• Title/Summary/Keyword: Immunomagnetic separation

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Microdevice for Separation of Circulating Tumor Cells Using Embedded Magnetophoresis with V-shaped Ni-Co Nanowires and Immuno-nanomagnetic Beads

  • Park, Jeong Won;Lee, Nae-Rym;Cho, Sung Mok;Jung, Moon Youn;Ihm, Chunhwa;Lee, Dae-Sik
    • ETRI Journal
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    • v.37 no.2
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    • pp.233-240
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    • 2015
  • The novelty of this study resides in a 6"-wafer-level microfabrication protocol for a microdevice with a fluidic control system for the separation of circulating tumor cells (CTCs) from human whole blood cells. The microdevice utilizes a lateral magnetophoresis method based on immunomagnetic nanobeads with anti-epithelial cell adhesive molecule antibodies that selectively bind to epithelial cancer cells. The device consists of a top polydimethylsiloxane substrate for microfluidic control and a bottom substrate for lateral magnetophoretic force generation with embedded v-shaped soft magnetic microwires. The microdevice can isolate about 93% of the spiked cancer cells (MCF-7, a breast cancer cell line) at a flow rate of 40/100 mL/min with respect to a whole human blood/buffer solution. For all isolation, it takes only 10 min to process 400 mL of whole human blood. The fabrication method is sufficiently simple and easy, allowing the microdevice to be a mass-producible clinical tool for cancer diagnosis, prognosis, and personalized medicine.

Synthesis of Model Microspheres and Adsorption Study of Bovine Albumin (모델 Microspheres의 합성 및 Bovine Albumin의 흡착)

  • 박영준;윤정열
    • Journal of Biomedical Engineering Research
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    • v.14 no.3
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    • pp.209-220
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    • 1993
  • Microspheres are expected to be applied to biomedical areas such as solid-phase immunoassays, drug delivery systems, Immunomagnetic cell separation. To synthesize micro-spheres for biomedical application, "two stage shot growth method" was developed. The uniformity ratio or synthesized microspheres was always smatter than 1.05. And the surface charge density (or the number of ionizable functional groups) of the microspheres synthesized by "two stage shot growth method" was 6-13 times higher than thats of the ml crospheres synthesized by conventional seeded batch copolymerization. As a previous step for biomedical application, adsorption experiments of bovine albumin on microspheres were carried out under various conditions. The maximum adsorbed amount was obtained in the neighborhood of pH 4.5. Isoelectric point of bovine albumin Is pH 5.0, so experimental result shows that it shifted to acid area. The adsorption Isotherm was obtained, the plateau region was always reached at 2.Og/L (bulk concentration of bovine albumin ) . The effect of the kind and the amount of surface functional group was also examined.p was also examined.

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초전도 NMR-CT의 영상 원리 및 그 응용

  • 조장희
    • Journal of Biomedical Engineering Research
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    • v.7 no.2
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    • pp.183-190
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    • 1986
  • Microspheres are expected to be applied to biomedical areas such as solid-phase immunoassays, drug delivery systems, immunomagnetic cell separation. To synthesize microspheres for biomedical application, "two stage shot growth method" was developed. The uniformity ratio of synthesized microspheres was always smaller than 1.05. And the surface charge density (or the number of ionizable functional groups) of the microspheres synthesized by "two stage shot growth method" was 6~13 times higher than that of the microspheres synthesized by conventional seeded batch copolymerization. As a previous step for biomedical application, adsorption experiments of bovine albumin on microspheres were carried out under various conditions. The maximum adsorbed amount was obtained in the neighborhood of pH 4.5. Isoelectric point of bovine albumin is pH 5.0, so experimental result shows that it shifted to acid area. The adsorption isotherm was obtained, the plateau region was always reached at 2.Og/L (bulk concentration of bovine albumin).The effect of the kind and the amount of surface functional group was also examined.p was also examined.

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Synthesis of Functional Microspheres and Biomedical Applications (기능성 Microspheres의 합성 및 생의학적 응용)

  • Kim, J.H.;Kim, W.S.
    • Proceedings of the KOSOMBE Conference
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    • v.1993 no.05
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    • pp.7-14
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    • 1993
  • Nowadays, microspheres are expected to be applied to biomedical areas and many studies are being performed. For biomedical applications, many kinds of microspheres were synthesized by emulsion polymerization, emulsifier-free emu]sion polymerization, and emulsifier-free emulsion polymerization with ionic surface-active comonomers. Further synthesis techniques about microencapsulation and magnetic microspheres are introduced. Among the practical applications of microspheres, some interesting subjects are introduced. These include solid-phase immunoassays, labeling and identification of lymphocyte populations, extracorporeal and hemoperfusion systems, drug delivery systems, and immunomagnetic cell separation. In addition, basic theories, problems and research trends are also introduced.

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Distributional Characteristics of Cryptosporidium and Giardia in Raw Water of Large Water Treatment Facility in Korea (국내 5만톤/일 이상 대규모 정수장 원수에서 크립토스포리디움 및 지아디아 분포특성)

  • Park, Sangjung;Kim, Jongmin;Kim, Taeseung;Chung, Dongil;Lee, Mokyoung;Nam, Sehee;Yoon, Jongho;Jung, Jongmoon;Cho, Eunjoo;Jeon, Hyensook;Jang, Eunsook;Chung, Mieun
    • Journal of Korean Society on Water Environment
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    • v.26 no.2
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    • pp.261-267
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    • 2010
  • In order to evaluate the distribution level of Cryptosporidium and Giardia in drinking water resources, distribution of Cryptosporidium and Giardia was studied in intake water of main water treatment facility with treatment capacity over 50,000 ton/day in Korea. 10 L samples from each study sites were collected quarterly for 2 years between Oct. '04 and Dec. '07. Cryptosporidium and Giardia were filtered and concentrated by capsule filter and centrifugation, and analyzed by immunomagnetic separation process and fluorescent assay. Cryptosporidium oocysts and Giardia cysts were detected in 7.0% and 9.3%, respectively, of a total 776 samples from 97 study sites. And mean detection number of Cryptosporidium oocysts and Giardia cysts in total 776 samples was 0.11/10 L, 0.16/10 L, detection range of Cryptosporidium oocysts and Giardia cysts was 0~7/10 L, 0~4/10 L, respectively. In seasonal distribution, Cryptosporidium was more frequent in spring as 9.2% than other season, Giardia was more frequent in winter as 14.6% than other season, but there was not shown significant seasonal characteristics. In correlation analysis with total 776 data, Cryptosporidium had significant correlation to total coliforms at the 0.05 level, but correlation value was too low as 0.07 (r=0.07). In case of Giardia, what had significant correlation at the 0.05 level was total coliforms and fecal coliforms, but correlation value was too low as 0.26, 0.27 respectively.

Improvement of Validity and Efficiency for Detection of Cryptosporidium Ocysts and Giardia Cysts in Environmental Water Samples (환경수 중 크립토스포리디움 오시스트 및 지아디아 시스트 검출의 정확도 및 회수율 향상을 위한 연구)

  • 이목영;조은주;김도연;변승헌;이의광;오세종;안승구
    • Korean Journal of Microbiology
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    • v.39 no.1
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    • pp.27-35
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    • 2003
  • No currently available methods to monitor pathogenic protozoa, Cryptosporidium and Giardia in environmental water come close to acceptable sensitivity, specificity and reproducibility, and so it has to be accompanied by thorough quality control and performance evaluation to credibly predict the distribution of them. We collected surface water samples from the Han River and spiked our prepared (oo)cysts, determined Matrix Spike recoveries using USEPA Method 1623 and considered what factors influence MS recovery and validity. As a result, average 46% of spiked oocysts and 60% of spike cysts were recovered, but repetitive sampling and statistical approach seemed to be necessary to determine the environmental pollution level of two protozoa as their variation coefficients was so much as 35oio and 26%. And MS recoveries with two acid dissociations during immunomagnetic separation were improved more 10% than that with one dissociations and the use of spiked suspension enumerated by flow cytometry instead of manual preparation enhanced the validity and reliability in spiking tests. Because fluorescence characteristics of (oo)cysts stained on well slides with FITC-labeled monoclonal antibodies and DAPI was not always same, well Elides from spiked field samples were helpful to evaluate the performance of staining. We found many (oo)cyst-like objects with typical fluorescence, not (co)cysts, from the Han River water samples, and then it was concluded that nuclei staining by DAPI (4',6-diamidino-2-phenylindole) and examination by Differential Interference Contrast Microscope should be critical for valid identification.

Interlaboratory Study for Cryptosporidium and Giardia Test Methods in Water (정수장 원생동물 분석방법의 국내 실험실간 교차시험 연구)

  • Chung, Hyenmi;Park, Sangjung;Kim, Daekyun;Park, Sanghee;Kim, Reeho;Lee, Sangho;Cho, Joorae;Lee, Kyeunghee;Park, Jonggeun;Lee, Mokyoung;Byun, Seungheon;Cho, Eunju;Rhim, Yeontaek
    • Journal of Korean Society on Water Environment
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    • v.22 no.2
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    • pp.321-327
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    • 2006
  • This interlaboratory study was designed to evaluate protozoan test methods in water and to predict the major causes of deviation of the methods. Each of four laboratories with previous experience of protozoa analysis in water participated, and met the initial performance criteria of EPA 1623 method provided. The protozoan analysis procedure consists of filtrations, concentration, immunomagnetic separation, dyeing (staining) and counting with observation. We tested three different filtration equipments: capsule filter for 10 L of surface water, and high volume (HV) capsule filter and membrane filter for 100 L of finished water. When the recovery of each step of the procedure was evaluated with EasySeed, the commercial stock of each 100 Cryptosporidium and Giardia, immunomagnetic separation and filtration step were the most crucial steps affecting the stability of the recovery, especially for Cryptosporidium. There was no significant difference of recovery among the filtration methods. Recovery of protozoa from source water were evaluated with spiked EasySeed as matrix tests. The recoveries of Giardia increased significantly in the matrix tests compared those in the deionized water. We also applied red stained mixture stocks of Cryptosporidium and Giardia called ColorSeed as internal standards of water sample tests. The recoveries of both EasySeed and ColorSeed in samples tested were within the range of the criteria, however, the Giardia recoveries using ColorSeed decreased significantly. Further optimization study with ColorSeed will be necessary, considering the convenience of using the internal standard without additional sample analysis. The significant factors of the recovery variation were identified as the differences of laboratories as well as water quality and type of the stock for spiking. The results of this study emphasize the importance of the quality assurance program for protozoan analysis lab in water.

Comparison of Methods for Detection of Escherichia coli O157:H7 in Ground Beef and Radish Sprouts

  • Lee, Jae-Hoon;Hyeon, Ji-Yeon;Heo, Seok;Hwang, In-Gyun;Kwak, Hyo-Sun;Choi, In-Soo;Park, Chan-Kyu;Seo, Kun-Ho
    • Food Science of Animal Resources
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    • v.30 no.2
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    • pp.179-184
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    • 2010
  • Escherichia coli O157:H7 is a food-borne pathogen that causes bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome (HUS). We compared three selective media and evaluated the performance of immunomagnetic separation (IMS) for the detection of low levels of E. coli O157:H7 in ground beef and radish sprouts with different levels of background flora. Bulk food samples (500 g for each trial) were artificially inoculated with nalidixic acid-resistant E. coli O157:H7 at the lowest dose that would generate 20 partial-positive samples of 25 g each. All samples were homogenized in mTSB (225 mL) and incubated overnight at $37^{\circ}C$. IMS was performed using the enriched mTSB samples (1 mL) along with conventional spreads plated onto three different selective media: Sorbitol MacConkey agar (SMAC), Sorbitol MacConkey agar with cefixime and tellulite (CT-SMAC), and Sorbitol MacConkey agar with nalidixic acid (NAL-SMAC) as the gold standard. Two suspicious colonies from each medium were selected and confirmed usinga serological test after transfer to tryptic soy broth with yeast extract (TSAYE). CT-SMAC was better than SMAC for detecting E. coli O157:H7 in all food types. Although there was no statistical difference in the number of positive samples when using IMS vs. non-IMS techniques, more positive samples were detected when IMS was used in both ground beef and radish sprouts. It appears that the improvement was more significant in radish sprouts, which had a higher level of background flora than ground beef. The results also suggest that the combination of CT-SMAC and IMS is sufficient to recover low levels of E. coli O157:H7 in high background flora food samples.

표면특성이 제어된 기능성 나노 입자의 전자 및 의공학적 응용

  • 박영준;이준영;김중현
    • Proceedings of the Korea Crystallographic Association Conference
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    • 2002.11a
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    • pp.54-55
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    • 2002
  • The fabrication, characterization and manipulation of nanoparticle system brings together physics, chemistry, materials science and biology in an unprecedented way. Phenomena occurring in such systems are fundamental to the workings of electronic devices, but also to living organisms. The ability to fabricate the surface of nanoparticles Is essential in the further development of functional devices that incorporate nanoscale features. Even more essential is the ability to introduce a wide range of chemical and materials flexibility into these structures to build up more complex nanostructures that can ultimately rival biological nanosystems. In this respect, polymers are potentially ideal nanoscale building blocks because of their length scale, well-defined architecture, controlled synthesis, ease of processing and wide range of chemical functionality that can be incorporated. In this presentation, we will look at a number of promising polymer-based nanoparticle fabrication strategies that have been developed recently, with an emphasis on those techniques that incorporate nanostructured polymeric particles into electronic devices or biomedical applications. And functional nanoparticles deliberately designed using several powerful process methods and their application will be discussed. Nanostructured nanoparticles, what we called, implies dispersed colloids with the size ranged from several nanometers to hundreds of nanometer. They have extremely large surface area, thus it is very important to control the morphology or surface functionality fitted for adequate objectives and properties. Their properties should be controlled for various kind of bio-related technologies, such as immunomagnetic cell separation, drug delivery systems, labeling and identification of lymphocyte populations, extracorporeal and hemoperfusion systems, etc. Well-defined polymeric nanoparticles can be considered as smart bomb or MEMS.

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Detection of Campylobacter jejuni in food and poultry visors using immunomagnetic separation and microtitre hybridization

  • Simard, Ronald-E.
    • Proceedings of the Korean Society of Fisheries Technology Conference
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    • 2000.05a
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    • pp.71-73
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    • 2000
  • Campylobacter jejuni is most frequently identified cause of cause of acute diarrhoeal infections in developeed countries, exceeding rates of illness caused by both salmonella and shigilla(Skirrow, 1990 ; Lior 1994). Previous studies on campylobacter jejuni contamination of commercial broiler carcasses in u.s.(Stern, 1992). Most cases of the disease result from indirect transmission of Campylobactor from animals via milk, water and meat. In addition to Campylobactor jejuni. the closely relates species Campylobactor coli and Campylobactor lari have also been implicated as agents of gastroenteritis in humans. Campylobactor coli represented only approximately 3% of the Campylobactor isolates from patients with Campylobactor enteritis(Griffiths and Park, 1990) whereas Campylobactor coli is mainly isolated from pork(Lmmerding et al., 1988). Campylobactor jejuni has also been isolated from cases of bacteremia, appendicitis and, recently, has been associated with Guillai-Barre syndrome(Allos and Blaser, 1994; von Wulffen et al., 1994; Phillips, 1995). Studies in volunteers indicated that the infectious dose for Campylobactor jejuni is low(about 500 organisms)(Robinson, 1981). The methods traditionally used to detect Campylobactor ssp. in food require at least two days of incubation in an enrichment broth followed by plating and two days of incubation on complex culture media containing many antibiotics(Goossens and Butzler, 1992). Finnaly, several biochemical tests must be done to confirm the indentification at the species level. Therfore, sensitive and specific methods for the detection of small numbers of Campylobactor cells in food are needed. Polymerase chain reaction(PCR) assays targeting specific DNA sequences have been developed for the detection of Campylobactor(Giesendorf and Quint, 1995; Hemandex et al., 1995; Winter and Slavidk, 1995). In most cases, a short enrichment step is needed to enhance the sensitivity of the assay prior to detection by PCR as the number of bacteria in the food products is low in comparison with those found in dinical samples, and because the complex composition of food matrices can hinder the PCR and lower its sensitivity. However, these PCR systems are technically demanding to carry out and cumbersome when processing a large number of samples simutaneously. In this paper, an immunomagnetic method to concentrate Campylobactor cells present in food or clinical samples after an enrichment step is described. To detect specifically the thermophilic Campylobactor. a monoclonal antibody was adsorbed on the surface of the magnetic beads which react against a major porin of 45kDa present on the surface of the cells(Huyer et al., 1986). After this partial purification and concentration step, detection of bound cells was achieved using a simple, inexpensive microtitre plate-based hybridization system. We examined two alternative detection systems, one specific for thermophilic Campylobactor based on the detection of 23S rRNA using an immobilized DNA probe. The second system is less specific but more sensitive because of the high copy number of the rRNA present in bacterial cell($10^3-10^4$). By using specific immunomagnetic beads against thermophilic Campylobactor, it was possible to concentrate these cells from a heterogeneous media and obtain highly specific hybridization reactions with good sensitivity. There are several advantages in using microtitre plates instead of filter membranes or other matrices for hybridization techniques. Microtitre plates are much easier to handle than filter membranes during the adsorption, washing, hybridization and detection steps, and their use faciilitates the simultanuous analysis of multiple sample. Here we report on the use of a very simple detection procedure based on a monoclonal anti-RNA-DNA hybrid antibody(Fliss et al., 1999) for detection of the RNA-DNA hybrids formed in the wells.

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