• Korean Journal of Food Science and Technology
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• v.41 no.2
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• pp.215-219
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• 2009

To conduct a risk assessment of $AFB_1$ intake, $AFM_1$, which is a metabolite of $AFB_1$ in the human and porcine urine, was determined by competitive direct ELISA (cdELISA). The detection limit of cdELISA using anti-$AFM_1$ antibody and $AFB_1$-HRP conjugate was 10 pg/mL. The recoveries of $AFM_1$ were 117-167% after the addition of $AFM_1$ in the human urine in a range of 3-100 pg/mL. 165 samples (95.5%) of those obtained from 172 persons evidenced measurable levels of urinary $AFM_1$. The detected $AFM_1$ ranges were 0-11.6 pg/mL and the average level of $AFM_1$ contamination was 2.74${\pm}$ 1.89 pg/mL. The estimated amount of $AFM_1$ excretion in the human urine was 3.97 ng/day and the estimated $AFB_1$ intake amount was 79.4 ng/day. The probable daily intake (PDI) of $AFB_1$ by the subjects was estimated to be 1.28 ng/kg bw/day, which was higher than the tolerable daily intake (TDI, 0.15 ng/kg bw/day). In the case of porcine urine, the $AFM_1$ ranged between 0.97-26.7 pg/mL and the average contaminated $AFM_1$ was 10.62${\pm}$4.39 pg/mL. The estimated amount of $AFM_1$ excretion in the porcine urine was 27.6 ng/day, and the estimated $AFB_1$ intake amount was 551 ng/day.

• Journal of Food Hygiene and Safety
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• v.18 no.3
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• pp.95-100
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• 2003

Competitive direct enzyme-linked immunosorbent assay(cdELISA) of aflatoxin $B_1$ ($AFB_1$) in deonjang(Korean-style soybean paste) and kochujang(fermented hot peppersoybean paste) and the level of $AFB_1$ in modern or traditional style deonjang and gochujang, produced in Korea, was surveyed by cdELISA. From the standard curve of the cdELISA, the detection limit of $AFB_1$ was 0.2 ng/m/. The average recovery of $AFB_1$ was 71.5% in the range of 1~100 ng/g after spiking $AFB_1$ into deonjang and it means that it could be possible to detect the $AFB_1$ in these range by the cdELISA in deonjang. Among the 30 kochujangs tested, no $AFB_1$ was detected in kochujangs. Among the 30 deonjangs, $AFB_1$ was detected in 6 ones in the range of 1.0~6.0 ng/g. The occurrence of $AFB_1$ in deonjang and kochujang tested in this study was less than the Korea Standard and Specification of aflatoxin in foods (10 ppb).

• Environmental Analysis Health and Toxicology
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• v.5 no.3_4
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• pp.29-36
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• 1990

Aflatoxins, produced by strains of Aspergillus flavus and Aspergillus parasiticus, can be found worldwide in corn, barley, peanuts, and other commodities. Among this group of toxins, aflatoxin B$_1$was realized to be one of the most potent environmental carcinogens, mutagens and teratogens. It is routinely monitored by methods such as thin layer chromatography, liquid chromatography, fluorodensitometric technique and radioimmunoassay. However, these assays are expensive, necessitate radioactive reagents, and require overnight incubation. In this study, the determination of fungal flora in several sorts cereals has been carried out in order to obtain an appropriate information of the population of fungi. The quantitative analysis of aflatoxin B$_1$has been carried out by High Performance Liquid Chromatography (HPLC) method and Enzyme Linked Immunosorbent Assay (ELISA). The results were summarized as follow: 1) From the 100 samples,313 colonies of fungi were isolated. Among the 313 colonies, 274 were possible to identify into 11 genera. The identified genera were Aspergillus Penicillium, Mucor, Rhizopus, Alternaria, Cladosorium, Fusarium, Circinella, Chrysosporium, Paecilomyces and Phoma. 2) Six of Aspergillus flavus were aflatoxin-producing strains. Aspergillus flavus isolated from sample barleys was contained the highest content (21.8 $\mu\textrm{g}$/ml) of aflatoxin B$_1$. 3) The yield of aflatoxin B$_1$-oxime compound was appromately 75%. Aflatoxin B$_1$-oxime-Human serum albumin was approved by formal consent as complete antigen. 4) Direct competitive ELISA permitted detection of 0.15 ng levels. In the quantitative microanalysis, ELISA was superior to HPLC method.

• Journal of Food Hygiene and Safety
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• v.4 no.3
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• pp.171-176
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• 1989

A rapid, simple method of ELISA was applied for the determination of aflatoxin BI in cereals from Y oungnam districts. Antibodies obtained cross reacted with aflatoxin B2 and to a less extent with other aflatoxin BI analogs. Response range for a typical standard curve was between I and 100 ppb. Fewer interference by spiked methanol-PBSdimethylformamide extracts ofrice was evidenced. Contents of aflatoxin BI from rice (65) and barley (116) were determined by competitive direct enzyme- linked immunosorbent assay as follows. Three out of 65 rices samples were positive. Rice samples of R-IS, R-30, and R-59 represent the aflatoxin B1 levels of $7.5\;\mu\textrm{g}.kg,\;6.0\;\mu\textrm{g}/kg,\;3.5\;\mu\textrm{g}/kg,\;3.3\;\mu\textrm{g}/kg$, respectively, and showed 4.6% aflatoxin BI contamination in rice samples. Meanwhile, four out of 116 barley samples were positive. VB-37 showed the highest aflatoxin Bllevels of $9.6\;\mu\textrm{g}/kg$ and VB-35, VB-15 and VB-54 represent $7.5\;\mu\textrm{g}.kg,\;6.0\;\mu\textrm{g}/kg\;and\;3.6\;\mu\textrm{g}/kg$, respectively, and showed 3.4% aflatoxin B1 contamination in barley samples.

• Journal of Veterinary Science
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• v.23 no.4
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• pp.55.1-55.12
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• 2022

Background: ASF was first reported in Kenya in 1910 in 1921. In China, ASF spread to 31 provinces including Henan and Jiangsu within six months after it was first reported on August 3, 2018. The epidemic almost affected the whole China, causing direct economic losses of tens of billions of yuan. Cause great loss to our pig industry. As ELISA is cheap and easy to operate, OIE regards it as the preferred serological method for ASF detection. P54 protein has good antigenicity and is an ideal antigen for detection. Objective: To identify a conservative site in the African swine fever virus (ASFV) p54 protein and perform a Cloth-enzyme-linked immunosorbent assay (ELISA) for detecting the ASFV antibody in order to reduce risks posed by using the live virus in diagnostic assays. Method: We used bioinformatics methods to predict the antigen epitope of the ASFV p54 protein in combination with the antigenic index and artificially synthesized the predicted antigen epitope peptides. Using ASFV-positive serum and specific monoclonal antibodies (mAbs), we performed indirect ELISA and blocking ELISA to verify the immunological properties of the predicted epitope polypeptide. Results: The results of our prediction revealed that the possible antigen epitope regions were A23-29, A36-45, A72-94, A114-120, A124-130, and A137-150. The indirect ELISA showed that the peptides A23-29, A36-45, A72-94, A114-120, and A137-150 have good antigenicity. Moreover, the A36-45 polypeptide can react specifically with the mAb secreted by hybridoma cells, and its binding site contains a minimum number of essential amino acids in the sequence 37DIQFINPY44. Conclusions: Our study confirmed a conservative antigenic site in the ASFV p54 protein and its amino acid sequence. A competitive ELISA method for detecting ASFV antibodies was established based on recombinant p54 and matching mAb. Moreover, testing the protein sequence alignment verified that the method can theoretically detect antibodies produced by pigs affected by nearly all ASFVs worldwide.

• Journal of Food Hygiene and Safety
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• v.16 no.3
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• pp.206-211
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• 2001

This experiment was conducted to investigate the effects of mixed culture with mycotoxigenic and non-mycotoxigenic fungi on mycotoxin production. For this work, Aspegillus flavus (aflatoxin producing strain), Aspegillus niger (non-mycotoxigenic strain) and Penicillium griseofulvum (patulin producing strain)were cultured in 5 ml SLS medium for 15 days under single or mixed culture. Aflatoxin was determined by direct competitive ELISA, whereas, patulin was measured by HPLC. The mycelial growth, pH and total acidity were also observed by general methods. The mycelial growth was slightly decreased in the mixed culture, meanwhile total acidity was increased and pH was shown lower than that in single culture. Aspergillus flavus produced 145 $\mu\textrm{g}$/ml of aflatoxin for 12 days single culture, but in mixed culture, aflatoxin was decreased to 93%, and was shown as 10.16$\mu\textrm{g}$/ml level. Patulin production in mixed culture was also decreased to 69.3% and was shown only 23.72$\mu\textrm{g}$/ml level as compared with in single culture.

• Journal of Microbiology and Biotechnology
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• v.19 no.1
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• pp.83-92
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• 2009

Individual immunochromatographic assays (ICG) for ochratoxin A (OTA) and zearalenone (ZEA) were optimized and used in the development of a one-step simultaneous immunochromatographic assay (OS-ICG) for the rapid multianalysis of two mycotoxins in corn samples. The nitrocellulose membrane of the OS-ICG was treated with OTA-bovine serum albumin (BSA), ZEA-ovalbumin (OVA), and anti-mouse IgG in the OTA test, ZEA test, and control zones, respectively. Monoclonal antibody-gold conjugates (OTA3 MAb-gold and ZEA2C5 MAb-gold) were sprayed onto the conjugate pad. The visual detection limits were 2.5 and 5 ng/ml for OTA and ZEA, respectively, and the results were obtained within 15 min after starting the analysis. An efficient, simple, and rapid extraction method using 30% MeOH/PBS was established and validated by analyzing the corn samples spiked with OTA/ZEA mixtures (0/0, 5/10, 10/20, and $20/30\;{\mu}g/kg$). The cut-off values of the OS-ICG for the spiked corn were 5 and $10\;{\mu}g/kg$ for OTA and ZEA, respectively. Natural corn samples were analyzed by OS-ICG, direct competitive enzyme-linked immunosorbent assay (DC-ELISA), and HPLC. Results of the OS-ICG were in good agreement with those obtained by DC-ELISA and HPLC. The developed OS-ICG offers a rapid, easy-to-use, and portable analytical system and can be used as a convenient qualitative tool for the on-site simultaneous determination of OTA and ZEA in cereals, food, and agricultural products in one analytical cycle.

• Journal of Food Hygiene and Safety
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• v.16 no.4
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• pp.274-279
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• 2001

Generally, non-aflatoxigenic fungi, such as Aspergillus oryzae, and Aspergillus are main microflora in Korean traditional fermented foods including Meju and soybean paste, but sometimes, Aspergillus flavus and Aspergillus parasiticus can be contaminated and accumulated aflatoxins during fermentation and storage. So the screening of aflatoxigenic strains in fermented traditional food is very important to improve the sanitary quality of those foods. In this work, we screened aflatoxin producing fungi from commercial Meju and soybean paste in Western Gyeongnam by immunoassay. Samples were randomly purchased from market of the commercial Meju(10 EA) and soybean paste(20 EA) in nine areas of Western Gyeongnam. Of the samples collected,24 strains and 22 strains of Aspergillus sp. were isolated from Meju and soybean paste, respectively. The isolated strains were cultured on SLS media at $25^{\circ}C$ for 15 days. The cultured broth were extracted with ethyl acetate and were analysed to determine aflatoxin B$_1$(AFB$_1$) by direct competitive ELISA(DC-ELISA). Six strains(25%) isolated from Meju, and 2 strains(9%) isolated from saybean paste, were confined as aflatoxin producing strains. The average range of aflatoxin productivity of isolates from Meju was 54.6 $\pm$ 38.7 ng/ml and that from soybean paste was 11.1 $\pm$ 8.6 ng/ml, respectively. Among them, isolated strain No. M-5-4 produced a high level of AFBl and showed 98.26 ng/ml of AFB$_1$. Every isolates were also re-confined their AFB$_1$productivity by thin layer chromatography(TLC). The TLC results also showed same trend as DC-ELISA results. As the above results, the screening of hazard mycotoxigenic fungi from traditional fermented foods should be necessary for the safety and the application of HACCP system in the food manufactory in Korea.

• Animal cells and systems
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• v.15 no.4
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• pp.259-267
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• 2011

This study was undertaken to produce a $F_{ab}$ fragment of a human monoclonal antibody reactive to oxidized and carbamylated low-density lipoprotein (oxLDL and cLDL) using phage display technology. An analysis of DNA sequences of this $F_{ab}$, termed plaque 15,16-46 $F_{ab}$, revealed that the rearranged $V_H$ was highly mutated. Complementarity-determining regions of the $V_H$ showed a very high R/S ratio and contained many positively charged amino acids. In direct binding and competitive ELISA, the $F_{ab}$ reacted strongly with both MDA-LDL and Cu-oxLDL forms of oxLDL, and also showed high affinity for cLDL. Immunofluorescence and immunohistochemical analyses showed that this $F_{ab}$ positively stained atherosclerotic aortic plaques in $ApoE^{-/-}$ mice as well as those in patients with atherosclerosis. The $F_{ab}$ also showed positive staining in placental decidua from patients with preeclampsia. It is suggested that the plaque 15,16-46 $F_{ab}$ against oxLDL and cLDL might possibly be applicable for developing a diagnostic reagent for both human and rodent animal research to detect and characterize atherosclerotic disease progression in atherosclerotic lesions as well as exploring the pathogenesis of atherogenic diseases such as preeclampsia.