• JSTS:Journal of Semiconductor Technology and Science
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• v.7 no.3
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• pp.140-150
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• 2007

A simple, label-free microfluidic cell purification method is presented for separation of cancer cells by exploiting difference in cell adhesion. To maximize the adhesion difference, three types of polymeric nanostructures (50nm pillars, 50nm perpendicular and 50nm parallel lines with respect to the direction of flow) were fabricated using UV-assisted capillary moulding and included inside a polydimethylsiloxane (PDMS) microfluidic channel bonded onto glass substrate. The adhesion force of human breast epithelial cells (MCF10A) and human breast carcinoma (MCF7) was measured independently by injecting each cell line into the microfluidic device followed by culture for a period of time (e.g., one, two, and three hours). Then, the cells bound to the floor of a microfluidic channel were detached by increasing the flow rate of medium in a stepwise fashion. It was found that the adhesion force of MCF10A was always higher than that of MCF cells regardless of culture time and surface nanotopography at all flow rates, resulting in a label-free detection and separation of cancer cells. For the cell types used in our study, the optimum separation was found for 2 hours culture on 50nm parallel line pattern followed by flow-induced detachment at a flow rate of $300{\mu}l/min$.

• KSBB Journal
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• v.15 no.4
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• pp.342-345
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• 2000

A novel pre-purification method was developed for producing peroxidase to guarantee high purity and yield from plant cell cultures in large-scale process. This method was a simple and efficient procedure for the isolation and pre-purification of peroxidase from the biomass consisting of active clay treatment followed by cationic exchange chromatography. The use of active clay in the pre-purification process allows for rapid and efficient separation of peroxidase from interfering compounds and dramatically increases yield and purity of crude peroxidase for purification steps compared to alternative processes. This pre-purification process serves to minimize the buffer usage size and complexity of the HPLC operations for peroxidase purification. This process is readily scalable to a pilot plant and eventually to a production environment where mass production of material are expected to be produced.

• KSBB Journal
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• v.21 no.1 s.96
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• pp.1-10
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• 2006

The recovery and purification of a paclitaxel from plant cell cultures is essential to commercial process. This review describes a large-scale recovery and purification method for producing paclitaxel, to guarantee high purity and yield from plant cell cultures. Also, the process of separation and purification is optimized in conjunction with a extraction step, pre-purification, purification, and polishing (drying) as an integrated process to meet final product quality requirements such as purity, residual solvents, product morphologies, impurities, bacterial endotoxin, etc. This information is very useful for production and quality control of pharmaceuticals in commercialization step.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2013.10a
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• pp.943-945
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• 2013

The study of Schwann cell myelination has been facilitated by the availability to isolate and establish pure population of primary Schwann cells. Dorsal root ganglia (DRG) of mouse embryo as source of Schwann cells were used in this study. This method includes three steps: first step of dissociation of the embryonic DRG, second step of expansion of Schwann cell precursors, followed by mechanical separation of the Schwann cell-neuronal network from the underlying fibroblasts, and third step of purification of Schwann cells from the associated neurons and subsequent expansion of the purified Schwann cells. We made a highly purified population of Schwann cells and Schwann cell-neuron networks in a short period using this procedure.

• Proceedings of the Korean Vacuum Society Conference
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• 2013.08a
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• pp.276-276
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• 2013

Here we report an integrated microdevice consisting of an efficient passive mixer, a magnetic separation chamber, and a capillary electrophoretic microchannel in which DNA barcode assay, target pathogen separation, and barcode DNA capillary electrophoretic analysis were performed sequentially within 30 min for multiplex pathogen detection at the single-cell level. The intestine-shaped serpentine 3D micromixer provides a high mixing rate to generate magnetic particle-pathogenic bacteria-DNA barcode labelled AuNP complexes quantitatively. After magnetic separation and purification of those complexes, the barcode DNA strands were released and analyzed by the microfluidic capillary electrophoresis within 5 min. The size of the barcode DNA strand was controlled depending on the target bacteria (Staphylococcus aureus, Escherichia coli O157:H7, and Salmonella typhimurium), and the different elution time of the barcode DNA peak in the electropherogram allows us to recognize the target pathogen with ease in the monoplex as well as in the multiplex analysis. In addition, the quantity of the DNA barcode strand (~104) per AuNP is enough to be observed in the laser-induced confocal fluorescence detector, thereby making single-cell analysis possible. This novel integrated microdevice enables us to perform rapid, sensitive, and multiplex pathogen detection with sample-in-answer-out capability to be applied for biosafety testing, environmental screening, and clinical trials.

• KSBB Journal
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• v.15 no.5
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• pp.537-540
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• 2000

In developing a HPLC purification process, it was hoped that a single chromatographic system would be sufficient to abtain pure paclitaxel in high yield. However, no such system was found, due in part to the complex taxoid profile of crude paclitaxel and to the rigorous nature of the product specification. A two step HPLC purification was adopted using reverse-phase separation on C(sub)18 as a first step, and normal-phase separation on silica as the final polishing step. Impurity profiles were established and maintained for paclitaxel, which identified and quantified each impurity observed in purified paclitaxel from these two steps, all impurities at or above 0.1% were identified. Results provide information for improving the quality control of paclitaxel production.

• Proceedings of the Korean Institute of Resources Recycling Conference
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• 2003.10a
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• pp.196-200
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• 2003

A novel prepurification method was developed aiming at increasing yield and purity, also reducing solvent usage for purification of paclitaxel. The use of a micelle and precipitation in the prepurification process allows for rapid separation of paclitaxel from interfering compounds and dramatically reduces solvent usage compared to alternative methodologies. The prepurification process serves to minimize the size and complexity of the HPLC operations for paclitaxel purification. The process is readily scalable to a pilot plant and eventually to a production environment where multikilogram quantities of material are expected to be produced. As much as possible, the process has been optimized to minimize solvent usage, complexity, and operating costs.

• KSBB Journal
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• v.30 no.3
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• pp.103-108
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• 2015

Production of foreign proteins by transgenic plant cell cultures has several advantages such as post-translational modification, low risk of product contamination and low-cost production and purification. However, target proteins are degraded by extracellular proteases existing in the media. A solution to this problem is the use of perfusion culture and ion exchange chromatography for the application of integrated bioprocess using in situ recovery. With this method, production of human granulocyte-macrophage colony-stimulating factor (hGM-CSF) was investigated in this study. First, optimization of cell concentration during the induction phase for the production of hGM-CSF was examined. As cell concentration increased, the level of hGM-CSF was decreased due to the presence of extracellular proteases. Induction using sugarfree media produced 33% more hGM-CSF. The effects of pH on the binding of hGM-CSF to cationic and anionic exchange resins were also investigated. In terms of stability, optimal pH was found to be 5~7. In the case of using buffer exchange when CM-Sepharose was used as a cationic exchange resin, optimal pH for binding was 4.8 and adsorption yield was 77%. When DEAE-Sepharose was used as an anionic exchange resin, it was 5.5 (74%). Without buffer exchange, optimal pH was 4.6 (84%). From these results, an integrated bioprocess using in situ recovery with simultaneous production and separation of foreign protein in transgenic plant cell suspension cultures was found to be feasible.

• Journal of Applied Biological Chemistry
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• v.43 no.3
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• pp.176-179
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• 2000

Nonpolar taxoides extracted from a large-scale cell culture of Taxus chinensis were isolated through the normal and reverse phase column chromatographies, and their compounds were identified via NMR spectroscopy. The complete separation method was systematically established and described. In dichloromethane, dissolved paclitaxel and other taxoids with hexane were precipitated during the purification of paclitaxel from the plant cell culture of T. chinensis through a large-scale process while the relatively nonpolar taxane derivatives remained dissolved in the hexane phase. 13-Deoxy baccatin III (I), baccatin VI (II), taxchinin I (III), $2{\alpha}$, $5{\alpha}$, $10{\beta}$, $14{\beta}$-tetraacetoxy-4(20), 11-taxadiene(IV), 1-deoxy baccatinVI(V), and taxayuntin C (VI) were isolated through column chromatography and identified via NMR spectroscopy. Compounds I and IV were found to the major components, aside from paclitaxel, in the plant cell culture of T. chinensis. The concentrations of I and IV were compared with the that concentration of the paclitaxel in each of plant cell culture. The possible applications of compounds I, II, IV, and V were discussed.

• 한국생물공학회:학술대회논문집
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• 2000.11a
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• pp.260-263
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• 2000

Membranes are widely used to separate target solute molecules such as proteins on the basis of their size in cell broth mixture to minimize the loss of target compounds. In this study, membrane separation system using ultrafilters of MWCO 100 K and 50 K, was operated for concentration and purification of microbial transglutaminase. Fermentation broth containing MTGase was prefiltered by using pore size 1.6 and $0.7\;{\mu}m$ pre-filter made of cellulose fiber and $0.45\;{\mu}m$ microfilter made of cellulose acetate. The prefiltered solution was concentrated by 100 K and 50 K ultrafilter. The final enzyme concentration was 1.29 units/ml and enzyme specific activity was 0.2 units/mg protein. This specific activity were 3.7 times higher than that of initial cell broth mixture. Membrane separation process of microfiltration and ultrafiltration was proved to be very economic, energy efficient and effective separation method used to concentrate MTGase.