• Bulletin of the Korean Chemical Society
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• 제18권5호
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• pp.515-519
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• 1997

In a competitive enzyme immunoassay, the performance was tested with different analyte-enzyme conjugates (signal generators) in their binding constants to antibody. Analyte (progesterone)-enzyme (glucose oxidase; GO) conjugates were chemically synthesized and purified by using a gel column with an immobilized antibody to progesterone. In an elution range from the column, four peaks were detected by measuring total enzyme activities. Results from further analysis indicated that the first peak contained mainly unreacted GO while the next three peaks conjugated GO with progesterone. These three conjugate preparations were compared in dose-response curves along with the unpurified mixture. The purified conjugates showed higher detection capabilities than did the mixture. Especially, the preparation in the second peak next to the free GO peak improved the detection limit five times. This performance was comparable to that of a progesterone-horseradish peroxidase conjugate that has been identified to have one progesterone ligand.

• 한국발생생물학회지:발생과생식
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• 제5권1호
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• pp.59-71
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• 2001

Testosterone의 radioimmunoassay (RIA) 가 1970년대 개발된 이래, 경제적이고, 측정 한계가 낮고, 감도가 높은 호르몬의 측정방법을 개발하여 왔다. 방사면역측정법 (RIA와 IRMA)은 비방사면역측정법 (NIA)로 대체되어 정밀도, 정확도, 특이도와 실용성 등이 크게 발전되었다. 최근 혈액, 타액, 분뇨 등에서 호르몬을 측정하여 남성과 여성의 성기능장애, 심리적스트레스나 기분의 변화, 폐경 및 남성호르몬 결핍증 등의 노화 현상, 자연 및 소 실험동물의 연구 등에서 적은 시료에서 여러가지 호르몬의 변화를 측정하기 위해, femtogram의 초감도 측정법의 개발이 요구되고 있다. 본 논문은 저자들이 지난 20여년간 Testosterone의 측정법을 개발하고 응용하던 결과를 토대로, 스테로이드의 측정법을 개발 정립하거나, 기존 방법을 연구에 변형할 때,측정의 신빙성을 높히려 할 때, 실험을 계획할 때 등에서 고려할 사항을 요약하고자 하였다. 또한 초감도 측정법을 정릴하고 측정의 질을 높힐 수 있는 방향을 제시하고자 하였다. 국내에서 대부분의 연구자가 선진 제국의 학자에게 의존하거나, 상업적 kit를 사용하여, 측정의 일관성이 없고, 비교가 어려우며, 외화를 낭비하고 있다. 표준화된 호르몬, 질이 좋은 항체를 생산 공급하고, 측정방법을 국내 실정에 맞추어 표준화하고, 정립하여 평가하고, 실험실간, 연구자간에 서로 신뢰할 수 있는 정보를 교환할 수 있는 정도 관리 계획이 만들어져야 한다. 또한 선진국처럼 후학들을 위하여 매년 학회에서 지원하는 Training programe 매년 실시되어야 하며, 국가적 차원의 자원이 필요하다고 판단된다.

• Clinical and Experimental Reproductive Medicine
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• 제14권2호
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• pp.138-148
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• 1987

5${\alpha}$-dihydrotestosterone(DHT)과 testosterone(T)은 남성 생식기관의 주 생성 호르몬들로 그 구조가 매우 비슷하여 이들 각 개를 특이하게 측정(specific determination)하는 방법이 개발되지 않고 있다. 본 연구는 고속액체 크로마토그라피(HPLC)를 이용하여 이들을 분리한 후 섬광면역측 정법(Luminescence immunoassay, LIA)으로 정량하는 방법을 개발하여 이들의 응용 가능성을 검토하고져 하였다. DHT와 T의 retention time은 각각 10.3min, 17.6min이었다. DHT-LIA와 T-LIA에 서 다른 스테로이드들과의 교차반응도는 방사면역측정법(RIA)과 대동소이하였다. 정도관리(quality control) 시료의 intra-assay variation은 DHT-LIA가 8.7%, T-LIA가 6.0%의 변이계수를 나타내었고, inter-assay variation의 변이계수는 각각 12.0% 및 15.3%이었다. 실측치(y)와 기대치(x)간의 관계를 보면, DHT-LIA경우는 Y=0.94X+0.9(r=0.989), T-LIA는 Y=1.01X+0.06(r =0.988)로 두 측정치 사이에는 통계적으로 유의한 차이가 없었다. 위의-측정방법을 이용하여 DHT-enanthate와 T-enanthate 처리후 혈청내 DHT 및 T의 농도변화를 조사한 실험결과 LIA와 RIA의 값사이에 유의한 차이가 없었다. 위의 결과로 보아 본 실험에서 개발된 DHT와 T의 섬광면역측정법은 정립되었다고 사려된다.

• 대한수의학회지
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• 제31권2호
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• pp.217-222
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• 1991

Most progesterone enzyme immunoassays(EIA) are used liquid phase double-antibody separation. These methods consume considerable time and reagents because of the requirements for several washing and centrifugation steps involving the reactants. Because of there several problems, we were prompted to develop an effective EIA system by the use of higher titer of progesterone antiserum free of anti-bovine serum albumin antibodies (anti-BSA). The results obtained were as follows. 1. The antibody of progesterone antiserum was high as $1.5{\times}10^5$. 2. Percent activity bound of progesterone antiserum was about 77 at a dilution to $5{\times}10^3$ times. 3. Progesterone antiserum was contained a large amount of anti-BSA antibodies. 4. The anti-BSA was completely absorbed by using of polymerised BSA. 5. The molecular weight of albumin polymer (polymerised BSA) obtained by using 2.5% glut. araldehyde was $5{\times}10^5$.

• 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.

• 대한한방내과학회지
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• 제21권3호
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• pp.433-441
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• 2000

Objective : To analyze the effects of Yukmijihwang-tang, Taeksa-tang, Silbi-um on mesangial cell proliferation, fibronectin synthesis and MHC-class II expression. Methods : Laboratory studies were performed with the method of surface enzyme immunoassays or flow cytometry after addition of peripheral blood mononuclear cells(PBMC) supernatants treated with medications using the cultured human mesangial cells. Results : 1. Silbi-um produces more suppressive effect than control group and hydrocortisone group on the mesangial cell proliferation. In Yukmijihwang-tang, Taeksa-tang and Silbi-um, mesangial cell proliferation significantly decreased than in hydrocortisone group 2. In the 'without fetal bovine serum' study, Yukmijihwang-tang take more suppressive effect than Control group on the fibronectin synthesis. In the 'with fetal bovine serum' study, Yukmijihwang-tang, Taeksa-tang, Silbi-um all have suppressive effect, but it hasn' t any statistical significance. 3. Yukmijihwang-tang, Taeksa-tang, Silbi-um all have a suppressive effect on the MHC-class II expression. Conclusions : Herb medicine generally show a suppressive effect on the suppression of the mesangial cell proliferation, fibronectin synthesis and MHC-class II expression.

• The Plant Pathology Journal
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• 제26권1호
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• pp.25-31
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• 2010

Papaya ringspot virus (PRSV) causes the most widespread and devastating disease in papaya. Isolates of PRSV originating from different geographical regions in south India were collected and maintained on natural host papaya. The entire coat protein (CP) gene of Papaya ringspot virus-P biotype (PRSV-P) was amplified by RTPCR. The amplicon was inserted into pGEM-T vector, sequenced and sub cloned into a bacterial expression vector pRSET-A using a directional cloning strategy. The PRSV coat protein was over-expressed as a fusion protein in Escherichia coli. SDS-PAGE gel revealed that CP expressed as a ~40 kDa protein. The recombinant coat protein (rCP) fused with 6x His-tag was purified from E.coli using Ni-NTA resin. The antigenicity of the fusion protein was determined by western blot analysis using antibodies raised against purified PRSV. The purified rCP was used as an antigen to produce high titer PRSV specific polyclonal antiserum. The resulting antiserum was used to develop an immunocapture reverse transcription-polymerase chain reaction (IC-RT-PCR) assay and compared its sensitivity levels with ELISA based assays for detection of PRSV isolates. IC-RT-PCR was shown to be the most sensitive test followed by dot-blot immunobinding assay (DBIA) and plate trapped ELISA.

• Interdisciplinary Bio Central
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• 제2권2호
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• pp.1.1-1.8
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• 2010

Microdevices have been used as effective experimental tools for the rapid and multiplexed analysis of individual cells in single-cell assays. Technological advances for miniaturizing such systems and the optimization of delicate controls in micron-sized space homing cells have motivated many researchers from diverse fields (e.g., cancer research, stem cell research, therapeutic agent development, etc.) to employ microtools in their scientific research. Microtools allow high-throughput, multiplexed analysis of single cells, and they are not limited by the lack of large samples. These characteristics may significantly benefit the study of immune cells, where the number of cells available for testing is usually limited. In this review, I present an overview of several microtools that are currently available for single-cell analyses in two popular formats: microarrays and microfluidic microdevices. Then, I discuss the potential to study human immunology on the single-cell level, and I highlight several recent examples of immunoassays performed with single-cell microdevice assays. Finally, I discuss the outlook for the development of optimized assay platforms to study human immune cells. The development and application of microdevices for studies on single immune cells presents novel opportunities for the qualitative and quantitative characterization of immune cells and may lead to a comprehensive understanding of fundamental aspects of human immunology.

• 대한한방소아과학회지
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• 제23권1호
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• pp.159-171
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• 2009

Objectives This study was evaluated the effects of CSE the regulatory mechanism of NO and cytokines in the LPS-stimulated Raw 264.7 cells. Methods The Coicis Semen MeOH extract dissolved in EMEM for 1 hour prior to the addition of LPS(1${mu}g/ml$). The cell viability was measured by MTT assay, and Nitric Oxide production was monitored by measuring the nitrite content in culture medium. The levels of cytokine and PGE2 were analyzed by sandwich immunoassays. Results CSE inhibited the production of NO (0.03 and 0.1 mg/ml), $TNF-{\alpha}$ (0.03 and 0.1 mg/ml), $IL-1{\beta}$ (0.03 and 0.1 mg/ml), IL-6 (0.03, 0.1 mg/ml) and PGE2(0.03 and 0.1 mg/ml) in Raw 264.7 cells activated with LPS(lipopolysaccharide). Conclusion According to the results above, Coicis Semen can produce anti-inflammatory effect, which may play a role in adjunctive therapy in Gram-negative bacterial infections.

• Bulletin of the Korean Chemical Society
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• 제34권3호
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• pp.922-926
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• 2013

3,3',4,4'-Tetrachlorobiphenyl (IUPAC PCB77) is one of seven indicative polychlorinated biphenyls (PCBs) in the surface sediments. The current study presents a novel polyclonal antibody for the determination of the PCB77 using indirect competitive enzyme-linked immunosorbent assay. Under optimum conditions, PCB77 was determined within the concentration range of 0.01-100 ${\mu}g\;L^{-1}$, with a detection limit of 0.057 ${\mu}g\;L^{-1}$. The assays were tested for their cross-reactivity profiles using 3 selected congeners and 4 Aroclor products. The assays were highly specific for coplanar PCB congeners, but less specific for Aroclor1248. The spiked recoveries from five sediment samples were 86%-114% for PCB77 from ELISA, which were satisfactory. The current study demonstrated that the developed antiserum and immunoassay procedure can be used to detect PCB77 in environmental samples. The results of the sediment analysis were confirmed by conventional GC/ECD.