• 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.

• 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.

• 대한수의학회지
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• 제31권2호
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• pp.223-228
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• 1991

Most progesterone enzyme immunoassays(EIA) are used liquid-phase double-antibody or single-antibody seperation. These methods consume considerable time and reagents because of the requirements for several washing and centrifugation steps involving the reactants. Because of these several problems, we were prompted to develop an effective enzyme-linked immunosorbent assay(ELISA) system that would be equal or superior to RIA for assay of progesterone. The results were obtained as follows. 1. Cross reaction of the progesterone antiserum with other steroids determined was shown with progesterone(100%), $11{\alpha}$-deoxycorti-costerone(2.271%), but the other steroids were shown below 0.9%. 2. Standard curve for progesterone ELISA was shown available difference according to progesterone concentration from 0 to 1,000pg/ml. 3. The lower limit of sensitivity was 0.2pg/well 4. Progesterone concentration was 1.6ng/ml for before parturition, and that was below 0.5ng/ml for after parturition. This development enzyme-linked immunosorbent assay for progesterone can be detected pregnancy diagnosis in cattle, and also applicable 10 research on physiological function including such as reproductive disorders.

• 한국수산과학회지
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• 제50권5호
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• pp.547-552
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• 2017

The viral hemorrhagic septicemia virus (VHSV) has an extensive host range, and infects farmed and wild fish inhabiting both freshwater and marine ecosystems. Enzyme-linked immunosorbent assay (ELISA) is highly useful in diagnosing viral hemorrhagic septicemia. However, ELISA shows high, non-specific background reaction with fish antibodies. In this study, we optimized the antigen and antibody concentrations used for detecting specific antibodies in VHSV-infected olive flounder to reduce non-specific binding, and improve the sensitivity of ELISA. The results suggested that OD (optical Density) values were valid when ELISA was performed with $0.1{\mu}g/well$ of virus, involving blocking with blocking buffer (Roth, Roti-Block), 1:300-1:600 dilution with flounder antisera, and 1:1000 dilution with anti-flounder IgM and HRP-conjugated goat anti-mouse IgG for detecting the VHSV antibody in flounder sera. Furthermore, 11 different VHSV strains isolated in Korea from 2012 to 2016 were used to infect the fish. The results showed no correlation between viral pathogenicity and antibody production. This research is a basic study on the application of antibody detection in the diagnosis of viral hemorrhagic septicemia in the olive flounder.

• 한국식품영양과학회지
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• 제27권6호
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• pp.1152-1159
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• 1998

In order to develop more rapid and reproducible analysis of biotin known as vitamin H, attempts were made to establish the condition for enzyme linked immunosorbent assay(ELISA) compared with traditional microbiological assay(MBA). Antibiotin and antiserum were obtained from the immunized rabbits injected with emulsion of biotin KLH conjugate and Freund's adjuvant. The antiserum showed cross reactivity on biocytin, a derivative of biotin, which is converted to biotin in intestine, at the rate of 177%(median inhibitory concentration(IC50)=12.58ppb), but not on other derivatives such as desthiobiotin, diaminobiotin and 2 imino biotin. Specific antibody for biotin was purified from the antiserum through protein A column and desalting column. The conditions of competitive direct ELISA (cdELISA) were established. Detection range of biotin concentration by cdELISA was 0.01∼300ng/ ml(ppb). In the spike test with milk, fruit flake and pine carrot juice, the correlation coefficient between two methods of MBA and ELISA was reliably consistent at the value of r=0.992. But detection of biotin by microbiological assay(MBA) was rather restricted in range and nonspecific. Detection range of biotin by MBA was 0.1∼0.5ng/ml(ppb). It showed cross reactivities on biocytin and desthiobiotin at the rate of 80.1% and 66.7%, respectively. In conclusion, ELISA revealed a significant improvement compared with MBA for the biotin detection in terms of sensitivity, detection range and cross reactivity. In addition, a variety of samples could be analyzed rapidly and conveniently at one time by using ELISA. These results strongly suggest that the ELISA is very promising for the practical application to detect biotin contents in a wide range of food stuffs.

• 한국식품과학회지
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• 제32권5호
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• pp.1009-1014
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• 2000

PA를 분석하기 위하여 효소면역측정법을 개발하고자 하였다. Bc방법과 Po방법으로 BSA에 PA를 conjugation하여 각각의 PA-BSA conjugate(PA-BSA[Bc]와 PA-BSA[Po])를 제조하였으며, 이를 토끼에 면역하여 항PA-BSA 항체를 얻었다. 항PA-BSA[Po] 항체를 사용한 ciELISA의 결과에서 경합반응이 제대로 일어나지 않았기 때문에, 식품 속에 있는 PA를 검출하기 위해서 PA-BSA[Po]을 코팅한 후 항PA-BSA[Bc] 항체를 사용하였다. 이 결과에서 PA의 검출한계가 1 ppm인 것을 확인할 수 있었으며, 교차반응을 통해 PA 유도체들에 대해서도 항PA-BSA[Bc] 항체가 PA에 대해 특이성이 매우 강하였다. 또한, MBA의 결과에서는 그 검출한계가 10ppb인 것을 확인할 수 있었다. 분석시료인 계란(109%), 상추(64%), 소간(344%)의 식품시료에 대한 실험에서 상추를 제외하고는 ciELISA는 MBA의 결과와 비교해 볼 때 양호한 결과를 얻을 수 있었다. 그러므로, ciELISA는 MBA보다 분석시간, 교차반응 등의 면에서 장점이 있어 식품 중 PA의 검출에 효과적으로 활용할 수 있을 것이다.

• Bulletin of the Korean Chemical Society
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• 제24권3호
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• pp.334-338
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• 2003

A simple synthetic method for haptens of organophosphorus (OP) pesticides with a spacer arm (aminocarboxylic acid) attached at the pesticide thiophosphate group was developed and was applied to the synthesis of haptens for the OP fungicide tolclofos-methyl. Using the haptens, a selective enzyme-linked immunosorbent assay (ELISA) for tolclofos-methyl was developed. One of the haptens was coupled to BSA to use as an immunogen. Rabbits were immunized with this conjugate to obtain polyclonal antibodies to tolclofos-methyl. The antisera were screened against another hapten coupled to ovalbumin (OVA). Using the serum with highest specificity, an antigen-coated ELISA was developed, which showed an $IC_{50}$ of 160 ng/mL with the detection limit of 20 ng/mL. The antibodies showed negligible cross-reactivity with other OP pesticides. An antibody-coated ELISA was also developed, which showed an $IC_{50}$ of 410 ng/mL with a detection limit of 130 ng/mL.

• 대한수의학회지
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• 제30권2호
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• pp.187-192
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• 1990

돼지로부터 분리한 Staphylococcus hyicus subsp hyicus 489주의 protein A 존재여부와 함량을 indirect hemagglutination 및 enzyme-linked immunosorbent assay(ELISA)법으로 조사하였다. Indirect hemagglutination text에 의하여 cell-bound protein A 및 extracellular protein A 보유균은 489주 중 각각 87.7% 및 36.0%로 나타났다. ELISA법에 의한 이들 균의 protein A함량 측정에서 전균주의 extracellular protein A는 1ng/ml미만이었으며, cell-bound protein A함량은 대부분의 균주에서 1ng/ml 미만이었고 11주가 25~108ng/ml 수준이었다.

• 식물병연구
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• 제25권3호
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• pp.131-135
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• 2019

Cylindrocarpon destructans causes ginseng root rot and produces radicicol that has an antifungal effect. In this study, we developed a method to detect this fungus using enzyme-linked immunosorbent assay (ELISA). Secreted proteins of C. destructans were used as antigens to obtain C. destructans-specific IgG from mouse. Out of 318 monoclonal antibodies generated from mouse, two antibodies (Cd7-2-2 and Cd7-2-10) showed highest specificity and sensitivity. Indirect ELISA using both antigens successfully detected C. destructans in soils, but direct ELISA using IgG conjugated with horseradish peroxidase failed to detect antigens in soils. The indirect ELISA developed here can efficiently detect the fungus and help manage ginseng root rot disease in fields.