• 약학회지
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• 제50권3호
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• pp.191-198
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• 2006

Acanthopanax species have traditionally been used as a tonic, a sedative as well as in the treatment of rheumatism, hypertension and diabetes. In the present study, oxidative stress was induced in Vero cells by incubating the cells with glucose and the cell viability was measured by MTT assay. The concentration of glucose which 50% of cell viability was 125 mM $(IC_{50})$ and the cell viability was increased to $87.6{\pm}8.8%$ by treatment of the extracts of Acanthopanax divaricatus var. albeofructus. The antioxidative activity of water extract of different parts of the Acanthopanax plant was investigated by DPPH (1,1-diphenyl-2-picrylhydrazyl) assay, xylenol orange assay, TBARS (thiobarbituric acid reactive substances) assay and enzyme (superoxide anion and catalase) assay. Each extract (leaves, root, stem and fruits) of the plant showed free radical and $H_2O_2$ scavenging activity. The extract also inhibited lipid peroxidation and recovered enzyme (superoxide anion dismutase and catalase) activity in Vero cells treated with glucose.

• 대한의생명과학회지
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• 제16권4호
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• pp.279-285
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• 2010

Endogenous cannabinoids (endocannabinoids) display various pharmacological effects including pain control, anti-inflammation, and neuroprotection. The synthesis and release of endocannabinoids are regulated under both physiological and pathological conditions. The main degrading enzyme of endocannabinoid is fatty acid amide hydrolase (FAAH). Therefore we have developed the fluorescence-based assay system for FAAH. We established stable CosM6 cell lines expressing human FAAH. We also synthesized 2-oxo-2H-chromen-7-yl decanoate (DAEC) as a fluorogenic substrate for FAAH. When crude membrane extracts stably expressing FAAH was incubated with DAEC at $25^{\circ}C$, FAAH reacted specifically to DAEC and catalyzes the hydrolysis of DAEC into decanoic acid and highly fluorescent coumarin. Furthermore, the serin hydrolase inhibitor, phenylmethanesulfonylfluoride, inhibited the coumarin release to the reaction buffer in concentration dependent manner. This assay system is suitable for high-throughput screening since this system has simple experimental procedure and measurement method.

• 한국양식학회지
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• 제16권3호
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• pp.196-201
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• 2003

Sessile organisms such as the oyster Crassostrea gigas have been given much attention as a potential biomonitoring indicator to assess the impact of toxicants on aquatic organism. In this study, we exposed cells isolated from gill of oyster (Crassostrea gigas) to hydrogen peroxide in vitro. In addition oysters were in vivo exposed to pyrene and benzo(a)pyrene at various concentrations for 2 weeks. Comet assay was used to detect DNA single strand breaks and to investigate the application of this technique as a tool for aquatic biomonitoring. Hydrogen peroxide increased DNA single strand break with increasing concentration after 30 minutes exposure in vitro. Pyrene and benzo(a)pyrene caused DNA damage only at very high concentration (100 $\mu\textrm{g}$/L or 1000 $\mu\textrm{g}$/L) at two week exposure in vivo. DNA damage was relatively higher at hemocyte than at gill. It suggested that metabolized PAHs are transferred to hemolymph from digestive gland which have a relatively high enzyme activity, and attacked the DNA of hemocyte, while gill accumulated PAHs without degrading them to their metabolites due to low enzyme activity at gill. Both in vitro and in vivo exposure experiments showed that the comet assay is an effective tool on screening whether the organism are exposed to genotoxic contaminants.

• Bulletin of the Korean Chemical Society
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• 제25권1호
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• pp.115-118
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• 2004

We have examined the feasibility of using the specific interaction between mistletoe lectin I (ML I) and ${\beta}$-Dgalactose instead of the anti-ML I antibody in developing a homogeneous type competitive binding assay for ML I. We also have examined the feasibility of adapting the biotin/avidin mediated homogeneous assay for this system. Alkaline phosphatase (AKP) was employed as a single substrate enzyme label. The dose-response curve shows a detection range of 1-25 ${\mu}$g/mL and a linear response with a correlation coefficient of 0.99. To demonstrate the analytical utility of this method, 10 ${\mu}$g/mL of ML I was spiked into distilled water. The results show that the mean recovery was 10.03 ${\mu}$g/mL with an SD of 0.18. The difference between the spiked value and the mean recovery was 0.03 ${\mu}$g/mL, with a relative error of 0.3 and 1.6 % of RSD.

• KSBB Journal
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• 제10권4호
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• pp.374-385
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• 1995

Gelatin 분해성 proteolytic enzyme 생성균인 B. subtilis 80021가 생산하는 세포외 단백칠 분해효소 생산을 위한 영양 요구성과 배양조건을 검토한 결과 는 다음과 같다. 영양요구성은 1.5%(w/v) glucose와 2.5% (w/v) yeast extract에서 효소활성도가 최고치를 나타내였고 metalion은 $Co^{2+}, Zn^{2+}$효소생성의 저해제로 작용한 반면 $Fe^{2+}와 Mn^{2+}$에서는 촉진제로 작용 하여 최고 3.5배 효소생산생이 증가하였다. 아미노산 첨가 영향에서는 대부분의 아미노산이 효소활생 을 촉진시키는 것으로 확인되였다. 배지의 최적초기 pH는 7.0 부근이며 배양온도는 $35^{\circ}C$에서 효소에 따 라 다소 차이는 였었으나 배양시간 30시간 정도에서 최고의 효소생산성을 나다내었고 30°C에서는 42시간째에 효소활성도가 최고치를 나타내였다. Tannic acid 첨가에 대한 만백질 분해효소 생산에서는 발효 애지에 tannic acid 첨가시 완전한 효소생산 저해제로 작용하였으며 salicin을 탄소원으로 한 salicinase assay에서는 온도 빛 배양시간에 관계없이 효소활성 을 보이지 않았고 복합탄소원 이용에 대한 amylase assay에서는 $35^{\circ}C$, 36시간째에 최대의 효소활성을 나타내였고 Invertase assay에서는 $35^{\circ}C$, 30시간째 에 최대 효소활성도 2.5unit/ml을 나타내였다. 또한 gelatin 가수분해능에서는 45% (w/v)의 gelatin 용 액의 점도가 발효조효소액에 의해 96% (mPas)의 정도저하를 가져오는 것으로 확인되었다.

• Archives of Pharmacal Research
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• 제10권1호
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• pp.18-24
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• 1987

Mouse monocolonal antibodies to Hepatitis B surface antien (HBsAg) were prepared and their functional capabilities tested by the method of solid phase enzyme linked immuno sorbent assay (ELISA). HBsAg binding studies inicated that one monoclonal antibody 6E-1-1 bound more HBsAg at a faster rate than the other monoclonal antibodies. Also, for the binding inhibition studies with the selected monoclonal antibody 6E-1-1, one monoclonal antibody 8D-3-6 didn't exhibit binding inhibition for HBsAg. Then, a simultaneous ELISA method was developed for the immunodiagnosis of HBsAg. Different combinations of two monoclonal antibodies as solid phase and horseradish peroxidase (HRPO) labeled phase were studied. The combination of monoclonal antibody of higher affinity constant (6E-1-1) immobilized in a solid phase and monoclonal antibody of lower affinity constant (8D-3-6) as a HRPO laeled phase was more sensitive when two monoclonal antibodies of different affinity constants for HBsAg were prepared.

• Journal of Microbiology and Biotechnology
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• 제13권5호
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• pp.794-799
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• 2003

Enzyme-linked immunosorbent assay (ELISA) was developed for the rapid detection of Fusarium species, known as harmful fungi in food. One of the hybridoma cell lines (lB8) which produced a monoclonal antibody (Mab) specific to Fusarium extracellular polysaccharide (EPS) was screened and the Mab was produced and purified. A detection limit of the sandwich ELISA against F. moniliforme EPS was $0.001\;\mu\textrm{g}/ml$ in the standard curve. Among the 59 strains tested, most Fusarium species showed hight reactivity with Mab lB8, even when the culture broths were diluted 100,000 times. On the other hand, the other genera, except A. versicolor and Trichoderma viride, had no reactivity. When 1 to $100\;\mu\textrm{g}$ of F. moniliforme EPS was spiked into rice, potato, and mandarine orange, the average recoveries were 151%, 84%, and 94%, respectively, determined by sandwich ELISA. The correlation coefficients between the EPS levels determined by sandwich ELISA and the dry mycelial weight of the liquid culture of F. moniliforme, as well as between the EPS and colony forming unit in solid culture of potato, were 0.97 and 0.91, respectively.

• Journal of Microbiology and Biotechnology
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• 제9권6호
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• pp.764-772
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• 1999

To develop an enzyme-linked immunosorbent assay (ELISA) for detecting mold, we produced anti-mold polyclonal antibodies by immunizing extracellular polysaccharide (EPS) of Aspergillus flavus or Penicillium citrinum in rabbits subcutaneously. Using the purified antibody (Ab) and Ab-HRP conjugate, a sandwich ELISA for EPS was established. The standard curve of the ELISA showed the detection limit for P citrinum EPS to be $0.003{\;}\mu\textrm{g}/ml$. The cross-reactivities of the anti-P citrinum EPS Ab toward components of P citrinum such as EPS, liquid, and solid culture mycelium were 100, 10.5, and 0.58%, respectively, and those toward components of A. flavus such as EPS, liquid and solid culture mycelium, and spore were 300, 0.67, 0.29, and 0%, respectively. When the reactivities toward culture broths of 59 mold strains were tested by the sandwich ELISA, most of the Aspergillus (16 of 18) and Penicillium (14 of 16) strains along with one of the two Cladosporium strains gave positive signals in the culture broths even when diluted 1,000 fold, while the rest of species such as Fusarium, Absidia, Alternaria, and Candida gave negative signals. When the water extracts of 30 corn samples were analyzed by the sandwich ELISA, the EPS in the com could be detected in the concentration range of $0.1-1.6{\;}\mu\textrm{g}/g$.

• 한국미생물·생명공학회지
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• 제22권4호
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• pp.430-435
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• 1994

In order to develop an enzyme-linked immunosorbent assay (ELISA) for zearalenone (ZEA) in corn, we produced antisera by immunizing rabbits with ZEA-6'-carboxymethyloxime-BSA, purified polyclonal anti ZEA antibodies, and subsequently established a competitive indirect ELISA. The antibodies showed low cross-reactivity of 9.6~1.4% against ZEA analogues such as $\alpha$-zearalenol, $\beta$-zearalenol, $\alpha$-zearalanol, and $\beta$-zearalanol. From the standard curve of the ELISA for ZEA in corn, the detection range was found to be 0.3~1, 000 ng/ml. When artificially contaminated corns were assayed by the ELISA, the average recovery of ZEA spiked to 30~1, 000 ng/g was 109% (96~123%), although that of ZEA spiked to 10 ng/g was somewhat high (258%). The average coefficient of variation (CV) of the recovery was 18.0% (0.9~28.3%). When 9 corn samples naturally contaminated were assayed 3 times, the average CV of the determinitions was 27.7% (9.3~52.4%). Therefore, the ELISA was elucidated to be a practical tool for the detection of ZEA of 30 ng/g and more from corn.

• 한국식품위생안전성학회지
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• 제8권4호
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• pp.205-214
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• 1993

The monoclonal antibody to chloramphenicol(CAP) was produced to develop an enzyme-linked immunosorbent assay(ELISA) for residual CAP. An immunogen(CAP-BSA) was prepared by immunogen, antibody titer was measured by indirect ELISA. Spleen cells form the immunized mouse were fused with SP2/OAg14 myeloma cells. Among hybridomas selected in HAT media, 6 clones shown high antibody titer to CAP were subjected to cloning by limit dilution, and all of the monoclonal antibodies(MCA1, 2, 3, 4, 5, 7 and 9) produced by each clone were identified as IgG1 by ELISA isotyping analysis. Competitive ELISA condition was established by using the purified monoclonal antibody MCA1 as primary antibody and CAP-HSA conjugate as coating antigen. Standard curve of CAP(n=28) showed that the lowest detection limit of CAP is 20ng/ml level. The cross-reactivities of the 6 monoclonal antibodies showed that CAP sodium succinate. CAP base, P-nitrophenol, and p-nitrobenzyl alcohol were 89∼178, 0.050∼2.237, 0.056∼0.794 and 0.013∼7.939%, respectively. No cross-reactivities were observed with phenylalanine, tyrosine, glutamine, thiamphenicol, neomycin, streptomycin, gentamicin, sulfamethazine, sulfathiazole, chlortetracycline and p-aminobenzoic acid.