• Bulletin of the Korean Chemical Society
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• v.31 no.2
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• pp.384-388
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• 2010

Human microsomal prostaglandin E synthase-1 (mPGES-1) catalyzes the conversion of prostaglandin $H_2$ ($PGH_2$) into prostaglandin $E_2$ ($PGE_2$). To establish a stable and efficient method to assess the activity of mPGES-1, a coupled enzyme assay system using mPGES-1, 15-hydroxyprostaglandin dehydrogenase (15-PGDH) and phosphomolybdic acid (PMA) was developed. In this assay system, $PGH_2$ was converted to $PGE_2$ by mPGES-1, and then $PGE_2$ was further transformed to the 15-keto-$PGE_2$ by 15-PGDH accompanying the production of NADH, which was easily detected by fluorescence spectrometry in a multi-well plate format. During the reaction, spontaneous oxidation of $PGH_2$ was prevented by PMA. Using this novel assay, the $K_m$ value of mPGES-1 for $PGH_2$ and the $IC_{50}$ value of the previously characterized inhibitor, MK-886, were determined to be 0.150 mM and $2.8\;{\mu}M$, respectively, which were consistent with the previously reported values. In addition, low backgrounds were observed in the multi-wall plate screening of chemical compounds.

• The Plant Pathology Journal
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• v.36 no.5
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• pp.497-502
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• 2020

Barley yellow dwarf virus (BYDV) is an economically important plant pathogen that causes stunted growth, delayed heading, leaf yellowing, and purple leaf tip, thereby reducing the yields of cereal crops worldwide. In the present study, a reverse transcription recombinase polymerase amplification (RT-RPA) assay was developed for the detection of BYDV in oat leaf samples. The RT-RPA assay involved incubation at an isothermal temperature (42℃) and could be performed rapidly in 5 min. In addition, no cross-reactivity was observed to occur with other cereal-infecting viruses, and the method was 100 times more sensitive than conventional reverse transcription polymerase chain reaction. Furthermore, the assay was validated for the detection of BYDV in both field-collected oat leaves and viruliferous aphids. Thus, the RT-RPA assay developed in the present study represents a simple, rapid, sensitive, and reliable method for detecting BYDV in oats.

• The Plant Pathology Journal
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• v.38 no.2
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• pp.159-166
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• 2022

Barley yellow dwarf virus (BYDV) has been a major viral pathogen causing significant losses of cereal crops including oats worldwide. It spreads naturally through aphids, and a rapid, specific, and reliable diagnostic method is imperative for disease monitoring and management. Here, we established a rapid and reliable method for isothermal reverse transcription recombinase polymerase amplification (RT-RPA) combined with a lateral flow strips (LFS) assay for the detection of BYDV-infected oat samples based on the conserved sequences of the BYDV coat protein gene. Specific primers and a probe for RT-RPA reacted and optimally incubated at 42℃ for 10 min, and the end-labeled amplification products were visualized on LFS within 10 min. The RT-RPA-LFS assay showed no cross-reactivity with other major cereal viruses, including barley mild mosaic virus, barley yellow mosaic virus, and rice black streaked dwarf virus, indicating high specificity of the assay. The sensitivity of the RT-RPA-LFS assay was similar to that of reverse transcription polymerase chain reaction, and it was successfully validated to detect BYDV in oat samples from six different regions and in individual aphids. These results confirm the outstanding potential of the RT-RPA-LFS assay for rapid detection of BYDV.

• Journal of Microbiology and Biotechnology
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• v.18 no.1
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• pp.118-123
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• 2008

A new method for antimicrobial susceptibility testing of vitro-cultured bacteria on an ordinary fluorescence spectrometer was developed. The viable bacteria reduced 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) to produce insoluble particles that displayed intense resonance scattering light. The assay showed a linear relationship between the number of viable bacteria and the intensity of resonance scattering light. Dead bacteria were unable to reduce MTT. Methicillin-resistant Staphylococcus aureus exposed to flavonoids from Marchantia convoluta showed a flavonoids concentration-dependent inhibition of the ability to reduce MTT. In the assay, less than 12 h was required to attain susceptibility results and fewer bacteria were utilized than in traditional methods. The RLS technique could, in combination with the MTT assay, be a rapid and sensitive measuring method to determine the in vitro activity of new antimicrobials.

• YAKHAK HOEJI
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• v.58 no.2
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• pp.125-128
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• 2014

We developed an alternative assay method without hazardous reagent of chloroform for urazamide tablets in Korean Pharmaceutical Codex. The HPLC analytical method was validated by system suitability, linearity, precision, accuracy and robustness. The linearity of the calibration curves in the desired concentration range is good ($r^2$ >0.999). Precision was obtained less than RSD 1.17%. Accuracy was obtained with recoveries in range of 98.12% and 99.47%. The developed assay could be expected to become valuable tools for revising the Korean Pharmaceutical Codex.

• Journal of Life Science
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• v.18 no.8
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• pp.1159-1163
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• 2008

The purpose of this study was to establish a high-throughput assay method capable of estimating specific lipase activity at oil-water interface. The method is based on the fact that fluorescence intensity of fluorescein is affected by pH. The pH-dependence might be used to monitor pH change caused by the release of fatty acid through the action of lipase. Assay was performed by incubating a reaction mixture containing oil emulsion, fluorescein and enzyme and by monitoring fluorescence intensity periodically. Fluorescence intensity decreased linearly with a rate proportional to the enzyme amount. Linear relationship was observed between enzyme amount and reaction rate which was calculated from a graph of fluorescence change against time. The assay was possible at different pH conditions in the range of pH 6.0-8.0.

• Parasites, Hosts and Diseases
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• v.59 no.1
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• pp.15-22
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• 2021

Artemisinin resistance (ART) has been confirmed in Greater Mekong Sub-region countries. Currently, C580Y mutation on Pfkelch13 gene is known as the molecular marker for the detection of ART. Rapid and accurate detection of ART in field study is essential to guide malaria containment and elimination interventions. A simple method for collection of malaria-infected blood is to spot the blood on filter paper and is fast and easy for transportation and storage in the field study. This study aims to evaluate LAMP-SNP assay for C580Y mutation detection by introducing an extra mismatched nucleotide at the 3' end of the FIP primer. The LAMP-SNP assay was performed in a water bath held at a temperature of 56℃ for 45 min. LAMP-SNP products were interpreted by both gel-electrophoresis and HNB-visualized changes in color. The method was then tested with 120 P. falciparum DNA from dried blood spot samples. In comparing the LAMP-SNP assay results with those from DNA sequencing of the clinical samples, the 2 results fully agreed to detect C580Y. The sensitivity and specificity of the LAMP-SNP assay showed 100%. There were no cross-reactions with other Plasmodium species and other Pfkelch13 mutations. The LAMP-SNP assay performed in this study was rapid, reliable, and useful in detecting artemisinin resistance in the field study.

• Journal of Applied Biological Chemistry
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• v.43 no.4
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• pp.230-234
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• 2000

A new method to assay the capsaicinoid synthetase (CS) activity was developed by utilizing NADHcoupled enzyme systems involving pyruvate kinase and lactate dehydrogenase. CS activities in Capsicum placenta, depending upon the kinetics of the NADH oxidation, revealed almost the same profile as compared with those shown using an HPLC-based method. When the substrates, 8-methyl nonanoic acid and vanillylamine, for the CS enzyme were employed separately or simultaneously, it appeared that the two-step reaction, acyl-CoA formation and condensation with vanillyla~ne, of the CS enzyme was a coupled reaction. Thus, this assay method of the CS enzyme can be considered as an alternative to the HPLC-based method, since it has the advantages of rapidity and simplicity as well as reliability when compared with the existing method.

• Bulletin of the Korean Chemical Society
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• v.31 no.9
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• pp.2557-2560
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• 2010

To assay for antioxidant capacity of natural products considered important in producing human health benefits, a practical and economical method using pellet techniques was developed. A standard visualizing reagent, 1,1diphenyl-2-picryl-hydrazyl (DPPH), was mixed with a water-miscible polyvinyl alcohol (PVA), serving as a solid phase support for the DPPH reagent. A DPPH pellet was prepared by dropping a small volume of the DPPH solution onto PET film, and drying in an oven. The PVA-based DPPH pellet was dissolved into water, in which the water-miscible PVA plays as a non-ionic surfactant to help the DPPH reagent to be dissolved into the solvent. Using the DPPH assay, the antioxidant capacity of water-soluble extracts of black soybean, barley, green tea, and green gram was examined. Among the natural products tested, green tea showed the highest antioxidant capacity. This PVA-based DPPH antioxidant assay can be further applied in the natural food, raw plant material, and health product inspection field.

• Biomedical Science Letters
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• v.10 no.3
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• pp.263-268
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• 2004

This study was carried out to evaluate the cytotoxic effect of phenolic compound on normal NIH 3T3 fibrolasts. The colorimetric assay for phenol compound, syringic acid was performed by MTT assay or XTT assay. MTT or XTT assays are known as a very sensitive method in measuring the cytotoxic effect of chemical agents in vitro. In the present study, syringic acid on normal Nlli 3T3 fibroblasts did not show any cytotoxicity for MTT assay or XTT assay compared with control after cells were treated with various concentrations of syringic acid for 48 hours. MTT/sub 50/ and XTT/sub 50/ were 3,340.9 μM and 2,462.4 μM of syringic acid, respectively. From the above the results, it is suggested that phenolic compound of syringic acid did not have any cytotoxicity on normal NIH 3T3 fibroblasts.