• Korean Journal of Pharmacognosy
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• v.46 no.4
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• pp.279-282
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• 2015

In order to quantify compound K(CK), anticancer component of Panax ginseng C. A. Meyer, high titer rabbit polyclonal antibodies (pAbs) were raised against a conjugate of CK and bovine serum albumin coupled by a periodate oxidation method. Coating antigen (CK-OVA) was also prepared by the same method with OVA. As a result of optimization of antiserum dilution (2,000 fold), coating antigen ($25{\mu}g/ml$) and other condition (incubation time, temperature and washing method), ELISA method for the determination of CK was established. The measuring range extended from 0.5 ng/ml to 25 ng/ml of CK. The antibodies exhibited minor or even no cross reactivities with protopanaxatriol (1.56%) and other tested ginsenosides, $GRb_1$ (0.11%), $GRg_1$ (0.07%) except protopanaxadiol (87.2%) from the structural similarity. And the antibody showed good correlation (r=0.987) between the assay values obtained by this ELISA method and HPLC. Therefore, the ELISA method could be very useful tools for the determination of CK in biological fluids because of their high sensitivity and specificity.

• Food Science of Animal Resources
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• v.32 no.6
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• pp.706-712
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• 2012

The present study was aimed to produce a chicken polyclonal antibody against Cronobacter muytjensii and to develop an immunoassay for its detection. Purification of anti-C. muytjensii IgY from egg yolk was accomplished using various methods such as water dilution and salt precipitation. As a result, sodium dodecyl sulfate-polyacrylamide gel electrophoresis produced two bands around 30 and 66 kDa, corresponding to a light and a heavy chain, respectively. Indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) was performed to determine the effectiveness of the chicken IgY against C. muytjensii. The optimum conditions for detecting C. muytjensii by indirect ELISA and checkerboard titration of the antigen revealed an optimum average absorbance at the concentration of 18 ${\mu}g/mL$, having ca. $10^8$ coated cells per well. The anti-C. muytjensii IgY antibody had high specificity for C. muytjensii and low cross-reactivity with other tested pathogens. In this assay, no cross-reactivity was observed with the other genera of pathogenic bacteria including Escherichia coli O157:H7, Salmonella Typhimurium, Staphylococcus aureus, Bacillus cereus, Enterobacter aerogenes, Salmonella Enteritidis and Listeria monocytogenes. In addition, detection of C. muytjensii in infant formula powder showed a low matrix effect on the detection curve of IC-ELISA for C. muytjensii, with similar detection limit of $10^5$ CFU/mL as shown in standard curve. These findings demonstrate that the developed method is able to detect C. muytjensii in infant formula powder. Due to the stable antibody supply without sacrificing animals, this IgY can have wide applications for the rapid and accurate detection of C. muytjensii in dairy foods samples.

• Parasites, Hosts and Diseases
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• v.28 no.4
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• pp.207-212
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• 1990

Enzyme-linked immunosorbent assay(ELISA) of paragonimiasis iloktsuenensis rat sera was performed using crude antigens of Paragonimus iloktsuenensis(PIA), P. westermani (PWA) and Clonorchis sinensis(CSA). Three crude antigens(PIA, PWA, CSA) were prepared to saline homogenated supernatants of whole adult worms. Infected rat sera were obtained biweekly from the albino rats fed 50∼.80 metacercariae of P. iloktsuenensis through gastric catheter. Experimental groups were divided into 4 groups: GI(controls), GII, GIII and GIV according to 1∼7 worms as GII, 10∼19 worms as GIII and 22∼40 worms as GIV, respectively, In ELISA, the mean OD values of each group for the homologous antigen(PIA) were increased significantly compared to the control sera at the 4th week of infection. With the progress of duration of infection, the mean OD values of infected sera of GII & GIV continuously increased up to the 12th week(last week), but in GIII the mean OD value increased until the loth week. No significance was noted among the infection dose groups (GII, GIII and GIV), after the 6th week of infection. Also, the OD values of all infected rats did not show any Proportional relytionships to the number of worms recovered. In brief, the antibody productivity of individual rats were strongly different. The rat sera infected with p. iloktsuenensis cross-reacted with those infected with P. westermani or C. sinensis, as identified by OD values.

• Development and Reproduction
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• v.9 no.2
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• pp.135-142
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• 2005

Vitellogenins(VTGs) are the precursor of egg-yolk proteins in most oviparous species from invertebrates to vertebrates. In oviparose vertebrates, VTGs are synthesized in the liver and transported through the blood to oocytes. In female fish, concentrations of plasma VTG increase rapidly at onset of vitellogenesis in the normal reproductive cycle. Male fishes also possess the gene for VTG, but plasma concentrations of the protein typically remain small, presumably due to low levels of endogenous estrogens. However, exposure of males to exogenous estrogenic mimics can result elevated. Therefore, the VTG in fish can be used as a useful biomarker for appropriate tools of endocrine disrupting compounds effects. In this studies, we prepared the test methods that can measure the plasma VTG level in the gobies that live in polluted area with mimic estrogen. For the purpose, we purified VTG of floating goby(Chaenogobius annularis) and prepared specific monoclonal and polyclonal antisera to yolk protein, then developed a sandwich competitive ELISA system for measurement of plasma VTG levels. Validation for the ELISA system using monoclonal and polyclonal antibodies against VTG was tested. The absorbance curve of serial dilutions of serum from vitellogenic female was paralleled to the standard curve of VTG, but normal male was not paralleled. The developed sandwich ELISA system was measured for VTG levels in plasma of common goby(Acanthogobius flaviman) and javeline goby(A. hasta) as well as in plasma of floating goby(C. annularis).

• Korean Journal of Food Science and Technology
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• v.30 no.6
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• pp.1415-1419
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• 1998

In order to evaluate the safety of greenhouse horticultures, a large number sample sources were collected, and the fungi of Aspergillus sp. and Penicillium sp. were isolated from them. Indirect competitive ELISA method and high performance liquid chromatography (HPLC) were applied to confirm the ochratoxin A producing abilities of isolated strains. One hundred ninety two sample sources including soil, pepper, strawberry and water mellon were collected for fungi isolation from western Gyeongnam, Andong and Gyeongbok. One hundred forty two strains of Aspergillus sp. and one hundred fifty three strains of Penicillium sp. were isolated respectively from them. The isolated fungi were tested for the production of ochratoxin A by ELISA. After culture of them on the modified sucrose low salt medium at $28^{\circ}C$ for 15 days, we found that five strains of Penicillium sp. produced ochratoxin A at the levels of $0.084{\sim}2.128\;{\mu}g/mL$. Among them, #129-2 strain isolated from water melon, showed the highest level of ochratoxin A as $2.128\;{\mu}g/mL$ broth. However, all of isolated Aspergillus sp. didn't produce ochratoxin A. When we compared the results of ELISA method with HPLC method, ochratoxin A production of each isolated strains showed very similar levels.

• The Plant Pathology Journal
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• v.22 no.1
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• pp.21-27
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• 2006

We immunized BALB/c mice with purified Cucumber green mottle mosaic virus isolate HY1 (CGMMV-HY1). Through the selection of positive clones that were grown on the HAT medium, four sensitive monoclonal clones (CG99-01, CG99-02, CG99-03, and CG99-04) were selected from 500 Hypoxanthine-guanine phosphoribosyltransferase positive hybridoma cells. Four sensitive clones of CGMMV-HYI were determined as IgM type of the subclass of mouse immunoglobulins Ig group. The titer of monoclonal antiserum against CGMMVHY1 was estimated 1:12,800 by the indirect ELISA. Although monoclonal antibodies (MAbs) from CG99-01 and from CG99-04 cross-reacted with Zucchini green mottle mosaic virus and Kyuri green mottle mosaic virus, MAb from the cell line CG99-03 was highly specific to CGMMV. No MAbs cross-reacted with Cucumber mosaic virus-Fny. Only CG99-04 reacted with Pepper mild mottle virus weakly and CG99-02 reacted with both CGMMV and KGMMV. CGMMV was detected from the rind of watermelon fruit by DAS-ELISA of CGMMV-HY1, but not from the flesh of watermelon. Average seed transmission rate of CGMMV in watermelon was $24\%$ from symptomatic watermelon collected from 5 regions of Gyeongnam province. CGMMV was detected by DAS-ELISA with specific MAb of CGMMVHY1 periodically from root stock, during the sequential process for nursery seedling in Haman. Necrotic spots on cotyledons of root stock seedling progressed to reveal the typical symptomatology on the primary leaves of scion upon grafting. Here, we have established MAb based ELISA system, which could accurately detect CGMMV from watermelon seeds, nursery seedlings, transplants and field samples from greenhouse or open out door field as well.

• Korean Journal of Veterinary Service
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• v.43 no.4
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• pp.237-244
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• 2020

Porcine transmissible gastroenteritis (TGE) has been a significant cause of economic losses in pig farming industry since 1950s. Although transmissible gastroenteritis virus (TGEV) has declined in recent years, it should not be excluded because of its characteristics; the frequency of gene mutation, the mortality in piglets, and the possibility for sudden incidence. Therefore, the herd-level monitoring of the virus is important to prevent further circulation of TGE. The aim of this study is to develop a large-scale sandwich enzyme-linked immunosorbent assay (ELISA) with high specificity to rapidly detect TGEV in feces by using monoclonal antibodies (Mabs). The TGEV specific Mabs were produced in hybridoma cells. Among the Mabs belonged to the IgG class developed by this study, the final selected 8H6, 1B7, 4G3, and 1F8 were identified to have the neutralization ability against TGEV. The sandwich ELISA was established using 8H6 as a reporter antibody and 1B7 and the reported 5C8 as a capture antibody. The developed sandwich ELISA was able to distinguish TGEV from other pathogenic diarrheal agents (porcine rotavirus, porcine reovirus, porcine epidemic diarrhea virus (PEDV), E. coli, and C. perfringens) in tissue culture as well as fecal samples. And the detection rate of TGEV in feces was 80% compared with RT-PCR. The results suggested that the developed sandwich ELISA may be useful in the herd-level monitoring for effective preventive measures due to the early diagnosis of TGEV using a large amount of samples.

• Research in Plant Disease
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• v.18 no.3
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• pp.231-235
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• 2012

Indian citrus ring spot virus (ICRSV) is known to cause a serious disease to citrus, especially to Kinnow mandarin, the popular cultivated citrus species in India. In this study, we developed diagnostic systems based on enzyme-linked immunosorbent assay (ELISA). In order to generate antibodies against ICRSV coat protein, we overexpressed the coat protein in Escherichia coli using the pET15b expression vector containing an optimized ICRSV coat protein gene. The recombinant ICRSV coat protein was overexpressed as soluble form at $37^{\circ}C$ upon IPTG induction. The protein was purified to 95% in purity by Ni-NTA column chromatography. The purified protein was immunized to rabbit for the generation of polyclonal antibody (PAb). The PAb showed a specific immunoreaction to recombinant ICRSV coat protein in western blot analysis and ELISA. Diluted rabbit antisera (10,000 fold) could detect less than 10 ng and 5 ng of the target protein in western blot and ELISA analysis, respectively.

• Korean Journal of Veterinary Research
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• v.41 no.1
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• pp.51-57
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• 2001

A procedure for extraction and purification of 8 kDa antigen of Brucella abortus RB51 was developed. Bacteria heat inactivated at $60^{\circ}C$, 30 min was extracted by 1% sarcosine and followed by fluid pressure liquid gel filtration chromatography of 2 series, Superose 12 HR 10/30 and Sephacryl S-100. There was produced $71.46{\mu}g/g$(wet) of 8 kDa antigen, and it resisted 1% trypsin, solved 1% triton X-100 higher than distilled water and inactivated 0.1% proteinase K. These results show that 8 kDa antigen may be a lipoprotein existed cell surface of B. abortus RB51. Also, we developed ELISA using purified 8 kDa surface antigen of Brucella abortus RB51 strain, its specificity and sensitivity was 95.0%, 98.6%, respectively. As compared with dot-blot assay using whole cell and ELISA using 8 kDa antigen, its correlation was 93.5%.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.13 no.12
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• pp.6208-6214
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• 2012

The most essential but missing components to understand and use toxic substances from marine microalgae are developing the fast, easy and economical determining technology for detecting it. In this paper we produced the antibodies against saxitoxin (STX). Mariculture keyhole limpet hemocyanin (mcKLH) and ovalbumin (OVA) were used as carrier proteins. mcKLH-STX conjugates were injected into the peritonial cavity of BALB/c mouse for immunization. After bleeding from mouse, anti-STX antiserum was isolated. Indirect enzyme-linked immunosorbent assays (ELISA) was performed to determine antiserum titer using the microtiter plate coated with free STX and OVA-STX. A goat anti-mouse IgG-phosphatase conjugate was used as secondary antibody to enable chromogenic reaction. Reactions of anti-STX antiserum were very specific on the OVA-STX and free STX. Sensitivity of anti-STX antiserum on STX was very high and STX detection limit was to be 64.9 ng/kg for indirect ELISA.