximately 70-80% in all cell types used in this study.

Active sealing for soft polymer microchips: method and practical applications

This paper presents a new sealing method for soft polymer (elastomer) microchips. A robust and reversible sealing method, which allows various materials to be bonded and sealed tightly with each other even in aqueous solutions, is developed. A poly (dimethylsiloxane) microchip system, which can actively generate bonding and sealing forces by itself, is invented. By inducing negative pressure into additional closed areas, an instant sucking disc is made. This disc is used to tighten up the conformal contact of soft polymers. Other functionalities of active sealing such as making reusable microchips, patterning cells and performing cellular assays in a single dish have also been examined and will be discussed hereunder. This technique gives a robust and universal solution for microchip sealing issues by sealing soft polymers with diverse materials under various conditions. Active sealing will simplify numerous assays in lab-on-a-chip industry and will open a new era for cellular microchip assays.

Combined microchannel-type erythrocyte deformability test with optical tweezers

A combined microchannel-type erythrocyte deformability test with optical tweezers has been developed especially for more sensitive detection of cancerous diseases. To demonstrate the performance and sensitivity of the microchannel-type method, we measured the transit velocity of individual erythrocytes passing through a specific confinement region and calculated the modified elongation index defined by the ratio of the width of the microchannel to the elongated length of the squeezed erythrocytes. To know exactly the effect of optical tweezers on erythrocytes, we investigated several morphologies of optically deformed erythrocytes and measured the shape recovery time of erythrocytes in a static aqueous solution under various powers (~ 24 mW) of 1064-nm laser by a dual-trap optical tweezers. Finally we combined these two methods by considering the key parameters of erythrocyte deformability. The results show that the ambiguity of the overlapped experimental data from microchannel-type erythrocyte deformability test was conspicuously reduced, and that the subtle change (≈ 100-200 ms) in shape recovery time which is one of mechanical properties of erythrocyte membrane surface was remarkably amplified to readily discriminate the difference (≈ 2-3 s) between normal and cancerous blood. This suggests the combined method is more sensitive enough to pinpoint the minor quantitative differences between individual erythrocytes, especially in the field of cancer and cardiovascular diseases.

A glasses-type wearable device for monitoring the patterns of food intake and facial activity

Here we present a new method for automatic and objective monitoring of ingestive behaviors in comparison with other facial activities through load cells embedded in a pair of glasses, named GlasSense. Typically, activated by subtle contraction and relaxation of a temporalis muscle, there is a cyclic movement of the temporomandibular joint during mastication. However, such muscular signals are, in general, too weak to sense without amplification or an electromyographic analysis. To detect these oscillatory facial signals without any use of obtrusive device, we incorporated a load cell into each hinge which was used as a lever mechanism on both sides of the glasses. Thus, the signal measured at the load cells can detect the force amplified mechanically by the hinge. We demonstrated a proof-of-concept validation of the amplification by differentiating the force signals between the hinge and the temple. A pattern recognition was applied to extract statistical features and classify featured behavioral patterns, such as natural head movement, chewing, talking, and wink. The overall results showed that the average F1 score of the classification was about 94.0% and the accuracy above 89%. We believe this approach will be helpful for designing a non-intrusive and un-obtrusive eyewear-based ingestive behavior monitoring system.

Single red blood cell deformability test using optical trapping in plastic microfluidic chip

In this study, single red blood cell (RBC) deformability test is performed by using optical trapping in poly(dimethylsiloxane) PDMS microfluidic chip. The salient feature of this method is to provide more extensional recovery time of individual RBCs in order to tell the minor difference of RBC deformability between normal and abnormal blood.

Design and Evaluation of Smart Glasses for Food Intake and Physical Activity Classification

This study presents a series of protocols of designing and manufacturing a glasses-type wearable device that detects the patterns of temporalis muscle activities during food intake and other physical activities. We fabricated a 3D-printed frame of the glasses and a load cell-integrated printed circuit board (PCB) module inserted in both hinges of the frame. The module was used to acquire the force signals, and transmit them wirelessly. These procedures provide the system with higher mobility, which can be evaluated in practical wearing conditions such as walking and waggling. A performance of the classification is also evaluated by distinguishing the patterns of food intake from those physical activities. A series of algorithms were used to preprocess the signals, generate feature vectors, and recognize the patterns of several featured activities (chewing and winking), and other physical activities (sedentary rest, talking, and walking). The results showed that the average F1 score of the classification among the featured activities was 91.4%. We believe this approach can be potentially useful for automatic and objective monitoring of ingestive behaviors with higher accuracy as practical means to treat ingestive problems.

Multiplexed Cell-Counting Methods by Using Functional Nanoparticles and Quantum Dots

This chapter mainly deals with investigation and development of intensity-based cellcounting methods using fluorescent silica nanoparticles (SiNPs) and quantum dots (QDs) for differential counting of leukocytes. The proposed cell-counting methods enable us to simultaneously measure multiple subsets of human blood cells using a single detector without fluorescence compensation due to an inherent signal overlap of emission spectra from multiple fluorescent labels. At the beginning of the chapter, brief history and theoretical background of multicolor flow cytometry and previous intensity-based cellcounting methods are reviewed. Subsequently, motivation and objectives of the proposed methods are introduced with current issues in this field.