• The Journal of the Korean Society for Microbiology
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• v.21 no.1
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• pp.151-161
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• 1986

In order to develop sensitive and sepcific assay methods for E. coli heat labile enterotoxin(LT) hybridoma cell lines secreting LT specific monoclonal antibody were obtained. LT was purified from cell lysate of E. coli O15H11. The steps included disruption of bacteria by French pressure, DEAE Sephacel ion exchange chromatography, Sephadex G200 gel filtration, and second DEAE Sephacel ion exchange chromatography, successively. Spleen cells from Balb/c mice immunized with the purified LT and $HGPRT^{(-)}$ plasmacytomas, $P3{\times}63Ag8.V653$ were mixed and fused by 50% (w/v) PEG. Hybrid cells were grown in 308 wells out of 360 wells, and 13 wells out of them secreted antibodies reacting to LT. Among these hybridoma cell 1G8-1D1 cell line was selected since it had produced high-titered monoclonal antibody continuously. By using culture supernatant and ascites from 1G8-1D1 cells the monoclonal antibody was characterized, and an assay system for detecting enterotoxigenic E. coli was established by double sandwich enzyme-linked immunosorbent assay (ELISA). The following results were obtained. 1. Antibody titers of culture supernatant and ascites from 1G8-1D1 hybridoma cells were 512, and 102, 400, respectively by GM1-ELISA and its immunoglobulin class was IgM. 2. The maximum absorption ratio of 1G8-1D1 cell culture supernatant to LT was 90% at $300\;{\mu}g/ml$ of LT concentration. LT concentration shown at 50% absorption ratio was $103.45{\mu}g$ and the absorption ratio was decreased with tile reduction of LT concentration. This result suggests that monoclonal antibody from 1G8-1D1 hybridoma cell bound with LT specifically. 3. The reactivities of 1G8-1D1 cell culture supernatant to LT and V. cholerae enterotoxin(CT) were 0.886 and 0.142(O.D. at 492nm) measured by the GM1-ELISA, indicating 1G8-1D1 monoclonal antibody reacted specifically with LT but not with CT. 4. The addition of 0.1ml of ascites to 0.6mg and 0.12mg of LT decreased the vascular permeability factor to 41% and 44% respectively, but it did not completely neutralize LT. 5. By double sandwich ELISA using monoclonal antibody, as little as 75ng of the purified LT per ml could be detected. 6. The results by assay of detecting LT in culture supernatants of 14 wild strains E. coli isolated from diarrhea patients by the double sandwich ELISA were almost the same level as those by reverse passive latex agglutination.

• Korean Journal of Microbiology
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• v.44 no.3
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• pp.228-236
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• 2008

Validation of viral safety is essential in ensuring the safety of mammalian cell culture-derived biopharmaceuticals, because numerous adventitious viruses have been contaminated during the manufacture of the products. Mammalian cells are highly susceptible to Reovirus type 3 (Reo-3), and there are several reports of Reo-3 contamination during the manufacture of biopharmaceuticals. In order to establish the validation system for the Reo-3 safety, a real-time RT-PCR method was developed for quantitative detection of Reo-3 in cell lines, raw materials, manufacturing processes, and final products as well as Reo-3 clearance validation. Specific primers for amplification of Reo-3 RNA was selected, and Reo-3 RNA was quantified by use of SYBR Green I. The sensitivity of the assay was calculated to be $3.2{\times}10^0\;TCID_{50}/ml$. The real-time RT-PCR method was proven to be reproducible and very specific to Reo-3. The established real-time RT-PCR assay was successfully applied to the validation of Chinese hamster ovary (CHO) cell artificially infected with Reo-3. Reo-3 RNA could be quantified in CHO cell as well as culture supernatant. When the real-time RT-PCR assay was applied to the validation of virus removal during a virus filtration process, the result was similar to that of virus infectivity assay. Therefore, it was concluded that this rapid, specific, sensitive, and robust assay could replace infectivity assay for detection and clearance validation of Reo-3.

• Journal of Korean Society of Environmental Engineers
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• v.28 no.3
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• pp.257-264
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• 2006

This study investigated the effects of hybrid process(chemical coagulation, Fenton oxidation and ceramic UF(ultrafiltration)) on COD and color removals of commercial reactive dyestuffs. In the case of chemical coagulation, the optimal concentrations of $Fe^{3+}$ coagulant for COD and color removals of RB49(reactive blue 49) and RY84(reactive yellow 84) were determined according to the different coagulant dose at the optimal pH. They were 2.78 mM(pH 7) in RB49 and 1.85 mM(pH 6) in RY84, respectively. In the case of Fenton oxidation, the optimal concentrations of $Fe^{3+}\;and\;H_2O_2$ were obtained. Optimal $[Fe^{2+}]:[H_2O_2]$ molar ratio of COD and color removals of RB49 and RY84 were 4.41:5.73 mM and 1.15:0.81 mM, respectively. In the case of ceramic UF, the flux and rejection of supernatant after Fenton oxidation were investigated. After ceramic UF for 9 hr, the average flux of RB49 and RY84 solutions were $53.4L/m^2hr\;and\;67.4L/m^2hr$ at 1 bar, respectively. In addition, the permeate flux increased and the average flux recovery were 98.5-99.9%(RB49) and 91.0-97.3%(RY84) according to adopting off-line cleaning(5% $H_2SO_4$). Finally, COD and color removals were 91.6-95.7% and 99.8% by hybrid process, respectively.

• Journal of Appropriate Technology
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• v.6 no.2
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• pp.174-182
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• 2020

The permissible limit of ammonia concentration in drinking water recommended by the World Health Organization (WHO) is 1.5 mg/L. However, in the case of groundwater in Kathmandu, Nepal, the concentration of ammonia fluctuates dramatically from 0 to 120 mg/L at different locations and groundwater depths (Chapagain et al., 2010). Such a high concentration of ammonia causes aesthetic problems in drinking water, such as bad taste and odor; hence, prior treatment is required. In Kathmandu, half of the population utilizes groundwater, which is also employed for drinking water, but owing to a lack of knowledge of household water filters, residents of Kathmandu tend to depend greatly on commercially available jar water than on the installation of a proper household filtration method. Thus, in our study, we employed adsorption and reverse osmosis (RO) as two of the most viable decentralized/household treatment options to address the issue of high contamination of ammonia in drinking water. We evaluated their performances from technical and the economic perspectives using synthetically prepared groundwater at varying ammonia concentrations (50 mg/L and 15 mg/L). Consequently, it was found that adsorption via ion exchange (IE) resin was a comparatively better ammonia removal technology than RO, with 100% ammonia removal even after regeneration; the removal by RO was limited to up to 90%. Furthermore, our study suggests that IE is the most suitable ammonia removal technology for places with lower water consumption (< 50 L/day), whereas RO seemed to be a cost-effective technology for places with higher water consumption, where the daily water demand exceeds 50 L/day. Lastly, these assessments suggest that installing a suitable household treatment system would be more efficient and sustainable from both technical and economic points of view than purchasing commercially bottled water.

• Korean Journal of Ichthyology
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• v.35 no.2
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• pp.91-100
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• 2023

The early life history of Silurus microdorsalis living in Jahocheon Stream was studied by observing egg and morphological development. Live fish were captured in June 2018, then reared in a circulating filtration system under a 14L : 10D photoperiod with a water temperature of 18℃. To artificially induce spawning, females were injected with 0.5 mL of Ovaprim (Syndel, Nanaimo, BC, Canada) per kg of body weight, and males were injected with 10,000 IU/kg body weight of human chorionic gonadotropin. Approximately 15 h later, eggs were artificially inseminated by the dry method. Mature eggs were light pale yellow, which separated them from immature eggs. Fertilized eggs were 2.16±0.06 mm (n=8) in diameter and fully hatched at 181 h after fertilization. The fertilization rate was 63.1±2.2%, and 10.0±3.7% of the embryos were malformed at 18℃. The rates of development were 181 h at 18℃, 109 h at 21℃, and 76 h at 24℃. The larval size immediately after hatching was 4.64±0.22 mm (n=8), and the larvae displayed negative phototaxis at 1 day after hatching. The total larval length on 7 days after hatching was 12.47±0.53 mm, with 25~30 basal anal fin rays and 14~16 basal caudal fin rays observed. The total larval length was 14.13±0.51 mm on 9 days after hatching, and approximately 90% of the black endoplasmic reticulum was deposited on the head and body. The dorsal fin had formed, and a single basal body was observed. On 15 days after hatching, the total larval length was 16.69±0.31 mm; the number of basal caudal fin rays (18 poles) was an integer because 2 dorsal fin basal rays and 60~63 anal fin basal rays were observed. The total larval length was 28.96±1.10 mm on 50 days after hatching; the numbers of caudal fins (n=18), dorsal fins (n=3), pectoral fins (n=11), and anal fin basal rays (n=67~73) were integers.