• Bulletin of the Korean Chemical Society
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• v.25 no.1
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• pp.115-118
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• 2004

We have examined the feasibility of using the specific interaction between mistletoe lectin I (ML I) and ${\beta}$-Dgalactose instead of the anti-ML I antibody in developing a homogeneous type competitive binding assay for ML I. We also have examined the feasibility of adapting the biotin/avidin mediated homogeneous assay for this system. Alkaline phosphatase (AKP) was employed as a single substrate enzyme label. The dose-response curve shows a detection range of 1-25 ${\mu}$g/mL and a linear response with a correlation coefficient of 0.99. To demonstrate the analytical utility of this method, 10 ${\mu}$g/mL of ML I was spiked into distilled water. The results show that the mean recovery was 10.03 ${\mu}$g/mL with an SD of 0.18. The difference between the spiked value and the mean recovery was 0.03 ${\mu}$g/mL, with a relative error of 0.3 and 1.6 % of RSD.

• Bulletin of the Korean Chemical Society
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• v.24 no.12
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• pp.1859-1861
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• 2003

• Bulletin of the Korean Chemical Society
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• v.17 no.11
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• pp.1018-1022
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• 1996

Adenosine deaminase (ADA) has been utilized as the label in devising a potentiometric homogeneous assay for riboflavin and riboflavin binding protein (RBP). The proposed homogeneous assay method employs an ADA-biotin conjugate as the signal generator and an avidin-riboflavin conjugate as the signal modulator in the solution phase. The catalytic activity of the ADA-biotin conjugate is inhibited in the presence of an excess amount of the avidin-riboflavin conjugate, and the observed inhibition is reversed in an amount proportional to the concentration of RBP added. When the analyte riboflavin is added to this mixture of ADA-biotin, avidin-riboflavin and RBP, the activity of the enzyme conjugate is re-inhibited in an amount proportional to the concentration of riboflavin. Since the enzyme label used in this system is ADA, an ammonia-producing enzyme, a potentiometric rather than photometric detection scheme is used to monitor the enzymatic activity in the assay.

• Analytical Science and Technology
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• v.13 no.5
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• pp.624-629
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• 2000

A simple and rapid homogeneous enzyme-linked binding assay for mistletoe lectin I(ML I) was developed using a coupled enzyme system of malate dehydrogenase (MDH) and D-galactose. A highly substituted MDH-galactose conjugate was prepared by employing an isothiocyanate method for formation of thiourea bond. In the presence of ML I, ML I inhibits the activity of the conjugate based on the ML I/D-galactose specific interaction. Thus, the concentration of ML I can be related to the homogeneous inhibition of the MDH-galactose conjugate. Using this method. ML I can be measured at the level of microgram per milliliter within 10 minutes.

• Journal of Microbiology and Biotechnology
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• v.17 no.6
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• pp.1002-1009
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• 2007

A rapid, convenient homogeneous competitive enzyme immunoassay for estimating the amount of fenthion is described. The assay utilizes glucose-6-phosphate dehydrogenase-hapten conjugates that are inhibited in solution by antibodies obtained from bovine serum albumin-hapten conjugates. In order to investigate the effects of bridging group recognition on the sensitivity of dose response characteristics, the bridging groups of varying alkyl chain length were attached at the phosphate position of fenthion. Among the antibodies used, the one obtained from the use of hapten (fenthion analog) with the same bridging group structure that was used in preparing the enzyme-fenthion conjugates showed maximum inhibition (up to 51.8%) in the absence of fenthion. In the presence of fenthion, the activity of the enzyme-hapten conjugate is regained in an amount proportional to the fenthion concentration. Under the optimized condition, the $ED_{50}$ value for fenthion was $0.809{\mu}g/ml$. The assay developed in this study is a rapid effective screening method for fenthion prior to precise analysis.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.04a
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• pp.281-281
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• 1996

The immunoassay method is frequently used for the identification and quantitation of ultratrace amount of bioactive substances. Homogeneous liposome immunoassays, which can avoid the use of radioisotopes and separation steps, have recently been reported in many publications. Cytolysin-mediated liposome immunoassay using melittin ever been studied but showed limited applications. Here, we designed a homogeneous liposome immunoassay using Clostridium perfringens phospholipase C (PLC), an enzyme which catalyzes the hydrolysis of phosphatidylcholine in biological membranes, as a cytolysin.

• Journal of Microbiology and Biotechnology
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• v.26 no.3
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• pp.588-595
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• 2016

IFN-γ release assays (IGRAs) have been developed as viable alternative diagnostic tools for detecting latent tuberculosis infection (LTBI). A customized homogeneous sandwich luminescent oxygen channeling immunoassay (LOCI) was used to quantify IFN-γ levels in IGRAs. Samples were collected from healthy volunteers (n = 40) who were T-Spot-negative and T-Spot-positive patients (n = 32) at rest. Then the amount of IFN-γ in the supernatant of IGRAs was measured by LOCI. The results demonstrated a low background, and high sensitivity, specificity, accuracy, and reproducibility, and a short assay time (only 30 min) with LOCI for IFN-γ. The recovery range was 81.63-102.06%, the coefficients of variation were below 5%, and the limit of detection was 19.0 mIU/ml. Excellent agreement between LOCI IFN-γ and the T-SPOT.TB test was obtained (97.2% agreement, κ = 0.94). The LOCI IFN-γ concentrations were significantly higher in T-Spot-positive patients than in the healthy group (p < 0.001). Moreover, as observed for the comparative LOCI IFN-γ assay, IFN-γ concentrations were related to the numbers of T-SPOT.TB spots. We have established an in vitro blood test for LTBI diagnosis, defined as LOCI IFN-γ. A high level of agreement between the LOCI IFN-γ method and T-SPOT.TB assay was observed in clinical studies that showed the LOCI IFN-γ method could determine LTBI. This study shows acceptable performance characteristics of the LOCI IFN-γ assay to diagnose LTBI.

• Proceedings of the PSK Conference
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• 2001.10a
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• pp.218.1-218.1
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• 2001

• YAKHAK HOEJI
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• v.27 no.2
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• pp.117-124
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• 1983

For development of $EMIT-T_{3}$ assay, the conjugation of 3, 5, 3'-triiodothyroformic acid NHS ester to G6PDH was attempted in various reaction conditions. Up to now, the best conjugation condition was the ratio of $T_{3}$-NHS:G6PDH=100 in 25% carbitol-Tris buffer at pH 9, $0^{\circ}C$ during overnight. The obtained $T_{3}$-G6PDH conjugates usually had 20% residual enzyme activity which was inhibited by 40-70% with various $anti-T_{3}$ antibodies. Utilizing the conjugate I and an antibody (S2633G), a useful standard curve for $T_{3}$ assay was obtained in the range of 0 to 5ng/ml with 499 EMIT units of separation.

• Bulletin of the Korean Chemical Society
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• v.31 no.10
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• pp.2813-2818
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• 2010

A novel method was developed for the speciation of chromium in natural water samples based on homogeneous liquid-liquid extraction and determination by flame atomic absorption spectrometry (FAAS). In this method, Cr(III) reacts with a new Schiff's base ligand to form the hydrophobic complex, which is subsequently entrapped in the sediment phase, whereas Cr(VI) remained in aqueous phase. The Cr(VI) assay is based on its reduction to Cr(III) by the addition of sodium sulfite to the sample solution. Thus, separation of Cr(III) and Cr(VI) could be realized. Homogeneous liquid-liquid extraction based on the pH-independent phase-separation process was investigated using a ternary solvent system (water-tetrabutylammonium ion ($TBA^+$)-chloroform) for the preconcentration of chromium. The phase separation phenomenon occurred by an ion-pair formation of TBA and perchlorate ion. Then sedimented phase was separated using a $100\;{\mu}L$ micro-syringe and diluted to 1.0 mL with ethanol. The sample was introduced into the flame by conventional aspiration. After the optimization of complexation and extraction conditions such as pH = 9.5, [ligand] = $1.0{\times}10^{-4}\;M$, [$TBA^+$] = $2.0{\times}10^{-2}\;M$, [$CHCl_3$] = $100.0\;{\mu}L$ and [$ClO_4$] = $2.0{\times}10{-2}\;M$, a preconcentration factor (Va/Vs) of 100 was obtained for only 10 mL of the sample. The relative standard deviation was 2.8% (n = 10). The limit of detection was sufficiently low and lie at ppb level. The proposed method was applied for the extraction and determination of chromium in natural water samples with satisfactory results.