• Biomolecules & Therapeutics
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• 제20권1호
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• pp.19-26
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• 2012

There are many cyclic peptides with diverse biological activities, such as antibacterial activity, immunosuppressive activity, and anti-tumor activity, and so on. Encouraged by natural cyclic peptides with biological activity, efforts have been made to develop cyclic peptides with both genetic and synthetic methods. The genetic methods include phage display, intein-based cyclic peptides, and mRNA display. The synthetic methods involve individual synthesis, parallel synthesis, as well as split-and-pool synthesis. Recent development of cyclic peptide library based on split-and-pool synthesis allows on-bead screening, in-solution screening, and microarray screening of cyclic peptides for biological activity. Cyclic peptides will be useful as receptor agonist/antagonist, RNA binding molecule, enzyme inhibitor and so on, and more cyclic peptides will emerge as therapeutic agents and biochemical tools.

• Bulletin of the Korean Chemical Society
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• 제32권1호
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• pp.247-250
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• 2011

We designed an effective screening method for double strand DNA (dsDNA) binders using DNA-modified magnetic particles. Hairpin DNA was immobilized on the surface of magnetic particle for a simple screening of dsDNA binding materials in a solution containing various compounds. Through several magnetic separation and incubation processes, four DNA-binding materials, DAPI, 9AA, AQ2A, and DNR, were successfully screened from among five candidates. Efficiency of screening was demonstrated by HPLC analysis using a C2/18 reverse-phase column. In addition, their relative binding strengths were verified by measuring the melting temperature ($T_m$). If hairpin DNA sequence is modified for other uses, this magnetic bead-based approach can be applied as a high-throughput screening method for various functional materials such as anti-cancer drugs.

• KSBB Journal
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• 제28권3호
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• pp.185-190
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• 2013

For the selection of peptide specifically binding to $Pb^{2+}$, the biopanning with the commercially available Ph.D.-7 phage displayed heptapeptide library was carried out against $Pb^{2+}$ immobilized on a metal-chelating IDA (iminodiacetic acid) resin. After four rounds of screening against $Pb^{2+}$-IDA including negative selections against charged bead with metal ions other than $Pb^{2+}$ and uncharged bead, several candidate lead-binding phage peptides were initially determined based on the order of frequency from the screened phage clones. Of the selected phage peptide sequences, the peptide of the highest frequency, CysSerIleArgThrLeuHisGlnCys (CSIRTLHQC) also exhibited the strongest affinity for $Pb^{2+}$ in binding assays for individual phage clones. However, there was not a significant difference in $Pb^{2+}$ affinity between selected peptides when using synthetic heptapeptides corresponding to the displayed peptide sequences of phage clones.

• 대한의생명과학회지
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• 제2권2호
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• pp.231-240
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• 1996

인체 혈액 응고 9인자는 간에서 생성되며 461개의 아미노산으로 구성된 당 단백질이다. 따라서 인체 혈액 응고 9인자 cDNA를 찾기 위해 태아의 간(fetal liver) cDNA library를 PCR(Polymerase Chain reaction) 방법으로 screening하였으며, 그 결과 ATG개시 코돈으로부터 TAA종료 코돈까지 포함하는 1.4 kb의 9인자 cDNA를 찾았다. 또한 클론된 9인자 cDNA를 박테리아에서 발현시키기 위해 박테리아 발현 벡터인 pGEX-2T 플라스미드에 클로닝하므로써 pGEX-F9 플라스미드를 제조하였다. pGEX-F9로 형질전환된 E. coli에서 PGEX-F9의 발현을 유도하면 73 kDa 크기의 GST-factor9 융합 단백질이 다량생성되며 , 이 단백질이 혈액 응고 9인자 단백질을 함유하는 융합 단잭질임을 혈액 응고 9인자 항체를 이용한 Western blot으로 입증하였다. E. coli에서 발현된 GST-factor 9 융합 단백질은 전체 단백질의 약 20%를 차지하며 GST agarose bead를 이용한 one step purificarion 방법을 통해 GST-factor9 융합 단백질을 쉽게 분리 할 수 있다.

• 대한기계학회:학술대회논문집
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• 대한기계학회 2008년도 추계학술대회A
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• pp.1685-1686
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• 2008

This paper presents the design and experiment results of a multi-layered microsystem for magnetic bead applications. The magneto-microfluidic device is designed for capable of separating magnetic beads. In the presence of the magnetic field, magnetic beads are attracted and moved to high gradient magnetic fields. A multi-layered microfluidic channel consists of top and bottom layers in order to separate magnetic beads in the vertical direction. Our channel is easily integrated magnetic cell sorter, especially on-chip microelectromagnet or permanent magnet device. Fast separation of magnetic beads in top and bottom channels can be used in high throughput screening to monitor the efficiency of blood and drug compounds.

• Asian Pacific Journal of Cancer Prevention
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• 제14권10호
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• pp.6069-6075
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• 2013

Background: At present, the diagnosis of colorectal cancer (CRC) requires a colorectal biopsy which is an invasive procedure. We undertook this pilot study to develop an alternative method and potential new biomarkers for diagnosis, and validated a set of well-integrated tools called ClinProt to investigate the serum peptidome in CRC patients. Methods: Fasting blood samples from 67 patients diagnosed with CRC by histological diagnosis, 55 patients diagnosed with colorectal adenoma by biopsy, and 65 healthy volunteers were collected. Division was into a model construction group and an external validation group randomly. The present work focused on serum proteomic analysis of model construction group by ClinProt Kit combined with mass spectrometry. This approach allowed construction of a peptide pattern able to differentiate the studied populations. An external validation group was used to verify the diagnostic capability of the peptidome pattern blindly. An immunoassay method was used to determine serum CEA of CRC and controls. Results: The results showed 59 differential peptide peaks in CRC, colorectal adenoma and health volunteers. A genetic algorithm was used to set up the classification models. Four of the identified peaks at m/z 797, 810, 4078 and 5343 were used to construct peptidome patterns, achieving an accuracy of 100% (> CEA, P<0.05). Furthermore, the peptidome patterns could differentiate the validation group with high accuracy close to 100%. Conclusions: Our results showed that proteomic analysis of serum with MALDI-TOF MS is a fast and reproducible approach, which may provide a novel approach to screening for CRC.

• 한국진공학회:학술대회논문집
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• 한국진공학회 2013년도 제45회 하계 정기학술대회 초록집
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• pp.88-89
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• 2013

A variety of influenza A viruses from animal hosts are continuously prevalent throughout the world which cause human epidemics resulting millions of human infections and enormous industrial and economic damages. Thus, early diagnosis of such pathogen is of paramount importance for biomedical examination and public healthcare screening. To approach this issue, here we propose a fully integrated Rotary genetic analysis system, called Rotary Genetic Analyzer, for on-site detection of influenza A viruses with high speed. The Rotary Genetic Analyzer is made up of four parts including a disposable microchip, a servo motor for precise and high rate spinning of the chip, thermal blocks for temperature control, and a miniaturized optical fluorescence detector as shown Fig. 1. A thermal block made from duralumin is integrated with a film heater at the bottom and a resistance temperature detector (RTD) in the middle. For the efficient performance of RT-PCR, three thermal blocks are placed on the Rotary stage and the temperature of each block is corresponded to the thermal cycling, namely $95^{\circ}C$ (denature), $58^{\circ}C$ (annealing), and $72^{\circ}C$ (extension). Rotary RT-PCR was performed to amplify the target gene which was monitored by an optical fluorescent detector above the extension block. A disposable microdevice (10 cm diameter) consists of a solid-phase extraction based sample pretreatment unit, bead chamber, and 4 ${\mu}L$ of the PCR chamber as shown Fig. 2. The microchip is fabricated using a patterned polycarbonate (PC) sheet with 1 mm thickness and a PC film with 130 ${\mu}m$ thickness, which layers are thermally bonded at $138^{\circ}C$ using acetone vapour. Silicatreated microglass beads with 150~212 ${\mu}L$ diameter are introduced into the sample pretreatment chambers and held in place by weir structure for construction of solid-phase extraction system. Fig. 3 shows strobed images of sequential loading of three samples. Three samples were loaded into the reservoir simultaneously (Fig. 3A), then the influenza A H3N2 viral RNA sample was loaded at 5000 RPM for 10 sec (Fig. 3B). Washing buffer was followed at 5000 RPM for 5 min (Fig. 3C), and angular frequency was decreased to 100 RPM for siphon priming of PCR cocktail to the channel as shown in Figure 3D. Finally the PCR cocktail was loaded to the bead chamber at 2000 RPM for 10 sec, and then RPM was increased up to 5000 RPM for 1 min to obtain the as much as PCR cocktail containing the RNA template (Fig. 3E). In this system, the wastes from RNA samples and washing buffer were transported to the waste chamber, which is fully filled to the chamber with precise optimization. Then, the PCR cocktail was able to transport to the PCR chamber. Fig. 3F shows the final image of the sample pretreatment. PCR cocktail containing RNA template is successfully isolated from waste. To detect the influenza A H3N2 virus, the purified RNA with PCR cocktail in the PCR chamber was amplified by using performed the RNA capture on the proposed microdevice. The fluorescence images were described in Figure 4A at the 0, 40 cycles. The fluorescence signal (40 cycle) was drastically increased confirming the influenza A H3N2 virus. The real-time profiles were successfully obtained using the optical fluorescence detector as shown in Figure 4B. The Rotary PCR and off-chip PCR were compared with same amount of influenza A H3N2 virus. The Ct value of Rotary PCR was smaller than the off-chip PCR without contamination. The whole process of the sample pretreatment and RT-PCR could be accomplished in 30 min on the fully integrated Rotary Genetic Analyzer system. We have demonstrated a fully integrated and portable Rotary Genetic Analyzer for detection of the gene expression of influenza A virus, which has 'Sample-in-answer-out' capability including sample pretreatment, rotary amplification, and optical detection. Target gene amplification was real-time monitored using the integrated Rotary Genetic Analyzer system.