• Title/Summary/Keyword: Label-free detection

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Real-Time Monitoring of Mitochondrial ATP Synthesis and Hydrolysis by Surface Infrared Spectroscopy

  • Yamaguchi, Ryo-Taro;Hirano-Iwata, Ayumi;Aonuma, Yuki;Yoshimura, Yuya;Shinohara, Yasuo;Kimura, Yasuo;Niwano, Michio
    • Proceedings of the Korean Vacuum Society Conference
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    • 2013.02a
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    • pp.108-109
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    • 2013
  • Mitochondria play key roles in the production of cell's energy. Their dominant function is the synthesis of adenosine 5'-triphosphate (ATP) from adenosine diphosphate (ADP) and phosphate (Pi) through the oxidative phosphorylation. Evaluation of drug-induced mitochondrial toxicity has become increasingly important since mitochondrial dysfunction has recently been implicated in numerous diseases including cancer and diabetes mellitus. Mitochondrial functions have been monitored via oxygen consumption, mitochondrial membrane potential, and more importantly via ATP synthesis since ATP synthesis is the most essential function of mitochondria. Various analytical methods have been employed to investigate ATP synthesis in mitochondria, including high performance liquid chromatography (HPLC), bioluminescence technique, and pH measurement. However, most of these methods are based on destructive analysis or indirect monitoring through the enzymatic reaction. Infrared absorption spectroscopy (IRAS) is one of the useful techniques for real-time, label-free, and direct monitoring of biological reactions [1,2]. However, the strong water absorption requires very short path length in the order of several micrometers. Transmission measurements with thin path length are not suitable for mitochondrial assays because solution handlings necessary for evaluating mitochondrial toxicity, such as rapid mixing of drugs and oxygen supply, are difficult in such a narrow space. On the other hand, IRAS in the multiple internal reflection (MIR) geometry provides an ideal optical configuration to combine solution handling and aqueous-phase measurement. We have recently reportedon a real-time monitoring of drug-induced necrotic and apoptotic cell death using MIR-IRAS [3,4]. Clear discrimination between viable and damaged cells has been demonstrated, showing a promise as a label-free and real-time detection for cell-based assays. In the present study, we have applied our MIR-IRAS system to mitochondria-based assays by monitoring ATP synthesis in isolated mitochondria from rat livers. Mitochondrial ATP synthesis and hydrolysis were in situ monitored with MIR-IRAS, while dissolved oxygen level and solution pH were simultaneously monitored with O2 and pH electrodes, respectively. It is demonstrated that ATP synthesis and hydrolysis can be monitored by the IR spectral changes in phosphate groups in adenine nucleotides and MIR-IRAS is useful for evaluating time-dependent drug effects of mitochondrial toxicants.

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Enhanced pH Response of Solution-gated Graphene FET by Using Vertically Grown ZnO Nanorods on Graphene Channel

  • Kim, B.Y;Jang, M.;Shin, K.-S.;Sohn, I.Y;Kim, S.-W.;Lee, N.-E
    • Proceedings of the Korean Vacuum Society Conference
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    • 2014.02a
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    • pp.434.2-434.2
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    • 2014
  • We observe enhanced pH response of solution-gated field-effect transistors (SG-FET) having 1D-2D hybrid channel of vertical grown ZnO nanorods grown on CVD graphene (Gr). In recent years, SG-FET based on Gr has received a lot of attention for biochemical sensing applications, because Gr has outstanding properties such as high sensitivity, low detection limit, label-free electrical detection, and so on. However, low-defect CVD Gr has hardly pH responsive due to lack of hydroxyl group on Gr surface. On the other hand, ZnO, consists of stable wurtzite structure, has attracted much interest due to its unique properties and wide range of applications in optoelectronics, biosensors, medical sciences, etc. Especially, ZnO were easily grown as vertical nanorods by hydrothermal method and ZnO nanostructures have higher sensitivity to environments than planar structures due to plentiful hydroxyl group on their surface. We prepared for ZnO nanorods vertically grown on CVD Gr (ZnO nanorods/Gr hybrid channel) and to fabricate SG-FET subsequently. We have analyzed hybrid channel FETs showing transfer characteristics similar to that of pristine Gr FETs and charge neutrality point (CNP) shifts along proton concentration in solution, which can determine pH level of solution. Hybrid channel SG-FET sensors led to increase in pH sensitivity up to 500%, compared to pristine Gr SG-FET sensors. We confirmed plentiful hydroxyl groups on ZnO nanorod surface interact with protons in solution, which causes shifts of CNP. The morphology and electrical characteristics of hybrid channel SG-FET were characterized by FE-SEM and semiconductor parameter analyzer, respectively. Sensitivity and sensing mechanism of ZnO nanorods/Gr hybrid channel FET will be discussed in detail.

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Nano SPR Biosensor for Detecting Lung Cancer-Specific Biomarker (폐암 바이오마커 검출용 나노SPR 바이오센서)

  • Jang, Eun-Yoon;Yeom, Se-Hyuk;Eum, Nyeon-Sik;Han, Jung-Hyun;Kim, Hyung-Kyung;Shin, Yong-Beom;Kang, Shin-Won
    • Journal of Sensor Science and Technology
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    • v.22 no.2
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    • pp.144-149
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    • 2013
  • In this research, we developed a biosensor to detect lung cancer-specific biomarker using Anodic Aluminum Oxide (AAO) chip based on interference and nano surface plasmon resonance (nanoSPR). The nano-porous AAO chip was fabricated $2{\mu}m$ of pore-depth by two-step anodizing method for surface uniformity. NanoSPR has sensitivity to the refractive index (RI) of the surrounding medium and also provides simple and label-free detection when specific antibodies are immobilized to the Au-deposited surface of nano-porous AAO chip. To detect the lung cancer-specific biomarker, antibodies were immobilized on the surface of the chip by Self Assembled Monolayer (SAM) method. Since then lung cancer-specific biomarker was applied atop the antibodies immobilized layer. The specific reaction of the antigen-antibody contributed to the change in the refractive index that cause shift of resonance spectrum in the interference pattern. The Limit of Detection (LOD) was 1 fg/ml by using our nano-porous AAO biosensor chip.

Biosensor System for the Detection of Agrichemicals and Its Applications (농약 검출을 위한 바이오센서 시스템 연구 및 그 응용)

  • Park, Tae-Jung;Yang, Min-Ho;Lee, Sang-Yup;Kim, Soo-Hyun
    • KSBB Journal
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    • v.24 no.3
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    • pp.227-238
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    • 2009
  • In the recent years, some organic toxic chemicals were used for obtaining high-yield productivity in agriculture. The undegraded pesticides may remain in the agricultural foods through atmosphere, water, and soil and cause public health problems to environmental resources and human beings even at very low concentrations. Small amounts of pesticides can affect a central nervous system, resulting in immunogenic diseases, infertility problems, respiratory diseases and born marrow diseases, which can lead even to death. Monitoring of the environmental pesticide is one of the important issues for the human well-being. Several kinds of biosensors have been successfully applied to the detection of agrichemical toxicity. Also, few platforms for biocide detection have been definitely developed for the degradation and reaction of pesticides. Biochip and electrochemistry experiments involve immobilizing a receptor molecule on a solid substrate surface, and monitoring its interaction with an analyze in a sample solution. Furthermore, nanotechnology can be applied to make high-throughput analyses that are smaller, faster and sensitive than conventional assays. Some nanomaterials or nanofabricated surfaces can be coupled to biomolecules and used in antibody-based assays and enzymatic methods for pesticide residues. The operation procedure has become more convenient as it does not require labeling procedure. In this paper, we review the recent advances in agrichemical defection research and also describe the label-free biosensor for pesticides using various useful detection methods.

Metal Oxide Thin Film Transistor with Porous Silver Nanowire Top Gate Electrode for Label-Free Bio-Relevant Molecules Detection

  • Yu, Tae-Hui;Kim, Jeong-Hyeok;Sang, Byeong-In;Choe, Won-Guk;Hwang, Do-Gyeong
    • Proceedings of the Korean Vacuum Society Conference
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    • 2016.02a
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    • pp.268-268
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    • 2016
  • Chemical sensors have attracted much attention due to their various applications such as agriculture product, cosmetic and pharmaceutical components and clinical control. A conventional chemical and biological sensor is consists of fluorescent dye, optical light sources, and photodetector to quantify the extent of concentration. Such complicated system leads to rising cost and slow response time. Until now, the most contemporary thin film transistors (TFTs) are used in the field of flat panel display technology for switching device. Some papers have reported that an interesting alternative to flat panel display technology is chemical sensor technology. Recent advances in chemical detection study for using TFTs, benefits from overwhelming progress made in organic thin film transistors (OTFTs) electronic, have been studied alternative to current optical detection system. However numerous problems still remain especially the long-term stability and lack of reliability. On the other hand, the utilization of metal oxide transistor technology in chemical sensors is substantially promising owing to many advantages such as outstanding electrical performance, flexible device, and transparency. The top-gate structure transistor indicated long-term atmosphere stability and reliability because insulator layer is deposited on the top of semiconductor layer, as an effective mechanical and chemical protection. We report on the fabrication of InGaZnO TFTs with silver nanowire as the top gate electrode for the aim of chemical materials detection by monitoring change of electrical properties. We demonstrated that the improved sensitivity characteristics are related to the employment of a unique combination of nano materials. The silver nanowire top-gate InGaZnO TFTs used in this study features the following advantages: i) high sensitivity, ii) long-term stability in atmosphere and buffer solution iii) no necessary additional electrode and iv) simple fabrication process by spray.

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Fabrication of Flexible Surface-enhanced Raman-Active Nanostructured Substrates Using Soft-Lithography

  • Park, Ji-Yun;Jang, Seok-Jin;Yeo, Jong-Seok
    • Proceedings of the Korean Vacuum Society Conference
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    • 2012.08a
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    • pp.411-411
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    • 2012
  • Over the recent years, surface enhanced Raman spectroscopy (SERS) has dramatically grown as a label-free detecting technique with the high level of selectivity and sensitivity. Conventional SERS-active nanostructured layers have been deposited or patterned on rigid substrates such as silicon wafers and glass slides. Such devices fabricated on a flexible platform may offer additional functionalities and potential applications. For example, flexible SERS-active substrates can be integrated into microfluidic diagnostic devices with round-shaped micro-channel, which has large surface area compared to the area of flat SERS-active substrates so that we may anticipate high sensitivity in a conformable device form. We demonstrate fabrication of flexible SERS-active nanostructured substrates based on soft-lithography for simple, low-cost processing. The SERS-active nanostructured substrates are fabricated using conventional Si fabrication process and inkjet printing methods. A Si mold is patterned by photolithography with an average height of 700 nm and an average pitch of 200 nm. Polydimethylsiloxane (PDMS), a mixture of Sylgard 184 elastomer and curing agnet (wt/wt = 10:1), is poured onto the mold that is coated with trichlorosilane for separating the PDMS easily from the mold. Then, the nano-pattern is transferred to the thin PDMS substrates. The soft lithographic methods enable the SERS-active nanostructured substrates to be repeatedly replicated. Silver layer is physically deposited on the PDMS. Then, gold nanoparticle (AuNP) inks are applied on the nanostructured PDMS using inkjet printer (Dimatix DMP 2831) to deposit AuNPs on the substrates. The characteristics of SERS-active substrates are measured; topology is provided by atomic force microscope (AFM, Park Systems XE-100) and Raman spectra are collected by Raman spectroscopy (Horiba LabRAM ARAMIS Spectrometer). We anticipate that the results may open up various possibilities of applying flexible platform to highly sensitive Raman detection.

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Detection of the Fungicide Iprovalicarb Residues Using a Surface Plasmon Resonance Biosensor (표면플라즈몬공명 바이오센서를 이용한 살균제 Iprovalicarb 잔류물의 검출)

  • Kim, Woon-Ho;Cho, Han-Keun;Kyung, Kee-Sung;Kim, Gi-Young
    • Journal of Biosystems Engineering
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    • v.34 no.1
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    • pp.50-56
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    • 2009
  • Surface plasmon resonance (SPR) biosensor has been used to detect many biochemical reactions, because this label-free sensor has high sensitivity and rapid response. The reactions are monitored by refractive index changes of the SPR biosensor. Iprovalicarb is protective, curative, and eradicative systemic fungicide introduced by Bayer AG in 1999. It has potential far control of downy mildew infesting onion, cucumber, grape and melon, late blight infesting tomato and potato, and anthracnose infesting watermelon and pepper. It is strictly limited to the maximum residue limit. In this study, the applicability of a portable SPR biosensor (Spreeta, Texas instrument, TX, USA) to detect the iprovalicarb residue was examined. The sensor chip was adopted to detect the reaction of iprovalicarb to immobilized iprovalicarb-antibody. The binding of the iprovalicarb onto the biosensor surface was measured by change of the refractive index (RI). Characteristics of the sensor chip including specificity, sensitivity, stability, and reusability were analyzed. In calibration test for seven levels of iprovalicarb concentration (0.32 to 5,000 mg/L) with three replications, a Sigmoidal model with Hill function was obtained between relative RI value and the iprovalicarb concentration with R-square of 0.998. It took 30 minutes to complete a set of detecting assay with the SPR biosensor.

Proposal of autonomous take-off drone algorithm using deep learning (딥러닝을 이용한 자율 이륙 드론 알고리즘 제안)

  • Lee, Jong-Gu;Jang, Min-Seok;Lee, Yon-Sik
    • Journal of the Korea Institute of Information and Communication Engineering
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    • v.25 no.2
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    • pp.187-192
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    • 2021
  • This study proposes a system for take-off in a forest or similar complex environment using an object detector. In the simulator, a raspberry pi is mounted on a quadcopter with a length of 550mm between motors on a diagonal line, and the experiment is conducted based on edge computing. As for the images to be used for learning, about 150 images of 640⁎480 size were obtained by selecting three points inside Kunsan University, and then converting them to black and white, and pre-processing the binarization by placing a boundary value of 127. After that, we trained the SSD_Inception model. In the simulation, as a result of the experiment of taking off the drone through the model trained with the verification image as an input, a trajectory similar to the takeoff was drawn using the label.

Inflammation Scan Using $^{99m}Tc-HMPAO$ Labelled Leukocytes ($^{99m}Tc-HMPAO$를 이용한 자가백혈구표지 및 그를 이용한 염증병소의 스캔)

  • Yang, Woo-Jin;Chung, Soo-Kyo;Shinn, Kyung-Sub;Bahk, Yong-Whee;Kim, Hoon-Kyo
    • The Korean Journal of Nuclear Medicine
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    • v.23 no.2
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    • pp.219-223
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    • 1989
  • Inflammation scan using radiolabelled leukocytes has high sensitivity and specificity. Several methods for labelling leukocytes have been evaluated using P-32 diisopropyl fluorophosphate (DFP-32), H-3 thymidine, Cr-51 chromate, Ga-67 citrate and Tc-99m-sulfur colloid. In-111-oxine has proved so far to be the most reliable agent for labelling leukocytes. In-111-oxine is, however, expensive, not easily available when needed, and its radiation dose to leukocytes is relatively high. Moreover, resolution of the resultant image is relatively poor. Tc-99m is still the agent of choice because of, as compared with the indium, its favorable physical characteristics, lower cost and availability. Now the technique for labelling the leukocytes with technetium is successfully obtained using the lipophilic HAPAO with higher efficiency for granulocytes than for other cells. With this technique it is possible to label leukocytes in plasma to improve the viability of the leukocytes. Inflammation scan using Tc-99m-HMPAO has been evaluated in several laboratories, and difference in methods for separation and labelling accounts for difference in efficiency, viability and biodistribution of the labelled leukocytes. We performed inflammation scan using leukocytes labelled with Tc-99m-HMPAO in three dogs 24 hours after inoculation of live E. Coli and A. Aureus in their right abdominal wall. We separated mixed leukocytes by simple sedimentation using 6% hetastarch (HES) and labelled the leukocytes with Tc-99m-HMPAO in 20% cell free plama diluted with phosphate buffer solution(Fig. 1). Uptake was high in the liver and spleen but is was minimal in the lungs on whole body scan. Kidneys and intestine showed minimal activity although it was high in the urinary bladder(Fig. 2). Uptake of labelled leukocytes in the inflammation site was do(mite on 2 hour-postinjection scan and abscess was clearly delineated on 24 hour-delayed scan with high target-to-nontarget ratio(Fig. 3, 4). Inflammation scan using mixed leukocytes labelled with Tc-99m-HMPAO is very sensitive and specific in early detection of inflammation.

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Detection of Undeclared Betamethasone Derivatives in Cosmetic Products Labeled to Contain Zinc Pyrithione as the Active Ingredient (아연피리치온을 유효성분으로 표기한 화장품류에서 미표기 성분인 베타메타손 유도체의 검출)

  • Lee, Jeong-Pyo;Park, Sung-Hwan;Yang, Seong-Jun;Kim, Sun-Mi;Son, Kyung-Hun;Yun, Mi-Ok;Choi, Sang-Sook
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.35 no.1
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    • pp.11-17
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    • 2009
  • Betamethasone propionate, an anti-inflammatory glucocorticosteroid, was detected in cosmetics with no indication on the label of this compound as an ingredient. The product was formulated as a topical spray or shampoo and labeled to contain zinc pyrithione as the active ingredient. A thin-layer chromatographic analysis was carried out on silica gel plates to provide a first indication about the presence of a compound with steroid structure and reactivity; then high-performance liquid chromatography (HPLC) separation allowed the identification of the corticosteroid agent and its quantification. To identify the corticosteroid agent from these commercial samples we collected the fractions suspected to have ketol steroids by prep HPLC and identified the compound as betamethasone propionate by NMR and MS spectrometry. Then we synthesized the standard for the betamethasone 17-propionate and 21-propionate and quantitate the corticosteroids from the sample by HPLC with that standards. By this method we identified the corticosteroid compounds from some commercial cosmetics such as zinc pyrithione sprays. The finding of betamethasone propionate in the products was shown by comparison to an authenticated standard of betamethasone propionate by retention time on reverse-phase HPLC. Two of the tested products contained betamethasone propionate at the levels of 0.005 ${\sim}$ 0.02% and the others were free of betamethasone propionate.