BioChip journal (BIOCHIP J)

Publications in this journal

• Article: Identification of survival factors in LPS-stimulated anthrax lethal toxin tolerant RAW 264.7 cells through proteomic approach

  Amitabh Das, Nando Dulal Das, Ji Hyun Park, Hyung Tae Lee, Mi Ran Choi, Kyoung Hwa Jung, Young Gyu Chai
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  ABSTRACT: Anthrax lethal toxin (LeTx; a combination of protective antigen and lethal factor) is secreted by the vegetative cells of Bacillus anthracis and is cytotoxic for certain macrophage cell lines. First-time exposure of murine macrophage cells (RAW 264.7) to lethal toxin (LeTx) (0.1+0.1 mg/mL) caused extensive cell death with a survival rate of approximately 40%, but upon secondary exposure to LeTx and lipopolysaccharide (LPS) (1 μg/mL), after a few passages, these cells had a survival rate of approximately 100%. The present study assessed protein expression changes after LPS exposure to LeTx-intoxication-resistant RAW 264.7 cells. To analyze the protein expression profile of LPS-treated LeTx-intoxication-resistant RAW 264.7 cells, we employed matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDITOF MS), and later, Ingenuity Pathways Analysis (IPA) was applied to establish the molecular networks. Among the differentially expressed proteins were voltage dependent anion channel 1, jip3 protein, heat shock protein 4, tubulin beta, 26S protease regulatory subunit 4, and DNA polymerase delta subunit 4 (DNA polδ) were significant. The molecular network signatures and functional proteomics obtained in this study may facilitate the evaluation of potential pathways such as PI3K signaling, NF-κB signaling, and LPSinduced MAPK signaling for the recovery of LPSinduced LeTx-intoxication-resistant RAW 264.7 cells.
  BioChip journal 03/2013; 7(1):75-84.
• Article: Design of Disposable DNA Biosensor Microchip with Amperometric Detection Featuring PCB substrate

  M. H. Ghanim and M. Z. Abdullah
  BioChip journal 03/2013; 7(1).
• Article: Axon orientation by gradient of cytochalasin D inside microfluidic device

  L Xiao, SK Mahto, SW Rhee
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  ABSTRACT: This paper describes the development of a microfluidic device for axon orientation. The device was fabricated with poly (dimethylsiloxane) by soft lithography and replica molding. There were two separated rooms in the device, and a control system and stable concentration gradient system were maintained in each room for at least 24 h. Finally, to demonstrate a practical application of the device, neuronal cells were cultured in both cell rooms, and the growth of cells and formation of axons were monitored under a concentration gradient of cytochalasin D for 8 h. Quantitative analysis of the orientation of axons was analyzed, and the data were plotted in a polar graph as a function of the direction. A figure of 33.8% of positive responding neurons were found in the cytochalasin D gradient room, while that figure was 23.3% of cells in control room. The extent of axon orientation shows a considerable difference between the two rooms, and the device developed here is capable of investigating axon orientation and can be used for other biological applications.
  BioChip journal 12/2012; 6(4):335-341.
• Article: Identification and characterization of flowering repressor-related genes in Chinese cabbage

  ChangKug Kim, Yeon-Hee Lee, Joon-Ki Hong, DongSuk Park, Mi-Kyoung Kim, MinSeok Cho, YongKab Kim, JangHo Hahn
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  ABSTRACT: In this study, we used a five-step process to identify those genes most likely involved in flowering repression in the Chinese cabbage (Brassica rapa). We tested 6,275 candidate genes with 300K microarrays, which included specific gene expression profiles of FLOWERING LOCUS C (FLC) mutants and normal cultivars during five cold vernalization stages. From that, we identified 289 transcription factor genes and 59 pathway network genes associated with floweringrelated metabolism. Then we compared the 348 genes to 1,287 genes from Gene Ontology and Clusters of Orthologous Groups analyses, which use similar orthologs to categorize conserved genes. Those analyses revealed 10 hypothetical genes for B. rapa, which we verified by reverse transcription-polymerase chain reaction. The final selected genes most likely play regulatory roles in either B. rapa flowering time control or flowering repression during vernalization. While these final genes require further characterization and validation, our study illustrates the usefulness of a multi-layered screening method after initially identifying genes from microarrays.
  BioChip journal 08/2012; 6(2):120-127.
• Article: Lipopolysaccharide-mediated protein expression profiling on neuronal differentiated SH-SY5Y cells

  Nando Dulal Das, Mi Ran Choi, Kyoung Hwa Jung, Ji Hyun Park, Hyung Tae Lee, Seung Hyun Kim, Young Gyu Chai
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  ABSTRACT: Neuroinflammation can contribute to neuronal dysfunction, death and several neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, and multiple sclerosis. Lipopolysaccharide (LPS)-induced neuroinflammation severely affects neurons and can contribute to neuronal dysfunction and degeneration by causing the release of inflammatory and neurotoxic factors. We evaluated the long-term effects of treating differentiated SH-SY5Y cells with LPS to mimic LPS-induced neuroinflammation. Using matrix assisted laser desorption ionization-time of flight mass spectrometry and MetaCore pathway analysis software (GeneGo), the proteomic expression profiles of differentiated SH-SY 5Y cells after LPS treatment was studied to determine the inflammatory effects on the process of SH-SY5Y differentiation. Long-term LPS treatment resulted in the upregulation of phosphodiesterase 4B (PDE4B), slit robo GTPase (SRGAP2), transcription repressor E2F-6, vimentin, and 70 kDa heat shock protein 9 (Mortalin/HSPA9). Taken together, our results suggest that LPS-treated differentiation of SH-SY5Y cells can lend insight into the multiple pathways involved in neurological diseases.
  BioChip journal 06/2012; 6(1):165-173.
• Article: Species identification of filefishes (Monacanthidae) using DNA microarray in Korean marketplace

  Ji-Hoon Kim, Jung Youn Park, Jin-Wook Jung, Mi-Jung Kim, Won Sun Lee, Cheul Min An, Jung-Ha Kang, Seung Yong Hwang
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  ABSTRACT: Two species of filefish, Spephanolepis cirrhifer and Thamnaconus modestus, are commonly dried, seasoned with sugar and salt, roasted, and sold as a snack called Jwipo in Korea. These species of fish are imported from China, Thailand, and Vietnam because of drastic catch reduction. However, imported materials have been identified as Aluterus monoceros, Paramonnacathus choirocephalus, and T. septentrionalis in a market survey. For these reasons, we developed a DNA microarray that distinguishes five species of filefish for quick and simple species identification. Species-specific oligonucleotide probes were designed by sequence analysis of mitochondrial cytochrome c oxidase subunit I. In this study, a DNA microarray system using species-specific probes successfully and rapidly identified five different filefish species, and shown potential for determining geographical origin. KeywordsFilefish–DNA microarray–Species identification–Jwipo–Cytochrome c oxidase subunit I
  BioChip journal 05/2012; 5(3):229-235.
• Article: Fluorescence detection by miniaturized instrumentation based on non-cooled CCD minicamera and dedicated for lab-on-a-chip applications

  Rafał Walczak
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  ABSTRACT: Due to high specificity and sensitivity, fluorometric detection is one of the main detection methods applied in analytical microsystems commonly known as lab-on-a-chip (LOC). In most cases, optical instrumentation for fluorescence induction and detection is based on configuration and components “borrowed” from large laboratory instruments based on an epifluorescence microscope. As a result, the optical instrumentation surrounding lab-on-a-chip is bulky, expensive and dedicated for operation only inside laboratories. In this paper a brief discussion on fluorescence detection in lab-on-a-chip is presented. Next, novel low-cost detection instrumentation utilising noncooled CCD image sensor and semiconductor laser light source is described. The instrumentation is dedicated for operation in portable and low-cost devices for different LOC-based life science applications. Finally, an example of application of the novel method and instrumentation co-working with lab-on-a-chip for real-time PCR detection of food pathogens is briefly described. KeywordsLab-on-a-chip–Fluorescence–Non-cooled CCD–Real-time PCR
  BioChip journal 05/2012; 5(3):271-279.
• Article: Knowledge based construction of functional modules for genetic network in Saccharomyces Cerevisiae

  Soo Young Cho, Sung Ho Park, Jin Choul Chai, Young Seek Lee
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  ABSTRACT: Methods of module prediction typically involve clustering of group genes and the respective transcription factors. These methods are based on the hypothesis that a gene is regulated by a transcription factor when the expression patterns of the gene and the transcription factor are similar. However, this method is not able to predict the direct target of a transcription factor or its effects on the gene expression. In this study, we propose a new approach that uses data integration in order to predict transcription mechanisms, i.e. the targets of transcription factors and the effects on gene expression. We analyzed yeast ChIPchip data, and DNA microarray data obtained under various physiological conditions. We predicted the functional classification of unknown genes using a Support Vector Machine. We validated our results by comparing with other module prediction programs, and found that our module prediction method shows a higher accuracy than others module prediction programs. KeywordsTranscription factor–Module network–Systems biology–Bioinformatics
  BioChip journal 05/2012; 5(2):145-150.
• Article: Fluorescence immunoassay of anti-cyclic citrulinated peptide (CCP) autoantibodies by using parylene-H film

  Hyuk Ko, Ga-yeon Lee, Byoung-Jin Jeon, Jae-Chul Pyun
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  ABSTRACT: A fluorescence immunoassay for rheumatoid arthritis was presented by immobilizing a short peptide cyclic citrullinated peptide (CCP) through parylene-H film. The parylene-H film is a polymer of pxylene modified with formyl groups which can react with primary amine groups of proteins or peptides. In this work, the covalent coupling of the peptide can be produced by only one step of incubation without additional coupling reagent, and the immobilization efficiency to the parylene-H film was compared with the immobilization of CCP through conventional physical adsorption. The applicability of this immobilization method for short peptides is demonstrated by detecting autoantibodies in rheumatoid arthritis patient serum. KeywordsParylene–Immobilization–Cyclic citrullinated peptide–Rheumatoid arthritis–Fluorescence immunoassay
  BioChip journal 04/2012; 5(3):242-245.
• Article: Differential gene expression following ionizing radiation in multicellular spheroid depending on p53 status: identification of potential targets and prediction of responsive signaling pathways

  Jee Young Kwon, Young Rok Seo
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  ABSTRACT: Ionizing radiation (IR) therapy is a potential treatment option of several solid tumors; however, the molecular responses of carcinoma cells to IR are not yet clarified. A multicellular spheroid (MCS)-in vivo mimic model was established in which allow us to in situ study of cellular response to IR in human carcinoma cells with different intrinsic p53 status. Here, relative cell survival rate was determined in p53-wild and-null type cells of MCS in comparison to that of monolayer culture in response to 10 Gy IR by trypan blue exclusion assay. It obviously showed that under MCS culture system radioresistance phenotype was dependent on p53 status since p53-null type MCS exhibited significantly higher cell survival rate, in contrast to p53-wild type MCS. In order to screen molecular targets in p53-proficient cancer cells upon IR exposure, we conducted microarray under 10 Gy IR to observe gene expression pattern change in MCS compared to monolayer cells, with and without p53. In addition, potential molecular network was analyzed using Pathway Studio software to define responsive signaling and interactions. Total 478 genes were notably altered at transcript level toward the IR treatment. Discovered genes were participated in several cellular major processes, including apoptosis (both caspase-dependent and-independent pathways), cell migration and proliferation. Among them ANXA11, C2, KCNE2, KIF3C, MSH5, and OSCAR were differentially expressed in p53-proficient MCS but constant in p53-deficient MCS. These genes might be considered as target molecules for evaluation of IR efficiency in p53-proficient MCS as a typical 3D-in vivo mimic model. Our findings demonstrate complex responses of p53-carcinoma cells following IR exposure including putative signaling pathways, leading to emphasis on the importance of in vivo mimic MCS model rather than conventional monolayer culture system. KeywordsApoptosis–Ionizing radiation (IR)–Multicellular spheroid (MCS)–p53–Radioresistance
  BioChip journal 04/2012; 5(3):280-288.
• Article: Genome-wide gene expression analysis of Patrinia scabiosaefolia reveals an antibiotic effect

  Eun-Kyeong Choi, Yeo eun Park, Bo Won Choi, Ki-Suk Kim, Hea Jung Yang, Kwang Seok Ahn, Hyeung-Jin Jang
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  ABSTRACT: Patrinia scabiosaefolia (PS) clinically had been prescribed in oriental ethnopharmacology, but its exact mechanism is unknown. In this study, we analyzed molecular perspective of antibiotic effects of PS. Escherichia coli O157:H7 was changed in four main mechanisms; (1) Cell wall biosynthesis decreased, (2) DNA transcription consists of folic acid metabolism interruption, (3) Protein synthesis and folding declined, (4) Bacterial cell motility increased. With these results, antibiotic effect was demonstrated at a molecular perspective of PS. Safe and effective drugs for antibiotic resistance will promote with the multi-target herbal medicine. Keywords Patrinia scabiosaefolia –Herbal medicine– Escherichia coli O157–Antibiosis–Bacterial cell wall–Folic acid
  BioChip journal 04/2012; 5(3):246-254.
• Article: Compartmented microfluidic device for positioning and chemotactic migration of cells

  Seog Woo Rhee
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  ABSTRACT: This paper describes a simple approach to position cells and monitor chemotactic migrations of MDA-MB 231 human breast cancer cells. The device was fabricated in poly(dimethylsiloxane) using soft lithography and micromolding techniques. The device has three compartments with volumes less than 2 μL in each channel. The middle compartment is separated by a physical barrier in which a number of small microgrooves are embedded to allow diffusion of chemicals and migration of cells. The three-compartment diffusion system generated a steady state gradient with an exponential shape across the middle channel. It gives two extreme regions for real time monitoring of cellular behaviors: control and gradient regions. To demonstrate the utility of the device for cell migration, chemotaxis of breast cancer cells was successfully observed in a soluble gradient of chemoattractant (EGF). During the chemotaxis experiment, most cells in the control region remained along the barrier and a few cells (11.4%) migrated towards the side channel while many of the cells (31.6%) in the gradient region migrated towards the side channel containing EGF. This device is expected to be applicable as an alternative method for the investigation of chemotactic migration of cells. KeywordsMicrofluidics–Positioning cells–Suction technique–Chemotaxis–Breast cancer cells
  BioChip journal 04/2012; 5(2):129-136.
• Article: Analysis of chemical/biochemical conversions on gold microparticles using MALDI-TOF MS

  Jae Il Kim, Sohyun Kim, Woon-Seok Yeo
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  ABSTRACT: Chemical and biochemical analyses on solid supports are now in common because of the feasibility and simplicity of the process. For example, post-translational modifications of proteins such as phosphorylation, acetylation, and glycosylation have been assayed on several types of solid supports, mostly on 2-dimensional formats. Use of 3-dimensional solid supports, such as gold microparticles in this study, is beneficial over 2-dimensional biochips in terms of larger active surface areas and easier handling protocol. In this report, we present analyses of chemical conversions as well as enzymatic conversions on gold microparticles using matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). For immobilization of enzyme substrates, gold microparticles, which presented organic functional groups, were subjected to a series of chemical reactions encompassing amide coupling and Michael addition. The resulting particles were then treated with enzymes. Direct analysis of the resulting products on microparticles by MALDI-TOF MS was performed without additional labeling steps, called SAMDI (self-assembled monolayers for MALDI). The mass spectra clearly showed chemical modifications and enzymatic conversions of peptide substrates on gold microparticles. We expect that our method can be used routinely in biological research, such as enzyme inhibitor assays, and development of substrate peptide of enzymes. We believe that the combination of gold microparticles with MALDI-TOF MS will provide an efficient analytical platform for use in various biochemical studies. KeywordsGold microparticles–Kinase–Mass spectrometry–Monolayers–Peptidase
  BioChip journal 04/2012; 5(3):199-205.
• Article: Methylglyoxal-mediated alteration of gene expression in human endothelial cells

  Seung Eun Lee, Hana Yang, Seong Il Jeong, Young-Ho Jin, Cheung-Seog Park, Yong Seek Park
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  ABSTRACT: Endothelial dysfunction is an important factor in the development of vascular diseases such as atherosclerosis, hypertension and diabetes. Methylglyoxal (MG) is a highly reactive dicarbonyl metabolite that is an extremely toxic glucose degradation product with strong oxidative activity. MG is involved in the pathogenesis of vascular complications of diabetes. Several studies have reported increased MG levels in pathology models of vascular injury. The present study investigated the genome-wide transcriptional responses of human umbilical vein endothelial cells (HUVECs) exposed to MG by microarray gene expression profiling. As a result, we identified 1,624 genes that were 1.5-fold up-or down-regulated within 12 h of MG treatment. The differentially expressed genes that were dysregulated in many biological processes included inflammatory responses, cell cycle, apoptosis, and cell adhesion. These results demonstrate the MG induced genome-wide alterations in expression profile in human endothelial cells and indicate that MG may cause cytotoxicity and tissue injury in the human endothelium. The data supports the view that MG-stimulated changes in gene expression contribute to the development of vascular disease. KeywordsMethylglyoxal–Gene expression profile–Vascular disease–Endothelial cells
  BioChip journal 04/2012; 5(3):220-228.
• Article: Computational identification of Chinese cabbage anthocyaninspecific genes

  ChangKug Kim, JinA Kim, Shoshi Kikuchi, JiWeon Choi, YongKab Kim, HyunJu Park, YoungJoo Seol, DongSuk Park, JangHo Hahn, YongHwan Kim
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  ABSTRACT: Therefore, 3,895 candidate genes related to anthocyanin biosynthesis in Chinese cabbage were identified using a 300K Brassica rapa microarray analysis. Gene expression during six stages of leaf developmental stages were examined in FC (green leaf) and FA(red leaf) Chinese cabbage cultivars. The 317 transcription factor genes found to be associated with anthocyanin were classified into 11 functional groups. The ratio of expression levels of each transcription factor between the two cultivars was examined during the six leaf developmental stages. A total of 14 genes were found to be expressed in all developmental stages commonly. Among these genes, 10 unknown and hypothetical genes were differentially revealed to be expressed between the two cultivars at each developmental stage, as determined by microarray analysis, and were verified by RT-PCR validation. These genes most likely play regulatory roles in either anthocyanin production or metabolism during flavonoid biosynthesis. While these genes require further validation and characterization, our results illustrate the potential usefulness of this multi-layered screening method using Chinese cabbage (Brassica rapa) microarrays. KeywordsChinese cabbage– Brassica rapa –Microarray–Anthocyanin genes–Transcription factor
  BioChip journal 04/2012; 5(2):184-192.
• Article: Comparison of genomic profiles in human neuroblastic SH-SY5Y and substrate-adherent SH-EP cells using array comparative genomic hybridization

  Jin Hwan Do, In Su Kim, Jae Dong Lee, Dong-Kug Choi
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  ABSTRACT: Human neuroblastoma is the most commonly diagnosed solid tumor in children. The existence and development of S-type cells is important for the prognosis and malignant properties in neuroblastoma patients. However, their origin is controversial and the relationship between S- and N-type cells in neuroblastoma has not yet been clarified. To investigate the feasibility of inter-conversion and characteristic features between S-type cells lacking malignant potential and N-type cells having metastatic potential, the genomic profiles of neuroblastoma SH-EP (S-type) and SH-SY5Y (N-type) cells were compared at high resolution. Common gain segments (>10 Mb) between SH-EP and SH-SY5Y cells were observed at 1q21.1–q44 and 17q21.32–q25.3. The results of the fluorescent in situ hybridization (FISH) analysis showed good agreement with the array-CGH data. Genome-wide inspection of SH-EP and SH-SY5Y cells successfully identified not only common chromosomal aberrations but also genomic variations, suggesting that interconversion could not take place between S- and Ntype cells. The identified differences between less aggressive S-type cells (SH-EP) and highly aggressive neuroblastic N-type cells (SH-SY5Y) might be useful for understanding tumorigenicity and discovery of potential new markers in neuroblastoma. This is the first effort to compare genomic profiles between less aggressive S-type cells (SH-EP) and highly aggressive neuroblastic N-type cells (SH-SY5Y) at high resolution. KeywordsCopy number variation–Neuroblastoma SH-EP and SH-SY5Y–Array-based comparative genomic hybridization (array-CGH)–Inter-conversion
  BioChip journal 04/2012; 5(2):165-174.
• Article: Detection of central single-nucleotide mismatches in short duplex DNAs on hyper-branched amine surfaces

  Eung-Sam Kim, Chang Ok Kim, Joon Won Park, Kwan Yong Choi
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  ABSTRACT: High capability to distinguish single-nucleotide mismatches of genes using short oligonucleotide probes is essential in diagnostic methods for identification of point mutations and single nucleotide polymorphisms. To investigate the feasibility of using an aziridine-treated surface containing hyper-branched amine groups to discriminate single-nucleotide mismatches in a human gene, target probes for exons 5–8 of the p53 gene from liver cancer cells were hybridized with four types of surface-bound capture probes, one for perfect match and three for central single-nucleotide mismatches. The aziridine slide with high DNA-loading capacity exhibited greater ability to detect single-nucleotide mismatch than did the generic amine slide. When a T30 tether was linked to the capture probe, the mismatch discrimination capability increased when using a chemical cross-linker, but decreased when using UV irradiation for cross-linking. DNA duplexes had lower melting temperatures when the single-nucleotide mismatch was in the central region than when it was in the terminal region regardless of the type of mismatched nucleotide. Our results suggest that capture probes attached to the aziridine surface can effectively identify point mutations in a genomic sequence or and can estimate the affinity of gene-specific antisense oligonucleotide probes. Keywordsp53 gene–Point mutation–Melting temperature–Aziridine–DNA microarray
  BioChip journal 04/2012; 5(2):137-144.
• Article: Exposure of BALB/3T3 fibroblast cells to temporal concentration profile of toxicant inside microfluidic device

  Hyunjong Shin, Sanjeev Kumar Mahto, Jae-Hyun Kim, Seog Woo Rhee
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  ABSTRACT: This work describes a simple method that generates temporal concentration profiles of chemicals inside a microfluidic channel and exposes cultured cells to chemicals in order to mimic the exponential clearance curve of chemicals in the human body. The device fabricated with a poly (dimethylsiloxane) (PDMS) replica mold and a glass substrate was used to create the microfluidic channels. The fluid streams inside the channels were controlled by programmable syringe pumps. By controlling the relative flow rates of the fluidic inlets using separate syringe pumps, the resulting composition of the inlets that feed the concentration generator zone can be controlled to generate temporal profiles of chemicals in the observation zone. The concentration profiles of fluorescent dye were confirmed by analyzing the time-lapse fluorescence images. Finally, a temporal concentration profile of toxic cadmium solution with exponential clearance curve was exposed to BALB/3T3 fibroblast cells confluently cultured inside a microfluidic device for 3 h, and the behavior of the cells was monitored using a time-lapse microscope. The relative concentration of reactive oxygen species inside the exposed cells was measured indirectly by using a fluorescent probe to assess the degree of cell death. The method developed in this work offers a convenient way of controlling the concentration of chemicals inside microfluidic channels, and enables time-dependent exposure of chemicals to cells and monitoring of the behavior of cells for biological research applications. KeywordsTemporal concentration profile–Time-dependent exposure–Cell death–Microfluidics–Cell death
  BioChip journal 04/2012; 5(3):214-219.
• Article: Graphene-based electrochemical biosensor for pathogenic virus detection

  Fei Liu, Ki Seok Choi, Tae Jung Park, Sang Yup Lee, Tae Seok Seo
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  ABSTRACT: A graphene based biosensor system is presented for performing highly sensitive pathogenic virus detection. A free-standing conductive graphene film was prepared as a novel electrochemical sensor through two steps: synthesis of a graphene oxide (GO) film from GO colloidal suspensions by using a speed vacuum concentrator and thermal annealing process at 900°C with H2/Ar flow to generate a reduced GO film. The resultant graphene film shows an excellent electron transfer property on the surface in the [Fe(CN)6]3−/4− redox system and is used as a working electrode in the electrochemical biosensor. The surface of graphene is modified with pyrene derivatives, and then covalently linked with virus-specific antibodies. The target cell, rotavirus, is captured on the graphene film through antibody-antigen interaction, and the entire process was monitored by cyclic voltammetric responses. A 105 pfu/mL of input cells is detected with ca. 30.7% sensitivity, and ca. 1.3% sensitivity is measured with 103 pfu/mL of input cells, demonstrating that graphene film based electrode can be applied for electrochemical biosensor. KeywordsGraphene film–Electrochemical biosensor–Pathogen detection–Detection sensitivity
  BioChip journal 04/2012; 5(2):123-128.