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Lee, Hyungbeen,Lee, Sang Won,Lee, Gyudo,Lee, Wonseok,Nam, Kihwan,Lee, Jeong Hoon,Hwang, Kyo Seon,Yang, Jaemoon,Lee, Hyeyoung,Kim, Sangsig,Lee, Sang Woo,Yoon, Dae Sung The Royal Society of Chemistry 2018 Nanoscale Vol.10 No.2
<P>Here, we demonstrate a powerful method to discriminate DNA mismatches at single-nucleotide resolution from 0 to 5 mismatches (<I>χ</I>0 to <I>χ</I>5) using Kelvin probe force microscopy (KPFM). Using our previously developed method, we quantified the surface potentials (SPs) of individual DNA-capped nanoparticles (DCNPs, ∼100 nm). On each DCNP, DNA hybridization occurs between ∼2200 immobilized probe DNA (<I>p</I>DNA) and target DNA with mismatches (<I>t</I>DNA, ∼80 nM). Thus, each DCNP used in the bioassay (each <I>p</I>DNA-<I>t</I>DNA interaction) corresponds to a single ensemble in which a large number of <I>p</I>DNA-<I>t</I>DNA interactions take place. Moreover, one KPFM image can scan at least dozens of ensembles, which allows statistical analysis (<I>i.e.</I>, an ensemble average) of many bioassay cases (ensembles) under the same conditions. We found that as the <I>χ</I>n increased from <I>χ</I>0 to <I>χ</I>5 in the tDNA, the average SP of dozens of ensembles (DCNPs) was attenuated owing to fewer hybridization events between the <I>p</I>DNA and the <I>t</I>DNA. Remarkably, the SP attenuation <I>vs.</I> the <I>χ</I>n showed an inverse-linear correlation, albeit the equilibrium constant for DNA hybridization exponentially decreased asymptotically as the <I>χ</I>n increased. In addition, we observed a cascade reaction at a 100-fold lower concentration of <I>t</I>DNA (∼0.8 nM); the average SP of DCNPs exhibited no significant decrease but rather split into two separate states (no-hybridization <I>vs.</I> full-hybridization). Compared to complementary <I>t</I>DNA (<I>i.e.</I>, <I>χ</I>0), the ratio of no-hybridization/full-hybridization within a given set of DCNPs became ∼1.6 times higher in the presence of tDNA with single mismatches (<I>i.e.</I>, <I>χ</I>1). The results imply that our method opens new avenues not only in the research on the DNA hybridization mechanism in the presence of DNA mismatches but also in the development of a robust technology for DNA mismatch detection.</P>
Development of an infant formula certified reference material for the analysis of organic nutrients
Lee, Joonhee,Kim, Byungjoo,Lee, Sun Young,Choi, Jongoh,Kang, Dukjin,Lee, Hwasim,Choi, KiHwan,Lee, Hyeyoung,Sim, Hee-Jung,Baek, Song-Yee,Lee, Honghee,Hyung, Seok-Won,Ahn, Seonghee,Seo, Dongwon,Hwang, J Applied Science Publishers 2019 Food chemistry Vol.298 No.-
<P><B>Abstract</B></P> <P>Infant formula certified reference material (CRM, KRISS CRM 108-02-003) were developed for the analysis of organic nutrients. The CRM is a milk-based infant formula powder, packaged at 14 g per unit. Ten thousand units were prepared and stored at −70 °C. For the certification of each nutrient, ten units were analyzed for simultaneous value-assignment and homogeneity test. Analytical methods used were isotope dilution mass spectrometry (IDMS) based on liquid chromatography mass spectrometer (LC/MS) or gas chromatography mass spectrometer (GC/MS) as higher-order reference methods.13 vitamins, 3 fatty acids, and total cholesterol were certified. The between-unit relative standard deviation of measurement results for each nutrient ranged 0.2% to 2.5%, showing very good homogeneity. The expanded relative uncertainties of the certified values ranged from 1% to 8%, indicating that they have higher-order metrological quality. The values of proximates (proteins, lipids, carbohydrates, water, and ash) were assigned through inter-laboratory comparisons.</P> <P><B>Highlights</B></P> <P> <UL> <LI> An infant formula CRMs for the analysis of organic nutrients was developed. </LI> <LI> Organic nutrients were certified by IDMS approaches as higher-order reference methods. </LI> <LI> Homogeneities and stability of the CRM were evaluated by IDMS approaches. </LI> <LI> Metrological qualities of the certified values were proved by their small uncertainties. </LI> <LI> Five proximates were value-assigned by interlaboratory comparison. </LI> </UL> </P>
Seok, Seung-Hyeok,Koo, Hye Cheong,Kasuga, Asako,Kim, Yeun,Lee, Eun Gae,Lee, Hyeyoung,Park, Jong-Hwan,Baek, Min-Won,Lee, Hui-Young,Kim, Dong-Jae,Lee, Byeung-Hee,Lee, Yong-Soon,Cho, Sang-Nae,Park, Jae-H Elsevier 2006 Veterinary microbiology Vol.114 No.3
<P><B>Abstract</B></P><P>Skin ulcers, scoliosis, and dropsy-like scale edema were observed in laboratory-maintained zebrafish. Affected fish had multifocal granulomas not only in internal organs such as the liver, intestine, genital organs, kidney, muscle, and spleen but also in the fin, epithelium, gills, and sclera of the eyes. Large numbers of acid-fast-rod-shaped bacteria were observed within the necrotic centers of well-demarcated, multifocal granulomas with Gram's stain and Ziehl–Neelson's stain. The size of the <I>Mycobacterium</I> spp. was 1–2μm×2–3μm with a double-layered cell wall, based upon electron-microscopical features. Definitive diagnosis of these outbreaks was obtained by culture on selective media followed by PCR-restriction fragment length polymorphism analysis (PRA) of the <I>rpoB</I> gene for species identification. The amplified 360-bp products of the <I>rpoB</I> gene of mycobacteria isolated from zebrafish were digested with <I>Msp</I>I restriction enzyme, which revealed unique band patterns matching those of <I>Mycobacterium abscessus</I> and <I>Mycobacterium chelonae</I> which are responsible for skin and soft tissue infection caused by rapidly growing mycobacteria in humans. This is the first documentation of the precise identification of zoonotic non-tuberculous mycobacteria isolated from laboratory-maintained zebrafish by the PRA of the <I>rpoB</I> gene; this study thus provides a great deal of useful epidemiological information and reduces the likelihood that epizootics will occur.</P>
Lee, Hyeyoung,Lee, Joonhee,Choi, Kihwan,Kim, Byungjoo Applied Science Publishers 2017 Food chemistry Vol.221 No.-
<P><B>Abstract</B></P> <P>A method based on isotope dilution-liquid chromatography/tandem mass spectrometry (ID-LC/MS/MS) using a C30 column has been developed for the separate and accurate determination of <I>trans</I>- and <I>cis</I>-vitamin K<SUB>1</SUB> in infant formula. Vitamin K<SUB>1</SUB> and the deuterium-labeled internal standard eluted at slightly different retention times experiencing different matrix effects, and this possibly resulted in biased measurement. The matrix effect profiles obtained from post-column infusion experiments showed that atmospheric pressure chemical ionization (APCI) was less susceptible to matrix effects near the retention time than electrospray ionization (ESI); therefore, APCI was used in this study. The developed method was validated by measuring fortified samples, and the results agreed with the gravimetric values. Its repeatability and reproducibly were within 2% relative standard deviation. The relative expanded uncertainty was approximately 5%, indicating that the method was of higher-order metrological quality as a reference method.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Application of ID-LC/MS/MS for the accurate determination of vitamin K<SUB>1</SUB> isomers. </LI> <LI> Quantification of <I>trans</I>- and <I>cis</I>-vitamin K<SUB>1</SUB> in the primary reference material by NMR. </LI> <LI> Minimization of the measurement bias using APCI rather than ESI. </LI> <LI> Developed method was proven to have high-order metrological quality. </LI> </UL> </P>
Lee, Hyeyoung,Nobrega de Moura Bell, Juliana Maria Leite,Barile, Daniela American Chemical Society 2019 Journal of agricultural and food chemistry Vol.67 No.12
<P>Bovine milk oligosaccharides (BMOs) that resemble human milk oligosaccharides are found in whey permeate, indicating that dairy streams can be used as a potential source of bioactive oligosaccharides. Recovery of oligosaccharides from whey permeate is hindered by their low abundance and high concentration of lactose. In the present work, lactose in bovine colostrum whey permeate was hydrolyzed by <I>Aspergillus oryzae</I> β-galactosidase to facilitate subsequent monosaccharide removal by membrane separation. Chromatographic separation coupled with high-resolution mass spectrometry revealed β-galactosidase degradation of several β-linkage-containing BMOs and production of novel oligosaccharides that ranged in size from 5 to 11 monosaccharide units containing several galactose repeating units and <I>N</I>-acetylhexosamine at their reducing ends. Optimization of BMO hydrolysis and separation methodology could generate high amounts of hetero-oligosaccharides for improved recovery of potentially biotherapeutic oligosaccharides.</P> [FIG OMISSION]</BR>
HiComet: a high-throughput comet analysis tool for large-scale DNA damage assessment
Lee, Taehoon,Lee, Sungmin,Sim, Woo Young,Jung, Yu Mi,Han, Sunmi,Won, Joong-Ho,Min, Hyeyoung,Yoon, Sungroh BioMed Central 2018 BMC bioinformatics Vol.19 No.-
<P><B>Background</B></P><P>DNA damage causes aging, cancer, and other serious diseases. The comet assay can detect multiple types of DNA lesions with high sensitivity, and it has been widely applied. Although comet assay platforms have improved the limited throughput and reproducibility of traditional assays in recent times, analyzing large quantities of comet data often requires a tremendous human effort. To overcome this challenge, we proposed HiComet, a computational tool that can rapidly recognize and characterize a large number of comets, using little user intervention.</P><P><B>Results</B></P><P>We tested HiComet with real data from 35 high-throughput comet assay experiments, with over 700 comets in total. The proposed method provided unprecedented levels of performance as an automated comet recognition tool in terms of robustness (measured by precision and recall) and throughput.</P><P><B>Conclusions</B></P><P>HiComet is an automated tool for high-throughput comet-assay analysis and could significantly facilitate characterization of individual comets by accelerating its most rate-limiting step. An online implementation of HiComet is freely available at https://github.com/taehoonlee/HiComet/.</P>