• Parasites, Hosts and Diseases
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• 제52권3호
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• pp.305-310
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• 2014

Ascidian soft tunic syndrome (AsSTS) caused by Azumiobodo hoyamushi (A. hoyamushi) is a serious aquaculture problem that results in mass mortality of ascidians. Accordingly, the early and accurate detection of A. hoyamushi would contribute substantially to disease management and prevention of transmission. Recently, the loop-mediated isothermal amplification (LAMP) method was adopted for clinical diagnosis of a range of infectious diseases. Here, the authors describe a rapid and efficient LAMP-based method targeting the 18S rDNA gene for detection of A. hoyamushi using ascidian DNA for the diagnosis of AsSTS. A. hoyamushi LAMP assay amplified the DNA of 0.01 parasites per reaction and detected A. hoyamushi in 10 ng of ascidian DNA. To validate A. hoyamushi 18S rDNA LAMP assays, AsSTS-suspected and non-diseased ascidians were examined by microscopy, PCR, and by using the LAMP assay. When PCR was used as a gold standard, the LAMP assay showed good agreement in terms of sensitivity, positive predictive value (PPV), and negative predictive value (NPV). In the present study, a LAMP assay based on directly heat-treated samples was found to be as efficient as DNA extraction using a commercial kit for detecting A. hoyamushi. Taken together, this study shows the devised A. hoyamushi LAMP assay could be used to diagnose AsSTS in a straightforward, sensitive, and specific manner, that it could be used for forecasting, surveillance, and quarantine of AsSTS.

• 한국진공학회:학술대회논문집
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• 한국진공학회 2014년도 제46회 동계 정기학술대회 초록집
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• pp.126-126
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• 2014

Label-free biomolecular assay based localized surface plasmon resonance (LSPR) of noble metal nanoparticles enables simple and rapid detection with the use of simple equipment. Nanosized metal nanoparticles exhibit a strong absorption band when the incident light frequency is resonant with the collective oscillation of the electrons, which is known as the LSPR. Here we demonstrate localized surface plasmon resonance (LSPR) substrates such as plasmonic Au nanodisks fabricated by a nanoimprinting process and gold nanorod-immobilized surfaces and their applications to highly sensitive and/or label-free biosensing. To increase detection sensitivity various bioreceptors weree designed. A single chain variable fragment (scFv) was used as a receptor to bind C-reactive protein (CRP). The results of this effort showed that CRP in human serum could be quantitatively detected lower than 1 ng/ml. Aptamers, which were immobilized on gold nanorods, were used to detect mycotoxins. The specific binding of ochratoxin A (OTA) to the aptamer was monitored by the longitudinal wavelength shift of LSPR peak in the UV-Vis spectra resulting from the changes of local refractive index near the GNR surface induced by accumulation of OTA and G-quadruplex structure formation of the aptamer. According to our results, OTA could be quantitatively detected lower than 1 nM level. Additionally, aptamer-functionalized GNR substrate was quite robust and can be regenerated many times by rinsing at 70 OC to remove bound target. During seven times of washing steps, the developed OTA sensing system could be reusable. Moreover, the proposed biosensor exhibited selectivity over other mycotoxins with an excellent recovery for detection in grinded corn samples, suggesting that the proposed LSPR based aptasensor plays an important role in label-free detection of mycotoxins.

• 한국자원식물학회지
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• 제25권2호
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• pp.193-199
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• 2012

본 연구는 미국 수수 수집종의 작물학적 특성 및 생리활성을 검정하기 위하여 수행되었다. 대부분의 미국 수수 수집종은 200 cm 이상의 큰 초장을 보였으며, 간경 또한 2cm 이상의 값을 나타냈다. 수수 수집종 중에서는 Sweet-N-Sterile(#4)이 가장 우량한 수수의 생장 특성을 보였다. 수수의 DPPH를 이용한 활성 검정 결과, Premium stock(#1), Early Sumac(#7), SS Silage(#9), WGF Grain Sorghum(#11) 4개의 수집종에서 기존의 항산화제인 ${\alpha}$-tocopherol과 BHA와 비슷한 활성을 보였고 BHT보다는 높은 활성을 보였다. 항미생물 실험결과 BMR Gold I(#3)이 가장 높은 활성을 나타냈으며, 특히 $Escherichia$ $coli$에서 125 ${\mu}g$/ml으로 높은 활성수치를 나타냈다.

• 한국임상수의학회지
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• 제24권2호
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• pp.77-81
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• 2007

본 연구는 개에서 심장사상충을 검출하기 위하여 표준검사를 적용하지 않은 상황에서 중합연쇄반응검사 (PCR)의 진단 능력을 평가하였다. 효소면역검사법 (ELISA)과 PCR 검사를 동시에 사용한 경우 PCR 검사의 민감도와 특이도는 두 검사의 조건부 독립을 가정한 상태에서expectation-maximization (EM) 알고리즘을 이용한 최대우도법과 Bayesian 기법으로 두 집단 검사 모형으로 분석하였다 2002-2004년 기간 중 심장사상충검사 결과를 기록한 의무기록에서 무작위로 266개 결과를 추출하여 133개씩 2회의 시험으로 배치하였다. 2회의 분석결과를 종합할 때 EM 알고리즘에서 PCR 검사의 민감도와 특이도는 각각 96.4-96.7%와 97.6-98.8%, Bayesian기법에서는 94.4-94.8h와 97.1-98%로 추정되었다. PCR 검사는 심장사상충을 스크리닝하는 도구로 유용하며, 표준검사를 적용하지 않은 상황에서 진단검사의 특성을 추론하는 방법으로 Bayesian 기법은 매우 유용함을 확인하였다.

• 보존과학연구
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• 통권27호
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• pp.129-143
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• 2006

황남대총 남분에서 출토된 3쌍의 금제태환이식을 상대로 과학적 조사를 실시하였다. 태환이식의 주환인 태환은 사용된 금판의 수량에 따라 제작기법이 크게 4가지로 구분될 수 있는데 황남대총 남분출토 3쌍의 태환을 확인해 본 결과 3가지의 기법을 확인할 수 있었기에 이는 기술적인 발전에 의한 것이 아니라 제작하고자 하는 금속의 성분에 따라 각기 방법을 달리 하였음을 알 수 있다. 그리고 태환의 성분분석 결과 쌍임에도 많은 차이를 보였으니 한쪽은 금은 합금으로, 또 다른 쪽은 99.5%의 순은(Ag(에 금아말감을 이용한 금도금으로 되었다. 또 다른 한쌍은 33.0%라는 많은 량의 은(Ag)이 합금되어 다른 금제이식에 비해 황금색이 아닌 붉은 색을 띠고 있었다. 이번에 조사된 금제 태환이식에는 모두 소량의 구리(Cu)가 모두 함유된 것으로 나타났다.위와 같은 쌍임에도 불구하고 제작기법의 상이, 중량 및 금속의 성분에 있어서 상당한 차이를 보이고 있음은 황남대총 남분에서 출토된 금제 태환이식이 제작기간의 촉박함에 의한 급조임을 의미한다고 할 수 있다. 앞으로 지역적인 제한에서 벗어나 여러 지역과의 비교분석 즉 자연과학적인 방법을 통한 태환이식의 성분 및 제작기법을 검토한다면 유추가 아닌 과학적인 근거를 기초로 기존에 알 수 없었던 다양한 고대의 금속공예기술과 사회상을 파악하는데 큰 역할을 할 수 있을 것이다.

• 한국동물위생학회지
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• 제34권4호
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• pp.321-331
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• 2011

Mycobacterium bovis, a member of the M. tuberculosis complex (MTC), is the causative agent of bovine tuberculosis. Detection of M. bovis and M. tuberculosis using conventional culture- and biochemical-based assays is time-consuming. Therefore, a simple and sensitive molecular assay for rapid detection would be of great help in specific situations such as faster diagnosis of bovine tuberculosis (bTB) infection in the abattoirs. We developed a novel multiplex real-time PCR assay which was applied directly to biological samples with evidence of bTB and it was allowed to differentiate between M. bovis and M. tuberculosis. The primers and TaqMan probes were designed to target the IS1081 gene, the multi-copy insertion element in the MTC and the 12.7-kb fragment which presents in M. tuberculosis, not in the M. bovis genome. The assay was optimized and validated by testing 10 species of mycobacteria including M. bovis and M. tuberculosis, and 10 other bacterial species such as Escherichia coli, and cattle lymph nodes (n=113). The tests identified 96.4% (27/28) as M. bovis from the MTC-positive bTB samples using conventional PCR for specific insertion elements IS1081. And MTC-negative bTB samples (n=85) were tested using conventional PCR and the real-time PCR. When comparative analyses were conducted on all bovine samples, using conventional PCR as the gold standard, the relative accuracy of real-time PCR was 99.1%, the relative specificity was 100%, and the agreement quotient (kappa) was 0.976. The detection limits of the real-time PCR assays for M. bovis and M. tuberculosis genomic DNA were 10 fg and 0.1 pg per PCR reaction, respectively. Consequently, this multiplex real-time PCR assay is a useful diagnotic tool for the identification of MTC and differentiation of M. bovis and M. tuberculosis, as well as the epidemiologic surveillance of animals slaughtered in abattoir.

• 한국가축번식학회지
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• 제9권2호
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• pp.133-139
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• 1985

An immunoassay may be defined as an analytical procedure involving the competitive reaction between a limiting concentration of specific antibody and two populations of antigen, one of which is labelled or immobillized. The advent of immunoassay has revolutionised our knowledge of reproductive physiology and the practice of veterinary and clinical medicine. Radioimmunoassay (RIA) was the first of these methods to be developed, which meausred the analyte with good sensitivity, accuracy and precision (1,2). The essential components of RIA are:-(i) a limited concentration of antibodies, (ii) a reference preparation, and (iii) an antigen labelled with a radioisotope (usually tritium or iodine-125). Most procedures invelove isolating the antibody-bound fraction and measuring the amount of labelled antigen. Good facilities are available for scintilltion counting, data reduction nd statistical analysis. RIA is undergoing refinement through:-(i) the introduction of new techniques to separate the antibody-bound and free fractions which minimize the misclassification of labelled antigen into these compartments, and the amount of non-specfic binding. (3), (ii) the development of non-extration for the measurement of haptens (4), (iii) the determination of a, pp.rent free (i.e. non-protein bound) analytes (5), and (iv) the use of monoclonal antibodies(6). In 1968, Miles and Hales introduced in important new type of immunoassay which they termed immunora-diometric assay (IRMA) based on t도 use of isotopically labelled specific antibodies(7) in a move from limited to excess reagent systems. The concept of two-site IRMAs (with a capture antibody on a solid-phase, and a second labelled antibody to a different antigenic determinant of the analyte) has enabled the development of more sensitive and less-time consuming methods for the measurement of protein hormones ovar wide concentration of analyte (8). The increasing use of isotopic methos for diverse a, pp.ications has exposed several problems. For example, the radioactive half-life and radiolysis of the labelled reagent limits assay sensitivity and imposes a time limit on the usefulness of a kit. In addition, the potential health hazards associated with the use and disposal of radioactive cmpounds and the solvents and photofluors necessary for liquid scientillation counting are incompatable with the development of extra-laboratory tests. To date, the most practical alternative labels to radioisotopes, for the measurement of analytes in a concentration > 1 ng/ml, are erythrocytes, polystyrene particiles, gold sols, dyes and enzymes or cofactors with a visual or colorimetric end-point(9). Increased sensitivity to<1 pg/ml may be obtained with fluorescent and chemiluminescent labels, or enzymes with a fluorometric, chemiluminometric or bioluminometric end-point. The sensitivity of any immunoassay or immunometric assay depends on the affinity of the antibody-antigen reaction, the specific activity of the label, the precision with which the reagents are manipulated and the nonspecific background signal (10). The sensitivity of a limited reagent system for the measurement of haptens or proteins is mainly dependent upon the affinity of the antibodies and the smalleest amount of reagent that may be manipulated. Consequently, it is difficult in practice to improve on the sensitivity obtained with iodine-125 as the label. Conversely, with excess reagent systems for the measurement of proteins it is theoretically possible to increase assay sensitivity at least 1000 fold with alternative luminescent labels. To date, a 10-fold improvement has been achieved, and attempts are being made to reduce the influence of other variables on the specific signal from the immunoreaction.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제35권6호
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• pp.353-359
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• 2013

Purpose: Extensive research is actively ongoing for development of an ideal bone substitute that meets the gold standard. Tooth was selected as a donor site for evaluation of potentials in bone substitutes based on its similar chemical compositions to alveolar bone. Previous studies have evaluated inorganic components of autogenous tooth bone graft material (AutoBT) and osteoconductivity. In continuation from the previous studies, the current study was conducted for analysis of organic components and evaluation of osteoinductivity of AutoBT. Methods: Forty-six extracted teeth were collected from actual patients (Korea Tooth Bank, R&D Institute). Extracted teeth were processed into AutoBT and implanted in dorsal subcutaneous muscular tissues of 15 athymic mice. Biopsy samples were harvested at two, five, and eight weeks. The Bradford assay, sodium dodecyl sulphate polyacrylamide gradient gel, and western blotting were performed for investigation of organic contents of AutoBT. Results: Histology analyses showed signs of new bone formation as early as two weeks. Results of the Bradford assay indicated the existence of noncollagenous proteins (NCP). 0.29% (2.89 mg/g) of proteins were extracted by weight in the root portion of AutoBT; 0.02% (0.029 mg/g) and 1.79% (17.93 mg/g) of proteins were measured by weight in crown and block-form of AutoBT, respectively. However, recombinant human bone morphogenetic protein-2 was not observed in AutoBT. Conclusion: Within the limitation of the current study, AutoBT induced new bone formation by NCP embedded in dentin.

• Parasites, Hosts and Diseases
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• 제39권1호
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• pp.67-76
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• 2001

This experiment was focused on the characterization of anti- Toxoplasma monoclonal antibodies (mAbs) and the effect of mAbs on the parasite invasion of mouse peritoneal macrophages. Twenty eight mAbs including M110, M556, R7A6 and M62l were characterized by Ab titer, immunoglobulin isotyping and western blot pattern. Antibody titer (optical density) of 4 mAbs. Ml 10. M556. R7A6 and M62l. were 0.53,0.67, 0.45 and 0.39 (normal mouse serum; 0.19) with the same IgGl isotypes shown by Enzyme-linked immunosorbent assay (ELISA). Western blot analysis showed that Ml 10. M556. R7A6 and M62l reacted with the 33 kDa (p30),31 kDa (p28),43 kDa and 36 kDa protein. Immuno-gold labelling of mAbs M110, M556, R7A6 and M621 reacted with the surface membrane, dense granules and parasitophorous vacuolar membrane (PVM) , rhoptries and cytoplasm of tachyzoite, respectively. For in vitro assay, preincubation of tachyzoties with four mAbs, Ml 10, M556, R7A6 and M62l resulted in the decrease of the number of infected macrophages (p < 0.05) and the suppression of parasite multiplication at 18 h post-infection. Four monoclonal antibodies including Ml 10 (SAGI) were found to have an important role in the inhibition of macrophage invasion and T. gondii multiplication in vitro, and these mAbs may be suitable for vaccine candidates, diagnostic kit and for chemotherapy.

• Tuberculosis and Respiratory Diseases
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• 제83권4호
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• pp.283-288
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• 2020

Background: Latent tuberculosis (TB) infection among TB contacts is diagnosed using plain chest radiography and interferon-gamma release assays (IGRAs). However, plain chest radiographs often miss active TB, and the results of IGRA could fluctuate over time. The purpose of this study was to elucidate changes in the results of the serial IGRAs and in the findings of the serial submillisievert chest computed tomography (CT) scans among the close contacts of active pulmonary TB patients. Methods: Patients age 20 or older with active pulmonary TB and their close contacts were invited to participate in this study. Two types of IGRA (QuantiFERON-TB Gold In-Tube assay [QFT-GIT] and the T-SPOT.TB test [T-SPOT]) and submillisievert chest CT scanning were performed at baseline and at 3 and 12 months after enrollment. Results: In total, 19 close contacts participated in this study. One was diagnosed with active pulmonary TB and was excluded from further analysis. At baseline, four of 18 contacts (22.2%) showed positive results for QFT-GIT and T-SPOT; there were no discordant results. During the follow-up, transient and permanent positive or negative conversions and discordant results between the two types of IGRAs were observed in some patients. Among the 17 contacts who underwent submillisievert chest CT scanning, calcified nodules were identified in seven (41.2%), noncalcified nodules in 14 (82.4%), and bronchiectasis in four (23.5%). Some nodules disappeared over time. Conclusion: The results of the QFT-GIT and T-SPOT assays and the CT images may change during 1 year of observation of close contacts of the active TB patients.