• Title/Summary/Keyword: Fluorescence polarization

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Real Time Measurement of Protease Activity of Live Uronema marinum (Ciliata: Scuticociliatida) by Fluorescence Polarization Assay

  • Lee Eun Hye;Kwon Se Ryun;Kim Chun Soo;Chung Joon Ki;Lee Hyung Ho;Kim Ki Hong
    • Fisheries and Aquatic Sciences
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    • v.5 no.4
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    • pp.311-313
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    • 2002
  • Proteolytic activity of live Uronema manum was analyzed by fluorescence polarization (FP) technique. Protease activity was measured by a decrease in FP value using fluorescein isothiocynate (FITC)-casein as a protein substrate. The results demonstrated an inverse linear relationship between fluorescence polarization (FP) values and live ciliate concentration over the range $1\times10^4\;to\;2\times10^5$ cells/well. However, the FP values of $10-10^3$ live parasites were not different significantly from that of control. Time-dependent decrease in FP value was shown in the wells containing live U. marinum. In the present study, FP assay had the benefit to provide measurements of substrate hydrolysis by live parasites in real-time, and did not require separations, precipitations, or transfers of reaction mixture.

Oligomerization State of the Plasma Membrane Proteolipid Apoprotein Purified from the Bovine Kidney, Probed by the Fluorescence Polarization

  • Chae, Quae;Nam, Sang-Rye
    • Bulletin of the Korean Chemical Society
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    • v.9 no.4
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    • pp.202-206
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    • 1988
  • In order to investigate the oligomerization state of the plasma membrane proteolipid apoprotein purified from the bovine kidney, fluorescence polarization experiment was carried out in the two different solvent systems, i.e., water and organic solvent(chloroform-methanol). The molecular volumes of the proteins estimated from the Perrin equation, were to be 45,258$A^3$ and 17,608$A^3$ in water and organic solvent, respectively. These values indicate that a trimerization is possibly occurring in the aqueous environment. As an auxiliary experiment for the calculation of the molecular volume using Perrin equation, fluorescence quenching constants ($K_q$) with the quencher acrylamide and fluorescence lifetimes (${\tau}_F$) of the intrinsic fluorophore tryptophan residue were estimated in the two different solvent systems. $K_q$ in water was 18.21$M^{-1}$ and it was 46.24$M^{-1}$ in organic solvent. Fluorescence lifetimes of tryptophan residue were calculated to be 2.80 nsec. in water and 3.81 nsec. in organic solvent, respectively.

QUANTITATION OF BARBITURATES IN URINE BY GC/MS AND ITS COMPARISON TO FLUORESCENCE POLARIZATION IMMUNOASSAY

  • Choo, Hea-Young;Park, Jeongeun;Park, Myung-Ja
    • Toxicological Research
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    • v.7 no.1
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    • pp.29-35
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    • 1991
  • Barbiturates are commonly abused tranquilizer and a rapid method to determine these drugs in biological samples is needed. In this study, was screened barbiturates in urine specimens by the fluorescence polarization immunoassay method(FPIA) and the positive samples were confirmed and identified by the more definitive GC/MS method. Fifteen positive smples which have barbiturate values higher than 0.5 ng/ml were analyzed by the GC/MS method. Eight samples were identified as phenobarbital and five samples were identified as crotilbarbitone.

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Effect of iron on the proteolytic activity of live Uronema marinum (Ciliata: Scoticociliatida) measured by fluorescence polarization

  • Lee, Eun-Hye;Kwon, Se-Ryeon;Choi, Seung-Hyuk;Kim, Ki-Hong
    • Journal of fish pathology
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    • v.19 no.1
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    • pp.83-86
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    • 2006
  • Effect of iron on the extracellular proteolytic activity of live Uronema marium was determined by fluorescence polarization (FP) method. Supplementation of 0.5 and 5.0 μM iron significantly increased caseinolytic activity of live U. marinum. In contrast, supplementation of 50 μM iron showed no significant differences in FP values compared to the control. The present result suggests that iron in cultured water or skin tissue of olive flounder may influence on the penetration and establishment of U. marinum, correlating with modulation of extracellular protease activity of the ciliates.

Label-Free and Real-Time Monitoring of Phosphatase Reactions Using a Phosphate-Specific and Fluorescent Probe

  • Lee, Ji-Hoon;Ahn, Hee-Chul;Shin, Dong-Yun;Ahn, Dae-Ro
    • Bulletin of the Korean Chemical Society
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    • v.29 no.5
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    • pp.943-947
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    • 2008
  • A phosphate-specific and fluorescent probe was prepared for label-free phosphatase assays based on fluorescence polarization. By using the probe, dephosphorylation reactions of DNA and protein substrates by calf intestinal alkaline phosphatase (CIP) could effectively be monitored in real-time. Since this assay method does not require additional materials such as labeled substrates and phosphospecific antibodies to obtain fluorescence polarization signals, it is simple, cost-effective, and expected to be useful not only for measuring activity of phosphatases but also for high-throughput screening of phosphatase inhibitors.

Diffusion Coefficients of CdSe/CdS Quantum Rods in Water Measured Using Polarized Fluorescence Correlation Spectroscopy

  • Lee, Jaeran;Pack, Chan-Gi;Kim, Soo Yong;Kim, Sok Won
    • Journal of the Optical Society of Korea
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    • v.18 no.5
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    • pp.598-604
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    • 2014
  • A polarization fluorescence correlation spectroscopy system based on a confocal microscope was built to study the rotational and translational diffusion of CdSe/CdS quantum rods (Q-rods), with the same and different polarization states between the polarizer and the analyzer (i.e. the XXX and XYY states). The rotational diffusion amplitude showed the dependences on polarization of $0.75{\pm}0.05$ in the XXX state and $0.26{\pm}0.03$ in the XYY state, when the translational diffusion amplitude was 1. The diffusion coefficients of the Q-rods were found based on their translational and rotational diffusion times in the two polarization states, in solutions with viscosity ranging from 0.9 to 6.9 cP. The translational and rotational diffusion coefficients ranged from $1.5{\times}10^{-11}$ to $2.6{\times}10^{-12}m^2s^{-1}$ and from $2.9{\times}10^5$ to $5.6{\times}10^4s^{-1}$, respectively.

Measurement of membrane fluidity of rockfish (Sebastes schlegeli) phagocytes during the respiratory burst using fluorescence polarization assay

  • Jung, Jae-Hyuck;Kwon, Se-Ryun;Lee, Eun-Hye;Kim, Sung-Mi;Kim, Ki-Hong
    • Journal of fish pathology
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    • v.16 no.2
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    • pp.131-134
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    • 2003
  • The change of membrane fluidity in rockfish (Sebastes schlegeli) phagocytes during respiratory burst was investigated. Fluorescence polarization (FP) was used as a measure of membrane fluidity, and 1-(4-trimethylaminophenyl)-6-phenyl-1 .3 ,5-hexatriene (TMA.-DPH) was used us a fluorescent probe. The significantly higher FP values in phagocytes stimulated With zymosan or phurbol myristate acetate (PMA) than unstimulated control phagocytes suggests that membrane fluidity of phagocytcs is decreased during the respiratory burst. The faster decrease of FP value in PMA stimulated phagocytes than in zymosan sumulated phagocytes may be due to bypass of the receptor-mediated stages of functional modulation. which is needed in zymosan stimulated phagocytes.

Transdermal permeation-enhancing activity of N-adamantyl n-alkanamides for lbuprofen in the rabbit

  • Han, Suk-Kyu;Park, Yong-Hoon;Ko, Young-Ill;Kim, Young-Mi
    • Archives of Pharmacal Research
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    • v.19 no.2
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    • pp.95-99
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    • 1996
  • Four N-adamantyl n-alkanamides were prepared by amide condensation reaction between amantadine and n-alkanoic acid. Their enhancing activity on the penetration of ibuprofen through rabbit skin from petrolatum ointment was evaluated in in-vivo experiment. The experiments showed that the compounds have a strong transdermal penetration-enhancing activity, and their activities were comparable with that of Azone. The measurements of the fluorescence polarization of DP[-i-labelled DPPC liposomes showed that these compounds considerablly decreased the phase transition temperature of the liposomes. The mechanism of the transdermal penetration-enhancing activity of the compounds was ascribed to the reduction of the resistance to drug flux of the stratum corneum lipid layers due to the loose packing of the layers when the bulk head group of the enhancers inserts into the layers.

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Effects of Lipid Composition on the Properties of Phospholipid Liposomal Membranes (리포솜 지질막의 성질에 미치는 지질 조성의 영향)

  • Kim, Min;Han, Suk-Kyu;Kim, Chong-Kook
    • YAKHAK HOEJI
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    • v.38 no.2
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    • pp.131-139
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    • 1994
  • Calcein-encapsulated small unilamellar vesicles of various lipid composition were prepared using the sonication technique, and their stabilities at $20^{\circ}C$ were examined by measuring calcein leakage from the liposomes. The fluidity of these liposomal bilayers was also investigated by measuring the fluorescence polarization of DPH labelled into the liposomes. The results showed that liposomes made of PC mixtures with different acyl chain length were very stable, which may be due to the formation of interdigitated bilayer structure. The addition of cholesterol further stabilized these PC liposomes. However, addition of cholesterol reduced the encapsulation efficiences of liposomes. The fluidity of the liposomes was significantly decreased by cholesterol in the liquid crystalline state, but not changed in the gel state. These results suggest that the enhanced stability of PC mixture liposomes may be ascribed to the formation of stable interdigitated bilayer structure. In membrane-mimetic and drug-delivery studies, vesicles made of mixtures of various phospholipids are recommended instead of addition of cholesterol to the phospholipid.

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