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Characterization and Purification of the Bacteriocin Produced by Bacillus licheniformis Isolated from Soybean Sauce (간장에서 분리한 Bacillus licheniformis가 생산하는 박테리오신의 특성 및 정제)

  • Jung, Sung-Sub;Choi, Jung-I;Joo, Woo-Hong;Suh, Hyun-Hyo;Na, Ae-Sil;Cho, Yong-Kweon;Moon, Ja-Young;Ha, Kwon-Chul;Paik, Do-Hyeon;Kang, Dae-Ook
    • Journal of Life Science
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    • v.19 no.7
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    • pp.994-1002
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    • 2009
  • A bacteriocin-producing bacterium identified as Bacillus licheniformis was isolated from soybean sauce. Antibacterial activity was confirmed by paper disc diffusion method, using Micrococcus luteus as a test organism. The bacteriocin also showed antibacterial activities against Bacillus sphaericus, Lactobacillus bulgaricus, Lactobacillus planiarum, Paenibacillus polymyxa, and Pediococcus dextrinicus. Optimal culture conditions for the production of bacteriocin was attained by growing the cells in an MRS medium at a pH of 6.5~ 7.0 and a temperature of 37$^\circ$C for 36$\sim$48 hr. Solvents such as chloroform, ethanol, acetone, and acetonitrile had little effect on bacteriocin activity. However, about 50% of bacteriocin activity diminished with treatment of methanol and isopropanol at the final concentration of 50% at 25$^\circ$C for 1 hr. It was stable against a pH variation range from 3.0 and 7.0, but the activity reduced to 50% at a pH range from 9.0 to 11.0. It's activity was not affected by heat treatment at 100$^\circ$C for 30 min and 50% of activity was retained after heat treatment at 100$^\circ$C for 60 min, showing high thermostability. The bacteriocin was purified to a homogeneity through ammonium sulfate precipitation, SP-Sepharose ion-exchange chromatography, and reverse-phase high-performance liquid chromatography (HPLC). The entire purification protocol led to a 75-fold increase in specific activity and a 13.5% yield of bacteriocin activity. The molecular weight of purified bacteriocin was estimated to be about 2.5 kDa by tricine-SDS-PAGE.

ω3-Polyunsaturated Fatty Acids-induced Inhibition of Tumorigenicity and Invasion by Suppression of COX-2/MMPs/VEGF through NF-kB in Colon Cancer Cells (오메가-3 지방산에 의한 COX-2/MMPs/VEGF 억제에 따른 대장암세포의 종양 형성 및 침윤 억제)

  • Shin, Soyeon;Kim, Yong-Jo;Han, Seung-Hyeon;Silwal, Prashanta;Heo, Jun-Young;Jeon, Young-Joo;Park, Seung-Kiel;Kweon, Gi-Ryang;Park, Jong-Il;Lim, Kyu
    • Journal of Life Science
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    • v.27 no.9
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    • pp.1020-1030
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    • 2017
  • Epidemiology studies have reported a reduced incidence of colon cancer among populations that consume a large quantity of ${\omega}3-polyunsaturated$ fatty acids (${\omega}3-PUFAs$) of marine origin. Herein, we demonstrated a mechanism of anticancer action of ${\omega}3-PUFAs$, showing that they suppressed invasion and tumorigenicity in colon cancer cells. Docosahexaenoic acids (DHA) inhibited the cell growth of HT29 cells. This action likely involved apoptosis, given that the DHA treatment increased the cleaved form of PARP and sub G1 cells. Moreover, the invasiveness of HT29 cells was inhibited following DHA treatment, whereas arachidonic acid (AA) had no effect. The levels of Matrix-metalloproteinase-9 (MMP-9) and MMP-2 mRNA decreased after DHA pretreatment. DHA treatment inhibited MMP-9 and MMP-2 promoter activities and reduced VEGF promoter activity. DHA pretreatment also inhibited the activities of prostaglandin-2 (PGE2)-induced MMPs and the VEGF promoter. Cyclooxygenase-2 (COX-2) overexpression increased the activity of MMPs and that of the Vascular endotherial growth factor (VEGF) promoter in HT29 cells, and DHA inhibited NF-kB and COX-2 promoter reporter activities. As shown by in vivo experiments, when mouse colon cancer cells (MCA38) were implanted into Fat-1 and wild-type mice, both the tumoral size and volume were dramatically inhibited in Fat-1 transgenic mice. Furthermore, TUNEL-positive cells increased in tumors from Fat-1 mice compared with wild mice. In immunohistochemistry, the intensity of CD31 in Fat-1 tumors was weaker. These findings suggest that ${\omega}3-PUFAs$ may inhibit tumorigenicity and angiogenesis as well as cancer cell invasion by suppression of COX-2, MMPs and VEGF via the reduction of NF-kB in colon cancer.

Perfluoropolymer Membranes of Tetrafluoroethylene and 2,2,4Trifluofo- 5Trifluorometoxy- 1,3Dioxole.

  • Arcella, V.;Colaianna, P.;Brinati, G.;Gordano, A.;Clarizia, G.;Tocci, E.;Drioli, E.
    • Proceedings of the Membrane Society of Korea Conference
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    • 1999.07a
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    • pp.39-42
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    • 1999
  • Perfluoropolymers represent the ultimate resistance to hostile chemical environments and high service temperature, attributed to the presence of fluorine in the polymer backbone, i.e. to the high bond energy of C-F and C-C bonds of fluorocarbons. Copolymers of Tetrafluoroethylene (TEE) and 2, 2, 4Trifluoro-5Trifluorometoxy- 1, 3Dioxole (TTD), commercially known as HYFLON AD, are amorphous perfluoropolymers with glass transition temperature (Tg)higher than room temperature, showing a thermal decomposition temperature exceeding 40$0^{\circ}C$. These polymer systems are highly soluble in fluorinated solvents, with low solution viscosities. This property allows the preparation of self-supported and composite membranes with desired membrane thickness. Symmetric and asymmetric perfluoropolymer membranes, made with HYFLON AD, have been prepared and evaluated. Porous and not porous symmetric membranes have been obtained by solvent evaporation with various processing conditions. Asymmetric membranes have been prepared by th wet phase inversion method. Measure of contact angle to distilled water have been carried out. Figure 1 compares experimental results with those of other commercial membranes. Contact angles of about 120$^{\circ}$for our amorphous perfluoropolymer membranes demonstrate that they posses a high hydrophobic character. Measure of contact angles to hexandecane have been also carried out to evaluate the organophobic character. Rsults are reported in Figure 2. The observed strong organophobicity leads to excellent fouling resistance and inertness. Porous membranes with pore size between 30 and 80 nanometers have shown no permeation to water at pressures as high as 10 bars. However high permeation to gases, such as O2, N2 and CO2, and no selectivities were observed. Considering the porous structure of the membrane, this behavior was expected. In consideration of the above properties, possible useful uses in th field of gas- liquid separations are envisaged for these membranes. A particularly promising application is in the field of membrane contactors, equipments in which membranes are used to improve mass transfer coefficients in respect to traditional extraction and absorption processes. Gas permeation properties have been evaluated for asymmetric membranes and composite symmetric ones. Experimental permselectivity values, obtained at different pressure differences, to various single gases are reported in Tab. 1, 2 and 3. Experimental data have been compared with literature data obtained with membranes made with different amorphous perfluoropolymer systems, such as copolymers of Perfluoro2, 2dimethyl dioxole (PDD) and Tetrafluorethylene, commercialized by the Du Pont Company with the trade name of Teflon AF. An interesting linear relationship between permeability and the glass transition temperature of the polymer constituting the membrane has been observed. Results are descussed in terms of polymer chain structure, which affects the presence of voids at molecular scale and their size distribution. Molecular Dyanmics studies are in progress in order to support the understanding of these results. A modified Theodoru- Suter method provided by the Amorphous Cell module of InsightII/Discover was used to determine the chain packing. A completely amorphous polymer box of about 3.5 nm was considered. Last but not least the use of amorphous perfluoropolymer membranes appears to be ideal when separation processes have to be performed in hostile environments, i.e. high temperatures and aggressive non-aqueous media, such as chemicals and solvents. In these cases Hyflon AD membranes can exploit the outstanding resistance of perfluoropolymers.

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Deposition of Poly-$Si_{1-x}Ge_x$ Thin Film by RTCVD (RTCVD에 의한 다결정 $Si_{1-x}Ge_x$ 박막 증착)

  • Kim, Jae-Jung;Lee, Seung-Ho;So, Myeong-Gi
    • Korean Journal of Materials Research
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    • v.5 no.6
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    • pp.690-698
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    • 1995
  • The Poly-S $i_{1-x}$G $e_{x}$ thin films were deposited on oxidized Si wafer by RTCVD(rapid thermal chemical vapor deposition) using Si $H_4$and Ge $H_4$, at 450 ~5$50^{\circ}C$. The variation of Ge mole fraction and the deposition rate of S $i_{1-x}$G $e_{x}$ thin film were studied as a function of the deposition temperature and the Ge $H_4$/Si $H_4$input ratio, and the crystal phase and the surface roughness were studied by XRD and AFM(atomic force microscopy), respectively. The experimental results showed that the activation energy for the deposition of poly-S $i_{1-x}$G $e_{x}$ was about 32~37Kca /mol and the deposition rate of S $i_{1-x}$G $e_{x}$ thin films was increased with increasing the deposition temperature and the input ratio. From the analysis of composition, it was known that the Ge mole fraction within the poly-S $i_{1-x}$G $e_{x}$ thin film was decreased with decreasing the input ratio and increasing the deposition temperature. As-deposited S $i_{1-x}$G $e_{x}$ thin films were polycrystalline over the entire experimental range. But those were amorphous at the deposition temperature of 450, 475$^{\circ}C$ and the input ratio of 0.05. By adding the Ge $H_4$, poly-S $i_{1-x}$G $e_{x}$ thin film were deposited at relatively lower deposition temperatures($\leq$ 5$50^{\circ}C$) than those of conventional poly-Si(>$600^{\circ}C$). From surface roughness measurement of poly-S $i_{1-x}$G $e_{x}$ it was found that the surface roughness( $R_{i}$ ) increased with increasing the deposition temperature and input ratio.and input ratio.

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Enhanced Production of Oleanolic Acid by the Elicitation in Oldenlandia diffusa Suspension Cell Cultures (백화사설초의 현탁세포배양에서 Elicitation에 의한 Oleanolic acid 생산성 증대)

  • Lee Yong-Il;Kim Dong-Il
    • KSBB Journal
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    • v.19 no.6 s.89
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    • pp.471-477
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    • 2004
  • Oldenlandia diffusa is a Chinese medicinal herb with antitumor activity capable of suppressing the growth of some cancer cell lines. Oleanolic acid and ursolic acid are triterpenoid compounds that exist in Oldenlandia diffusa. Recently, these have been noted for anti-inflammatory, anti-cancer, and hepato-protective effects. Application of both plant growth regulators, 2,4-D and kinetin, was found to be essential for the initiation of callus and suspension cells. Leaf blades of Oldenlandia diffusa was transformed into callus on Schenk and Hildebrandt medium supplemented with 0.5 mg/L 2,4-D and 0.1 mg/L kinetin, while optimum initiation condition for suspension cells of Oldenlandia diffusa was determined to be 0.75 mg/L 2,4-D and 0.1 mg/L kinetin. Chromatographic separation of oleanolic acid from its derivatives was achieved using Rexchrom S5-100-ODS column. Analytical conditions for oleanolic acid were determined as follows: flow rate at 1.0 mL/min, UV length at 200 nm and mobile phase of $80\%$ acetonitrile and $20\%$ water. Production of secondary metabolites was found to be increased by the treatment with elicitors or signal transducers. The maximum production of oleanolic acid was 99.6 mg/L in cultures with 0.5 mM salicylic acid. It is 1.74 times higher than that of control.

Effect of Dandelion Juice Supplementation on Alcohol-Induced Oxidative Stress and Hangover in Healthy Male College Students (건강한 남자 대학생에서 민들레즙 보충이 알코올에 의한 산화적 스트레스 및 숙취에 미치는 효과)

  • Noh, Kyung-Hee;Jang, Ji-Hyun;Kim, Jin-Ju;Shin, Jin-Hyuk;Kim, Dong-Kyoo;Song, Young-Sun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.38 no.6
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    • pp.683-693
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    • 2009
  • This study was designed to investigate the effect of dandelion juice supplementation on attenuation of oxidative stress and hangover after drinking alcohol in healthy college male students. This human trial was conducted by two phase cross over design with two weeks wash out period. The subjects (age $24{\sim}28$ years) were volunteers who had more than 72 g of ethanol drinking capacity. Dandelion group was given dandelion juice 220 mL daily for 7 days. Biochemical markers were determined in blood samples taken at 0 and 150 minutes after administration 72 g of alcohol. The levels of plasma glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, lactate dehydrogenase and bilirubin, the indicators of liver cell damage, were not significantly different between groups. No significant differences in lymphocyte DNA damage level between groups was observed. However, plasma acetaldehyde dehydrogenase (ALDH) and high density lipoprotein cholesterol levels were significantly (p<0.01) increased in dandelion supplemented group compared to that of control group. Furthermore, activities and protein expressions of glutathione-reductase and catalase of erythrocytes were significantly elevated in dandelion supplemented group compared to that of control group. From the above results, it is concluded that dandelion juice supplementation can reduce oxidative stress and hangover syndrome through the elevation of ALDH and antioxidative enzyme system in healthy male adults.

Inhibitory Effects of Ethanolic Extracts from Aster glehni on Xanthine Oxidase and Content Determination of Bioactive Components Using HPLC-UV (섬쑥부쟁이 에탄올 추출물의 잔틴산화효소 저해 효능 및 HPLC-UV를 이용한 유효성분의 함량 분석)

  • Kang, Dong Hyeon;Han, Eun Hye;Jin, Changbae;Kim, Hyoung Ja
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.45 no.11
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    • pp.1610-1616
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    • 2016
  • This study aimed to establish an optimal extraction process and high performance liquid chromatography-ultraviolet (HPLC-UV) analytical method for determination of 3,5-dicaffeoylquinic acid (3,5-DCQA) as a part of materials standardization for the development of a xanthine oxidase inhibitor as a health functional food. The quantitative determination method of 3,5-DCQA as a marker compound was optimized by HPLC analysis using a Luna RP-18 column, and the correlation coefficient for the calibration curve showed good linearity of more than 0.9999 using a gradient eluent of water (1% acetic acid) and methanol as the mobile phase at a flow rate of 1.0 mL/min and a detection wavelength of 320 nm. The HPLC-UV method was applied successfully to quantification of the marker compound (3,5-DCQA) in Aster glehni extracts after validation of the method with linearity, accuracy, and precision. Ethanolic extracts of A. glehni (AGEs) were evaluated by reflux extraction at 70 and $80^{\circ}C$ with 30, 50, 70, and 80% ethanol for 3, 4, 5, and 6 h, respectively. Among AGEs, 70% AGE at $70^{\circ}C$ showed the highest content of 3,5-DCQA of $52.59{\pm}3.45mg/100g$ A. glehni. Furthermore, AGEs were analyzed for their inhibitory activities on uric acid production by the xanthine/xanthine oxidase system. The 70% AGE at $70^{\circ}C$ showed the most potent inhibitory activity with $IC_{50}$ values of $77.01{\pm}3.13{\sim}89.96{\pm}3.08{\mu}g/mL$. The results suggest that standardization of 3,5-DCQA in AGEs using HPLC-UV analysis would be an acceptable method for the development of health functional foods.

Application of the Artificial Mussel for Monitoring Heavy Metal Levels in Seawater of the Coastal Environments, Korea (Artificial mussel을 이용한 우리나라 연안환경의 중금속 오염도 연구)

  • Ra, Kongtae;Kim, Joung-Keun;Kim, Kyung-Tae;Lee, Seung-Yong;Kim, Eun-Soo;Lee, Jung-Moo;Wu, Rudolf S.S.
    • Journal of the Korean Society for Marine Environment & Energy
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    • v.17 no.2
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    • pp.131-145
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    • 2014
  • The new passive sampler called "artificial mussel (AM)" offers a potential device to study the spatiotemporal changes of metal concentrations in different marine environment worldwide. The purpose of this study is to characterize metal (Cd, Cr, Cu, Zn, Pb) accumulation on the AM and transplanted mussel (Mytilus edulis) at 5 sites of Lake Shihwa. Both the AMs and mussels showed increasing concentrations of all five metals during the 12 weeks exposure period. Higher concentrations of Zn were showed in both the AMs and Mytilus edulis relative to other metals. The AMs accumulated higher concentrations of Cd, Cr and Zn, but they presented lower levels of Cu and Pb than Mytilus edulis. The correlations for Cd, Cu and Pb were statistically significant between the AMs and Mytilus edulis, indicating that the accumulation patterns for those metals were similar. However, no similarities for Cr and Zn were observed between two monitoring devices across all of the sites in Shihwa Lake. According to relationship for metal concentrations between dissolve phase in seawater and both the AMs and Mytilus edulis, the AMs for Cd, Cu and Zn represent more metal contamination than Mytilus edulis. Our results indicated that the AMs give a better resolution to reveal the spatial differences in dissolved metal concentration. This study suggests that the AMs can provide a time-integrated estimate of metal pollution in marine environments as well as freshwater environments of Korea.

Study for the Synthesis of $[^{123}I]$Idoxifene and Its Uptake in the Breast Cancer Cell ($[^{123}I]$Idoxifene 합성과 유방암의 세포섭취에 관한 연구)

  • Cho, Young-Sub;Yang, Seung-Dae;Suh, Yong-Sup;Chun, Kwon-Soo;Ahn, Soon-Hyuk;Lim, Soo-Jung;Lim, Sang-Moo;Yu, Kook-Hyun
    • The Korean Journal of Nuclear Medicine
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    • v.34 no.5
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    • pp.410-417
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    • 2000
  • Purpose: Idoxifene is currently entering phase II clinical trials for the treatment of advanced breast cancer. The radiolabeled idoxifene using $[^{123}I]$ provides an opportunity for clinical pharmacology with single photon emission computed tomography (SPECT). The purpose of this study was to prepare radiolabeled idoxifene using $[^{123}I]$ and to determine its cell uptake of breast cancer cell line. Materials and Methods: With a view to evaluating new anticancer drugs, we are investigating the novel antiestrogen pyrrolidino-4-iodotamoxifen (idoxifene). $[^{123}I]$Idoxifene has been prepared in no-carrier-added form using a tributyl stannylated precursor which has been synthesized by means of (2-chloroethoxy)benzene with (${\pm}$)-2-phenylbutanoic acid on the basis of previously reported standard methods. The biodistribution and dynamic behavior of the compound were investigated using the comparative breast cancer cell line, MCF-7 (estrogen receptor-positive) and MDA-MB-468 (non-estrogen receptor). Results and Conclusion: Acylation of (2-chloroethoxy)benzene with (${\pm}$)-2-phenylbutanoic acid gave the versatile ketone (81%) which reacted with 1,4-diiodobenzene to give triphenylethylene as a mixture of E and Z geometric isomers, which were separated by the recrystallization in ethanol. The E-isomer was treated with pyrrolidine to give idoxifene (67%). In order to incorporate radioactive iodine into the 4-position, the 4-stannylated precursor was prepared (30%). The yield of radioiodination was 90-92% with a high radiochemical purity greater than 98%. The ratio of tumor uptake of the breast cancer cell line between MCF-7 and MDA-MB-468 was about 1.7.

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Radioimmunoassay for Determination of Serum Macrophage Migration Inhibitory Factor (혈중 대식세포 유주 저지 인자 측정을 위한 방사면역측정법)

  • Lee, Tae-Sup;Shin, Seok-Hwan;Song, Jee-In;Woo, Kwang-Sun;Chung, Wee-Sup;Choi, Chang-Woon;Lim, Sang-Moo
    • The Korean Journal of Nuclear Medicine
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    • v.38 no.6
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    • pp.532-539
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    • 2004
  • Purpose: There has been a renewal of interest in Macrophage migration inhibitory factor (MIF), especially correlation in pathogenesis of sepsis by many infectious diseases and in regulation of host inflammatory and immune response. We developed immunoradiometric assay (IRMA) to determine serum human MIF concentration. Materials and Methods: The IRMA system utilizes solid phase bound monoclonal anti-recombinant human MIF (rhMIF) antibody as a capture antibody, biotinylated polyclonal anti-rhMIF antibody as a detector antibody. We applied with rhMIF that concentration of standard solutions increased from 0 ng/ml to 100 ng/ml. We used $^{125}I$-streptavidin (SA) as radiotracer to determination of rhMIF concentration. Streptavidin was labeled with $^{125}I$ by Chloramine-T method and $^{125}I$-SA was purified by ultracentrifugation. $^{125}I$-SA stability was evaluated by ITLC analysis at $4^{\circ}C$ and room temperatures until 60days. To validate IRMA system for MIF, we experimented intra-assay and inter-assay coefficients of variation, recovery test and dilution test. Results: Radiolabeling yield of $^{125}I$-SA was 87% and purified $^{125}I$-SA retained above 99% radiochemical purity. $^{125}I$-SA showed above 93% stability in $4^{\circ}C$ until 60days that it is good for immunoradiometric assay as radiotracer. Plotted standard dose response curve showed that increased concentration of rhMIF linearly correlated (R2=0.99) with bound radioactivity of $^{125}I$-SA. The highest intra- and inter-assay coefficients of variation were 5.5% and 7.6%, respectively. The average of recovery of MIF in samples was 102%. In dilution test, linear response curves were obtained (R2=0.97). Conclusion: Radioimmunoassay using $^{125}I$-SA as radiotracer thought to be useful for the determination of serum MIF concentration, and further, its data will be used to evaluate the correlation between clinical significance and serum MIF concentration in patients with various inflammatory diseases.