• 한국어병학회지
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• 제12권1호
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• pp.24-31
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• 1999

어류의 edwardsiellosis를 진단하기 위한 solid phase ELISA법에 대한 최적화 분석을 실시하였다. 부분 정제한 E. tarda Edk-2에 대한 토끼 항혈청을 sodium bicarbonate 완충용액에 $50{\mu}g/ml$ 농도로 희석하여 overnight 반응시켜 주었을 때 polystyrene bead 표면의 항체 immobilization이 최적화되었다. $50{\mu}g/ml$의 biotin 표지화 항체와 1:2000으로 희석된 extravidin-peroxidase를 차례로 처리하였을 때 최적의 반응을 나타내었으며 이렇게 최적화시킨 정성적 solid phase ELISA법은 EDTA 추출법으로 조제된 항원에 대해서는 $1{\times}10^5$ cells/ml, 열 추출법으로 조제된 항원에 대해서는 $1{\times}10^5$ cells/ml의 검출한계를 나타내었다. E. tarda Edk-2에 대한 토끼 항혈청을 이용한 본 연구의 solid phase ELISA법은 우리나라 양어장의 넙치 병어로부터 분리한 여러 한국형 E. tarda 균주와도 높은 교차반응을 나타내었다. 이러한 것은 본 기법을 다른 지역에서 분리한 여러 strain의 E. tarda에 대한 진단을 위하여 본 실험의 항혈청을 사용하여 가능하다는 것을 보여 주었다.

• 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 Veterinary Science
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• 제23권5호
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• pp.32.1-32.11
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• 2022

Background: Classical swine fever (CSF) is a severe infectious disease of pigs that causes significant economic losses to the swine industry. Objectives: This study developed a solid-phase blocking enzyme-linked immunosorbent assay (spbELISA) method for the specific detection of antibodies against the CSF virus (CSFV) in porcine serum samples. Methods: A spbELISA method was developed based on the recombinant E2 expressed in Escherichia coli. The specificity of this established spbELISA method was evaluated using reference serum samples positive for antibodies against other common infectious diseases. The stability and sensitivity were evaluated using an accelerated thermostability test. Results: The spbELISA successfully detected the antibody levels in swine vaccinated with the C-strain of CSFV. In addition, the detection ability of spbELISA for CSFV antibodies was compared with that of other commercial ELISA kits and validated using an indirect immunofluorescence assay. The results suggested that the spbELISA provides an alternative, stable, and rapid serological detection method suitable for the large-scale screening of CSFV serum antibodies. Conclusions: The spbELISA has practical applications in assessing the vaccination status of large pig herds.

• 한국어병학회지
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• 제10권2호
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• pp.75-86
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• 1997

어류의 감염 조직으로부터 edwardsiellosis의 원인균인 Edwardsiella tarda를 whole cell 자체로 직접 검출할 수 있는 solid phase ELISA법을 연구하였다. A. hydrophila ATCC7966, V. anguillarum HUFP5001, Y. ruckeri 11-4, E. ictaluri 및 Streptococcus sp. NG8206 등의 어병세균에 대해 ELISA법으로 실시한 교차반응 분석에서 A. hydrophila ATCC7966 균주와 V. anguillarum HUFP5001 균주가 E. tarda Edk-2에 대한 토끼 항혈청에 대해 높은 교차반응을 나타내었으나, 항혈청을 A. hydrophila ATCC7966 FKC로 흡착시킴으로써 교차반응을 제거할 수가 있었다. 그러나, 응집항체가 측정 결과와는 달리, ELISA 분석에서는 E. tarda 분리 균주간의 교차반응이 매우 높은 것으로 나타났다. Tissue homogenate내에 있는 항원을 검출함에 있어, 조직내의 지질이나 단백질 성분이 함께 분석용 plate에 coating되어 감도가 훨씬 감소하므로 ELISA법의 적용을 위해서는 감염 조직의 homogenate를 PBS에 100배 이상 희석한 후 진단을 실시해야 하는 것으로 나타났다. Tissue homogenate내에 있는 생균을 항원으로 하여 직접 검출할 때에는 검출한계가 $1{\times}10^3$ cells/ml로 나타나 FKC 항원의 사용에 비하여 더 증가된 감도를 보여주었다. 결론적으로 본 ELISA법은 양식장에서 발생한 edwardsiellosis를 진단함에 있어서 특이적이고 신속하며 민감한 방법으로 확인되었다.

• Bulletin of the Korean Chemical Society
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• 제24권5호
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• pp.664-666
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• 2003

• 대한미생물학회지
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• 제20권1호
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• pp.91-102
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• 1985

The advantages of enzyme-linked immunosorbent assay(ELISA) are its senstivity and simplicity in detecting IgG, IgM and IgA antibody. To apply ELISA to diagnosis of typhoid fever, antigen such as lipopolysaccharide of Salmonella typhi or killed whole cell must be coated on solid phase. It is easy to coat lipopolysaccharide on ELISA plate but troublesome to purify it. As it is easy to obtain the killed whole cells, the development of the appropriate method by which those antigens of S. typhi are optimally coated on solid phase is needed. To establish the appropriate method, carbonate buffer, methanol or poly-L-lysine was applied as binding substance on polystyrene or polyvinylchloride plate as solid phase when the killed whole cell antigens of S. typhi varided as follows: $10^6$, $10^7$, $10^8$ and $10^9\;cell/ml$. The criteria of the optimal method were determined as follows: 1. The optical density of positive sera is above 1.0(0.6 in IgM) at 1:10 serum dilution and is 0.3(0.2 in IgM) higher than that of negative sera: 2. The O.D. of sera is flat or lowering according to serum dilution: 3. It must be that the O.D. of negative sera is lower than 0.2 at the point of serum dilution where the O.D. of positive sera is higher than 1.0(0.5 in IgM). The results obtained were summarized as follows: 1. The methods which fitted the above criteria were to use poly-L-lysine as binding substance, polyvinylchloride plate as solid phase and $10^7\;cell/ml$ as antigen concentration of S. typhi(poly-L-lysine/polyvinylchloride/$10^7$) and poly-L-lysine/polyvinylchloride/$10^8$ in detecting IgG antibody, methanol/polystyrene/$10^9$, poly-L-lysine/polyvinylchloride/$10^8$ and poly-L-lysine/polyvinylchloride/$10^9$ in IgM and carbonate buffer/polystyrene/$10^8$, carbonate buffer/polystyrene/$10^9$, methanol/polystyrene/$10^8$, methanol/polyvinylchloride/$10^8$, methanol/polyvinylchloride/$10^9$, poly-L-lysine/polyvinylchloride/$10^8$ and poly-L-lysine/polyvinylchloride/$10^9$ in IgA. 2. The coaling method using poly-L-lysine, polyvinylchloride plate and $10^8\;cell/ml$ was best to assay IgG, IgM and IgA antibody all in one. By this method, to assay the each immunoglobulin calss with an appropriate fixed serum dilution, 1:320 dilution was best.

• 대한수의학회지
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• 제31권4호
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• pp.403-409
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• 1991

Progesterone의 단크론성 항체를 생산, 이용하여 감도가 높으면서도 신속히 측정할 수 있는 ELISA 기법을 처음으로 개발코져 실시하였다. 단크론성 항체는 종래의 면역방법에 의해 획득한 항혈청에 비해 약 10배의 결합율을 보였고 titer 역시 높았다. Dot-blot 분석 결과 단크론성 항체는 IgM이었다. 경합반응은 2시간으로 충분하였고, progesterone 표준용액을 이용한 표준 곡선은 0~1000pg/well에서 거의 직선적이었다. Progesterone의 단크론성 항체를 이용한 ELISA는 임상적으로는 물론 연구용으로도 신속한 항체의 기능 측정에는 물론 각종 번식 관련의 지표로 충분히 활용될 수 있을 것으로 판단된다.

• Reproductive and Developmental Biology
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• 제34권3호
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• pp.161-167
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• 2010

Early diagnoses of pregnancy for animal such as swine and bovine is extremely important to increase income of a farmhouse and for the management of farm. For the development of immunoasaay system of pregnancy in swine, we report a competitive heterologous enzyme linked immunosorbent assay (ELISA) for the direct measurement of oestrone sulfate (E1S) in diluted urine using anti-E1G (glucuronide) monoclonal antibody which cross react with ElS. The principle of assay was based on the typical solid-phase competitive ELISA methods using E1G-HRP (horseradish peroxidase) as a tracer and E1S for standard. The method had a reasonable sensitivity for the detection of E1S with 0.15 ng/ml as a detection limit. The intra-assay and inter-assay precisions were raging coefficient of from 8.50~9.67% and 8.50~9.87%, respectively, which were quite acceptable. In a field trial with a group 37 sows (18 non-pregnancy and 19 pregnancy sows) after day 29~30 post service, the concentration of E1S were determined to be below 30 ng/ml in all non-pregnancy group and over 48 ng/ml in pregnancy group except one sample. The method described here, heterologous ELISA for the measurement of E1S in urine is good enough for monitoring the early pregnancy test of swine.

• 대한임상검사과학회지
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• 제39권1호
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• pp.42-48
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• 2007

This study was carried out to review and evaluate anti-CCP ELISA assay for diagnostic in RA patients from an early arthritis clinic (EAC). The subjects were obtained from patients visiting the outpatient clinic of the Dept. of Rhumatology med.of P hospital in Daegu, during 6 months from July 1, 2006 to December 31, 2006. The subjects were 140 cases : 80 cases from RA patients (60 women and 20 men; mean age 58 years; range, 32-68 years) confirmed by clinical diagnostic. 39 cases of these RA patients were classified as having early RA (EAC). 50 cases (non-RA) did not fulfill the criteria for RA, and 10 cases were from healthy individuals. We performed the analysis with solid phase-ELISA method (ETI-max3000, Diasorin; Italy) for anti-CCP and Nephelometry assay (Roche/Hitachi 902 analyzer; USA) for RF. The results obtained were summarized as follows ; anti-CCP ELISA is more specific than RF Nephelometry assay (specificity 94% vs 90%) to diagnose RA patients with suspected EAC (early arthritis clinic). The combination test "anti-CCP and RF" had a very high specificity (specificity 98.3%, PPV; RA group 96%, EAC 95%), the difference was statistically significant (p<0.05). Anti-CCP ELISA had more sensitivity in EAC (Early arthritis clinic) patients than chronic RA patients (sensitivity 64% vs 24%, respectively), anti-CCP of RA group and EAC group was more specific than RF (anti-CCP PPV; 92%, 89% vs 89%, 81% respectively), the difference was statistically significant (p<0.05). The difference of antibody concetration between anti-CCP and RF for RA and the control group is statistically significant (p<0.05). In conclusion, anti-CCP ELISA testing may be useful if performed concomitantly with RF Nephelometry assay to diagnose RA patients with suspected EAC (early arthritis clinic).

• Clinical and Experimental Reproductive Medicine
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• 제17권1호
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• pp.71-80
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• 1990

항정자항체를 검사할 수 있는 면역분석법개발을 위하여 immunoaffinity chromatography로 분리 정제한 정자표현항원을 microtiter plate에 고정화 시킨것과 효소와 화학발광체로 표지된 2차항체를 사용하여 ELISA법과 CIA법을 개발하고 이 방법의 이용 가능성을 검토하기 위하여 기존 방법인 Kibrick test법으로 임상소견이 다른 남성혈청을 분석 비교하여 다음과 같은 결과를 얻었다. 1. 인간정자 표면항원을 분리하기 위한 immunoaffinity column을 제작키 위하여 인간정자를 토끼에 주사하여 정자에 대한 항혈청을 생산하였으며 이를 Protein A-Sepharose column으로 분리 정제하여 CNBr activated Sepharose-4B에 coupling시켜 immunoaffinity column을 제작하였다. 이 column에 균질화된 정자를 반응시키고 SDS를 넣은 Tris-HCI buffer로 용출시켰을때 60KD정도의 분자량을 갖는 분획을 얻었다. 2. 분리 정제된 human IgG를 microtier plate에 농도를 달리하여 고정화 시키고 ELISA용 Goat anti-human IgG-HRP conjugate와 CIA용 Rabbit anti-human IgG-ABEI-H conjugate와 반응시켜 그 활성도를 측정하였던 바 농도에 따라 반응의 정도가 감소하였다. 3. ELISA법으로 양성혈청의 희석곡선을 작성하였을때 경사가 완만한 것과 급한 것의 두 종류의 경향을 띈 곡선으로 대별되었으며 완만한 경사를 나타내는 것에서 1:160 희석치에서 O.D가가 0.1이하를 음성, 0.1이상 0.2이하를 약양성(weak positive) 그리고 0.2이상을 양성으로 판별하였다. 4. ELISA, CIA 그리고 Kibrick test로 동일시료를 분석 비교하였던바 ELISA와 CIA는 거의 동일한 상관관계를 보였으나 Kibrick test와는 50% 수준만 일치함을 보였다.