• Title/Summary/Keyword: Biological sample

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Cloning, expression, and activity of type IV antifreeze protein from cultured subtropical olive flounder (Paralichthys olivaceus)

  • Lee, Jong Kyu;Kim, Hak Jun
    • Fisheries and Aquatic Sciences
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    • v.19 no.8
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    • pp.33.1-33.7
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    • 2016
  • Antifreeze proteins (AFPs) lower the freezing point but not the melting point of aqueous solutions by inhibiting the growth of ice crystals via an adsorption-inhibition mechanism. However, the function of type IV AFP (AFP IV) is questionable, as its antifreeze activity is on the verge of detectable limits, its physiological concentration in adult fish blood is too low to function as a biological antifreeze, and its homologues are present even in fish from tropic oceans as well as freshwater. Therefore, we speculated that AFP IV may have gained antifreeze activity not by selective pressure but by chance. To test this hypothesis, we cloned, expressed, and assayed AFP IV from cultured subtropical olive flounder (Paralichthys olivaceus), which do not require antifreeze protein for survival. Among the identified expressed sequence tags of the flounder liver sample, a 5'-deleted complementary DNA (cDNA) sequence similar to the afp4 gene of the longhorn sculpin was identified, and its full-length cDNA and genome structure were examined. The deduced amino acid sequence of flounder AFP IV shared 55, 53, 52, and 49 % identity with those of Pleuragramma antarcticum, Myoxocephalus octodecemspinosus, Myoxocephalus scorpius, and Notothenia coriiceps, respectively. Furthermore, the genomic structure of this gene was conserved with those of other known AFP IVs. Notably, the recombinant AFP IV showed a weak but distinct thermal hysteresis of $0.07{\pm}0.01^{\circ}C$ at the concentration of 0.5 mg/mL, and ice crystals in an AFP IV solution grew star-shaped, which are very similar to those obtained from other polar AFP IVs. Taken together, our results do not support the hypothesis of evolution of AFP IV by selective pressure, suggesting that the antifreeze activity of AFP IV may have been gained by chance.

Evaluation of Morphological Changes in Degenerative Cartilage Using 3-D Optical Coherence Tomography

  • Youn, Jong-In
    • Journal of the Optical Society of Korea
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    • v.12 no.2
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    • pp.98-102
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    • 2008
  • Optical Coherence Tomography (OCT) is an important noninvasive medical imaging technique that can reveal subsurface structures of biological tissue. OCT has demonstrated a good correlation with histology in sufficient resolution to identify morphological changes in articular cartilage to differentiate normal through progressive stages of degenerative joint disease. Current OCT systems provide individual cross-sectional images that are representative of the tissue directly under the scanning beam, but they may not fully demonstrate the degree of degeneration occurring within a region of a joint surface. For a full understanding of the nature and degree of cartilage degeneration within a joint, multiple OCT images must be obtained and an overall assessment of the joint surmised from multiple individual images. This study presents frequency domain three-dimensional (3-D) OCT imaging of degenerative joint cartilage extracted from bovine knees. The 3-D OCT imaging of articular cartilage enables the assembly of 126 individual, adjacent, rapid scanned OCT images into a full 3-D image representation of the tissue scanned, or these may be viewed in a progression of successive individual two-dimensional (2-D) OCT images arranged in 3-D orientation. A fiber-based frequency domain OCT system that provides cross-sectional images was used to acquire 126 successive adjacent images for a sample volume of $6{\times}3.2{\times}2.5\;mm^3$. The axial resolution was $8\;{\mu}m$ in air. The 3-D OCT was able to demonstrate surface topography and subsurface disruption of articular cartilage consistent with the gross image as well as with histological cross-sections of the specimen. The 3-D OCT volumetric imaging of articular cartilage provides an enhanced appreciation and better understanding of regional degenerative joint disease than may be realized by individual 2-D OCT sectional images.

Multimodal Nonlinear Optical Microscopy for Simultaneous 3-D Label-Free and Immunofluorescence Imaging of Biological Samples

  • Park, Joo Hyun;Lee, Eun-Soo;Lee, Jae Yong;Lee, Eun Seong;Lee, Tae Geol;Kim, Se-Hwa;Lee, Sang-Won
    • Journal of the Optical Society of Korea
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    • v.18 no.5
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    • pp.551-557
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    • 2014
  • In this study, we demonstrated multimodal nonlinear optical (NLO) microscopy integrated simultaneously with two-photon excitation fluorescence (TPEF), second-harmonic generation (SHG), and coherent anti-Stokes Raman scattering (CARS) in order to obtain targeted cellular and label-free images in an immunofluorescence assay of the atherosclerotic aorta from apolipoprotein E-deficient mice. The multimodal NLO microscope used two laser systems: picosecond (ps) and femtosecond (fs) pulsed lasers. A pair of ps-pulsed lights served for CARS (817 nm and 1064 nm) and SHG (817 nm) images; light from the fs-pulsed laser with the center wavelength of 720 nm was incident into the sample to obtain autofluorescence and targeted molecular TPEF images for high efficiency of fluorescence intensity without cross-talk. For multicolor-targeted TPEF imaging, we stained smooth-muscle cells and macrophages with fluorescent dyes (Alexa Fluor 350 and Alexa Fluor 594) for an immunofluorescence assay. Each depth-sectioned image consisted of $512{\times}512$ pixels with a field of view of $250{\times}250{\mu}m^2$, a lateral resolution of $0.4{\mu}m$, and an axial resolution of $1.3{\mu}m$. We obtained composite multicolor images with conventional label-free NLO images and targeted TPEF images in atherosclerotic-plaque samples. Multicolor 3-D imaging of atherosclerotic-plaque structural and functional composition will be helpful for understanding the pathogenesis of cardiovascular disease.

Coherent X-ray Diffraction Imaging with Single-pulse Table-top Soft X-ray Laser

  • Kang, Hyon-Chol;Kim, H.T.;Lee, S.K.;Kim, C.M.;Choi, I.W.;Yu, T.J.;Sung, J.H.;Hafz, N.;Jeong, T.M.;Kang, S.W.;Jin, Y.Y.;Noh, Y.C.;Ko, D.K.;Kim, S.S.;Marathe, S.;Kim, S.N.;Kim, C.;Noh, D.Y.;Lee, J.
    • Proceedings of the Optical Society of Korea Conference
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    • 2008.02a
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    • pp.429-430
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    • 2008
  • We demonstrate coherent x-ray diffraction imaging using table-top x-ray laser at a wavelength of 13.9nm driven by 10-Hz ti:Sapphire laser system at the Advanced Photonics Research Institute in Korea. Since the flux of x-ray photons reaches as high as $10^9$ photons/pulse in a $20{\times}20{\mu}m^2$ field of view, we measured a ingle-pulse diffraction pattern of a micrometer-scale object with high dynamic range of diffraction intensities and successfully reconstructed to the image using phase retrieval algorithm with an oversampling ratio of 1:6. the imaging resolution is $^{\sim}150$ nm, while that is much improved by stacking the many diffraction patterns. This demonstration can be extended to the biological sample with the diffraction limited resolution.

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Quorum Quenching Bacteria Isolated from the Sludge of a Wastewater Treatment Plant and Their Application for Controlling Biofilm Formation

  • Kim, A-Leum;Park, Son-Young;Lee, Chi-Ho;Lee, Chung-Hak;Lee, Jung-Kee
    • Journal of Microbiology and Biotechnology
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    • v.24 no.11
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    • pp.1574-1582
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    • 2014
  • Bacteria recognize changes in their population density by sensing the concentration of signal molecules, N-acyl-homoserine lactones (AHLs). AHL-mediated quorum sensing (QS) plays a key role in biofilm formation, so the interference of QS, referred to as quorum quenching (QQ), has received a great deal of attention. A QQ strategy can be applied to membrane bioreactors (MBRs) for advanced wastewater treatment to control biofouling. To isolate QQ bacteria that can inhibit biofilm formation, we isolated diverse AHL-degrading bacteria from a laboratory-scale MBR and sludge from real wastewater treatment plants. A total of 225 AHL-degrading bacteria were isolated from the sludge sample by enrichment culture. Afipia sp., Acinetobacter sp. and Streptococcus sp. strains produced the intracellular QQ enzyme, whereas Pseudomonas sp., Micrococcus sp. and Staphylococcus sp. produced the extracellular QQ enzyme. In case of Microbacterium sp. and Rhodococcus sp., AHL-degrading activities were detected in the whole-cell assay and Rhodococcus sp. showed AHL-degrading activity in cell-free lysate as well. There has been no report for AHL-degrading capability in the case of Streptococcus sp. and Afipia sp. strains. Finally, inhibition of biofilm formation by isolated QQ bacteria or enzymes was observed on glass slides and 96-well microtiter plates using crystal violet staining. QQ strains or enzymes not only inhibited initial biofilm development but also reduced established biofilms.

HPLC Determination and Pharmacokinetics of Endogenous Acetyl-L-Carnitine (ALC) in Human Volunteers Orally Administered a Single Dose of ALC

  • Kwon, Oh-Seung;Chung, Youn-Bok
    • Archives of Pharmacal Research
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    • v.27 no.6
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    • pp.676-681
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    • 2004
  • Acetyl-L-camitine (ALC), a naturally occurring endogenous compound, has been shown to improve the cognitive performance of patients with senile dementia Alzheimer's type, and to be involved in cholinergic neurotransmission. Because ALC is an endogenous compound, valida-tion of the analytical methods of ALC in the biological fluids is very important and difficult. This study was presented validation and correction for plasma ALC concentrations and pharmacok-inetics after oral administration of ALC to human volunteers. ALC concentrations in human plasma were corrected by subtracting the concentration of blank plasma from each sample. Precision and accuracy (bias %) for uncorrected ALC concentrations were below 2.6 and 6.5% for intra-days, and 4.0 and 9.4% for inter-days, respectively. Precision and accuracy (bias %)for corrected ALC concentrations were below 10.9 and 6.0% for intra-days, and 10.5 and 16.9% for inter-days, respectively. Quantitation limit was $0.1{\;}\mu\textrm{g}/mL$. After oral administration of a 500 mg ALC tablet to 8 healthy volunteers, the principle pharmacokinetic parameters were 4.2 h of the half-life$ (t_{1/2},{\beta})$, the area under the curve $(AUC_{0{\rightarrow}8){\;}of{\;}9.88{\;}\mu\textrm{g}{\cdot}h/mL$, and 3.1 h of the time ($T_{max}$) to reach $C_{max}$. This study first describes the pharmacokinetic study after oral admin-istration of a single dose of ALC in human volunteers.

Preparation of Metal-p-aminobenzyl-DOTA Complex Using Magnetic Particles for Bio-tagging in Laser Ablation ICP-MS

  • Yoon, S.Y.;Lim, H.B.
    • Bulletin of the Korean Chemical Society
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    • v.33 no.11
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    • pp.3665-3670
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    • 2012
  • Metal-p-$NH_2$-Bn-DOTA (paraammionobenzyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid: ABDOTA) complex was synthesized and purified for bio-tagging to quantify biological target materials using laser ablation (LA)-ICP-MS. Since the preparation of a pure and stable tagging complex is the key procedure for quantification, magnetic particles were used to purify the synthesized metal-ABDOTA complex. The magnetic particles immobilized with the complex attracted to a permanent magnet, resulting in fast separation from free un-reacted metal ions in solution. Gd ions formed the metal-complex with a higher yield of 64.3% (${\pm}3.9%$ relative standard deviation (RSD)) than Y ions, 52.3% (${\pm}2.5%$ RSD), in the pH range 4-7. The complex bound to the magnetic particles was released by treatment with a strong base, of which the recovery was 81.7%. As a reference, a solid phase extraction (SPE) column packed with Chelex-100 resin was employed for separation under similar conditions and produced comparable results. The tagging technique complemented polydimethylsiloxane (PDMS) microarray chip sampling in LA-ICP-MS, allowing determination of small sample volumes at high throughputs. For application, immunoglobulin G (IgG) was immobilized on the pillars of PDMS microarray chips and then tagged with the prepared Gd complex. IgG could then be determined through measurement of Gd by LA-ICP-MS. A detection limit of 1.61 ng/mL (${\pm}0.75%$ RSD) for Gd was obtained.

Genetic Epidemiology of Renin-Angiotensin System in Korean Population

  • Kang, Byung-Yong;Bae, Joon-Seol;Kim, Ki-Tae;Oh, Ju-Hyung;Lee, Kang-Oh;Ryu, Jae-Chun;Kim, Jae-Hyoun;Oh, Sang-Duk;Yoon, Moon-Young;Lee, Chung-Choo
    • Environmental Mutagens and Carcinogens
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    • v.22 no.1
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    • pp.12-21
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    • 2002
  • Genetic polymorphisms of the renin-angiotensin system (RAS) have been associated with hypertension in various ethnic groups, but no relation between these polymorphisms and hypertension has yet been systematically evaluated. To assess the relationship between allelic variation of RAS genes and hypertension, we performed the case-control studies using genetic markers in Korean normotensives and hypertensives. The allele and genotype frequencies of RAS genes in Korean population were not significantly different between normotensives and hypertensives. To investigate the distribution of allele frequencies among various populations, the data obtained in this study were compared to those in other ethnic groups studied previously. Except for T174M polymorphism of angiotensinogen (AGT) gene, allele frequencies of RAS genes were different among racial groups. The reason for these differences may be due to the difference in various genetic or environmental background or due to the effects by various sample size studied. In addition, it can be emphasized that carefully designed studies are required to minimize the ethnic heterogeneity of the case and control populations.

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Assessment of Korean Water Quality Standards for Effluent Discharged from the Dye Industry Based on Acute Aquatic Toxicity Tests Using Microbes and Macroinvertebrates (염색폐수의 수질독성시험을 이용한 한국의 수질배출허용기준 평가연구)

  • Kim, Young-Hee;Lee, Min-Jung;Choi, Kyung-Ho;Eo, Soo-Mi;Lee, Hong-Keun
    • Journal of Environmental Health Sciences
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    • v.30 no.3
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    • pp.185-190
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    • 2004
  • Acute aquatic toxicity of effluents discharged from five dyeing plants in Gyeong-gi province were evaluated to assess whether the current Korean water quality standards(KWQS) could protect aquatic life. Chemical analyses of all parameters regulated under KWQS, except for E-coli, were also carried out to determine regulation compliance of the samples. All the effluent samples were satisfied with KWQS except for the color in only one sample. In acute Daphnia magna toxicity tests, significant mortality was observed in one of five samples and EC50 was 12.1%(95% confidence interval 9.1-16.2), which was in compliance with KWQS. The result of the Microtox assay indicated that acute microbial toxicity existed in effluents from three out of five plants, two of which were in compliance with KWQS. The agreement between regulation compliance of chemical concentrations of effluent and observed toxicity from various biological toxicity tests was very poor to fair (kappa = 0.194~0.250). The data presented suggest that exposure to dyeing wastewater which were in compliance with Korean water quality standards may not be safe to aquatic biota, and multiple tropical levels should be considered in aquatic toxicity monitoring of dyeing industry.

The Nitrite-Scavenging Effects by Component of Green Tea Extracts (녹차 추출물의 아질산염 분해작용)

  • Yeo, Saeng-Gyu;Yeum, Dong-Min;Lee, Dong-Ho;Ahn, Cheol-Woo;Kim, Seon-Bong;Park, Yeung-Ho
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.23 no.2
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    • pp.287-292
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    • 1994
  • This paper was investigated to elucidate the nitrite-scavenging action as one of biological actions of green tea extracts. Green tea extracts exhibited remarkable nitrite-scavenging action. Among the green tea extracts, steamed green tea showed higher nitrite-scavenging action than roasted green tea. Nitrite-savenging actions of gren tea extracts were hardly influenced until three times extraction with hot water. Furthermore , the nitirte-scavenging action of green tea extracts showed ph dependent , highest at pH 1.2 and lowest at pH 6.0. The nitrite -scavenging rate in sample extracts /amine /nitrite systems exhibited faster than that in amine/nitrite systems. In order to screen the nitirte-scavenging factors, samples extracts were fractionated with methanol. in both samples, methanol-soluble fractions possessed greater nitrite-scavenging action than methanol-insoluble fractions .The nitrite-scavenging action of methanol-soluble fractions increased with proportion to absorbance at 420nm and at 280nm, while that of green tea extracts increased with the contents of total phenols and absorbance at 280nm.

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