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The Reactivity of Antiserum Raised against Native Glucose-6-phosphate Dehydrogenase with Denatured Glucose-6-phosphate Dehydrogenase in Competitive ELISA

  • Kim, Moon-Hee
    • BMB Reports
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    • v.31 no.5
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    • pp.519-523
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    • 1998
  • We have previously reported that anti-glucose-6-phosphate dehydrogenase (G6PD) serum raised against native G6PD (nG6PD) enzyme recognized nG6PD antigen poorly in competitive enzyme-linked immunosorbent assay (ELISA) (Kim, 1997). In the present study, we investigated whether anti-G6PD serum raised against nG6PD can react with denatured G6PD effectively in competitive ELISA. We used partially active G6PD (paG6PD) by repeated freeze-thawing or SDS-denatured G6PD (SDS-G6PD) as both immobilized and soluble antigens, and anti-G6PD serum raised against nG6PD for competitive ELISA. The polystyrene cuvettes coated with either paG6PD or SDS-G6PD were challenged with a mixture of a limiting amount of anti-G6PD serum and various doses of paG6PD or SDS-G6PD as competitors, followed by incubation with alkaline phosphatase-anti-IgG conjugate. The competitive ELISA with paG6PD or SDS-G6PD antigen exhibited the sigmoidal dose-response curve characteristic of competition immunoassays. Furthermore, Triton-denatured G6PD (Triton-G6PD) was used in competitive ELISA. The paG6PD, SDS-G6PD, or Triton-G6PD used as competitors increased the inhibition of antibody binding to immobilized either of nG6PD or denatured G6PD compared with nG6PD competitor. The inhibition by denatured G6PD competitors was more pronounced at high competitor concentrations than at low counterparts. We conclude that anti-G6PD serum raised against nG6PD can effectively react with denatured G6PD in competitive ELISA and that our anti-G6PD serum recognizes denatured enzymes better than active enzymes.

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Competitive Enzyme-Linked Immunosorbent Assay for Glucose-6-Phosphate Dehydrogenase

  • Kim, Moon-Hee
    • BMB Reports
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    • v.30 no.5
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    • pp.326-331
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    • 1997
  • To construct a competitive ELISA standard curve for the detection of glucose-6-phosphate debydrogenase (G6PD), we used highly purified native G6PD (nG6PD) as both immobilized and soluble antigens and anti-G6PD serum raised against nG6PD as antibody. The polystyrene cuvettes coated with nG6PD were challenged with a mixture of a limiting amount of anti-G6PD serum and various doses of nG6PD as competitors followed by incubation with alkaline phosphatase-anti-IgG conjugate. The competitive ELISA did not exhibit the typical sigmoidal dose-response curve characteristic of competition immunoassays under the optimal concentrations of antigen and antibody. The soluble nG6PD used as competitor failed to effectively inhibit the binding of antibodies to the immobilized nG6PD. The addition of NADP, a cofactor of G6PD enzyme, to coating buffer used for immobilizing nG6PD to the cuvettes and PBS-Tween-BSA buffer for diluting competitors did not improve the inhibition of antibody binding to immobilized nG6PD by soluble n/G6PD. The addition of BSA to coating buffer did not increase inhibition, either. Surprisingly, when partially active G6PD (paG6PD), obtained by repeated freeze-thawing, was used as competitor, the antibody binding to either immobilized nG6PD or immobilized paG6PD was inhibited 49-58%. We conclude that an effective competitive ELISA system with nG6PD enzyme and anti-G6PD serum for the detection of G6PD may not be established due to the poor inhibition of antibody binding to immobilized nG6PD by soluble nG6PD under the present assay conditions and that the inhibition may be improved by using an inactivated enzyme as competitor regardless of the type of immobilized antigen used. These results imply that the immobilized nG6PD may undergo denaturation upon binding to the polystyrene cuvettes and that our anti-G6PD serum may recognize denatured enzyme better than active enzyme.

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Isoforms of Glucose 6-Phosphate Dehydrogenase in Deinococcus radiophilus

  • Sung, Ji-Youn;Lee, Young-Nam
    • Journal of Microbiology
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    • v.45 no.4
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    • pp.318-325
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    • 2007
  • Glucose 6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49) in Deinococcus radiophilus, an extraordinarily UV-resistant bacterium, was investigated to gain insight into its resistance as it was shown to be involved in a scavenging system of superoxide $(O_2^{-1})$ and peroxide $(O_2^{-2})$ generated by UV and oxidative stresses. D. radiophilus possesses two G6PDH isoforms: G6PDH-1 and G6PDH-2, both showing dual coenzyme specificity for NAD and NADP. Both enzymes were detected throughout the growth phase; however, the substantial increase in G6PDH-1 observed at stationary phase or as the results of external oxidative stress indicates that this enzyme is inducible under stressful environmental conditions. The G6PDH-1 and G6PDH-2 were purified 122- and 44-fold (using NADP as cofactor), respectively. The purified G6PDH-1 and G6PDH-2 had the specific activity of 2,890 and 1,033 U/mg protein (using NADP as cofactor) and 3,078 and 1,076 U/mg protein (using NAD as cofactor), respectively. The isoforms also evidenced distinct structures; G6PDH-1 was a tetramer of 35 kDa subunits, whereas G6PDH-2 was a dimer of 60kDa subunits. The pIs of G6PDH-1 and G6PDH-2 were 6.4 and 5.7, respectively. Both G6PDH-1 and G6PDH-2 were inhibited by both ATP and oleic acid, but G6PDH-1 was found to be more susceptible to oleic acid than G6PDH-2. The profound inhibition of both enzymes by ${\beta}-naphthoquinone-4-sulfonic$ acid suggests the involvement of lysine at their active sites. $Cu^{2+}$ was a potent inhibitor to G6PDH-2, but a lesser degree to G6PDH-1. Both G6PDH-1 and G6PDH-2 showed an optimum activity at pH 8.0 and $30^{\circ}C$.

A study on Enhancement Effectiveness of Cyanoacrylate Fumed Fingermark by the Solvent of Rhodamine 6G (Rhodamine 6G 용매에 따른 CA 훈증 지문 증강 효과에 관한 연구)

  • Shim, Yea-Ra;Yu, Je-Seol
    • The Journal of the Korea Contents Association
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    • v.17 no.7
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    • pp.294-302
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    • 2017
  • In this study, latent fingermarks deposited in porous or non-porous surface was developed by cyanoacrylate fuming, and then the developed fingermark is enhanced by using Rhodamine 6G. Between water-based R6G and organic solvent-based R6G, author studied about which material have higher effectiveness in enhancing fingermark. In all seven types of surfaces depositing fingermark, water-based R6G have higher effectiveness in enhancing fingermark and lower surface coloring than organic solvent-based R6G. But because the surfaces found in crime scene have multicolor background and various quality, the additional study about various surfaces is needed.

Effect of Molecular Weight of Chitosan on Flocculation of Suspended Solids in Rhodamine 6G-Sodium Dodecyl Sulfate System (키토산 분자량에 따른 Rhodamine 6G-Sodium Dodecyl Sulfate계의 응집효과에 관한 연구)

  • Kim, Sung Hyun;Nah, Jae Woon;Oh, Jeong Sun;Song, Ki Dong
    • Journal of the Korean Chemical Society
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    • v.42 no.6
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    • pp.672-678
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    • 1998
  • Chitin was extracted from crab shell of Portuns triberculatus and deacethylated to yield chitosan with various molecular weights. The absorption and the fluorescence spectra of Rhodamine 6G(Rh 6G)-sodium dodecyl sulfate(SDS) and Rh 6G-chitosan systems were obtained. From the spectra, we observed that the absorption and the fluorescence intensity of Rh 6G-SDS system decreased when S/D(the concentration of SDS to that of Rh 6G ratio) was below or at 32, while they increased when S/D was above 32. From the suspended solid(SS) removal rate and the transmittance of Rh 6G-SDS-chitosan system, we found that when S/D ratio was 32 its flocculating behaviour was much stronger than Rh 6G-SDS system. As the concentration and the molecular weight of chitosan increased, we also found that S/D range was extended from 32 to 100. With increasing the molecular weight of chitosan, the SS removal rate increased around pH 2~9 but decreased remarkably at pH>12.

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Properties of PA 6,6/Elastomer Blends with Various Elastomer Content (탄성중합체 함유량 변화에 따른 폴리아미드-6,6/탄성중합체 블랜드물의 특성)

  • Lee, Yoong;Lee, Chang-Woo;Cho, Yoon-Ho;Hahm, Yeong-Min
    • Elastomers and Composites
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    • v.34 no.1
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    • pp.20-30
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    • 1999
  • In this study, binary polyamide 6,6(PA 6,6)/ethylene-propylene rubber(EPM) or EPM-g-maleic anhydride(EPM-g-MA) blends and ternary PA 6,6/EPM/EPM-g-MA blends with various elastomer content were prepared in order to investigate the degree of influence of elastomer content and average particle size, morphology, and distribution of dispersed elastomer on mechanical and thermal properties of blends. According to the results, notched Izod impact strength and relative crystallinity of binary blends modified with EPM-g-MA as well as average particle size and distribution of dispersed elastomer in such blends were more improved than those of binary blends modified with EPM. Notched Izod impact strength of blend whose composition ratio(wt % ) was 70:30(PA 6,6 : EPM-g-MA) was the highest among the binary PA 6,6/EPM-g-MA blends. The impact strength was increased by 25 times and its relative crystallinity was increased by 7 times when compared with those of polyamide 6,6. In the case of ternary PA 6,6/EPM/EPM-g-MA blend of which composition ratio was 70:15:15(PA 6,6:EPM:EPM-g-MA), the elastomer was finely distributed with the average particle size of $0.56{\mu}m$. The Izod impact strength of this blend was the highest of all blends prepared with different elastomer content.

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Output Enhancement of Coaxial Flashlamp Pumped Rhodamine 6G Dye Laser by Energy Transfer (에너지 전환에 의한 동축 섬광관 펌핑 Rhodamine 6G 색소 레이저의 출력 증가)

  • 장우권
    • Korean Journal of Optics and Photonics
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    • v.4 no.2
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    • pp.195-199
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    • 1993
  • The output energy of Rh-6G dye laser was enhanced by the energy transfer in the mixture of Rh-6G and C-545. The laser was pumped by coaxial flashlamp filled argon gas. The optimum concentration of Rh-6G was $10_-4mol/l$ without mixing. The output energy was enhanced about 70 % at 0.4 % C-545 mixture with respect to the concentration of Rh-6G. The peak output power and the output energy were 27 kW and 50 mJ at the pumping energy of 346 J.

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Monitoring on preparation properties of Bokbunja (Rubus coreanus Miquel) granule (복분자 과립차 가공특성 모니터링)

  • Lee, Gee-Dong
    • Food Science and Preservation
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    • v.20 no.4
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    • pp.532-538
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    • 2013
  • This study was done in order to monitor the quality properties of the granule using Bokbunja (Rubus coreanus Miquel) extracts. In order to prepare the granule depending on operational parameters such as content of Bokbunja extract ($X_1$, 0.4~1.2 g), sugar content ($X_2$, 6~10 g) and citric acid content ($X_3$, 0.1~0.3 g), a response surface methodology was applied to monitor the optimum recipes on the organoleptic properties and Hunter's color. The optimum recipe on the organoleptic color showed extract content of 0.96 g, sugar content of 7.05 g and citric acid content of 0.232 g. The optimum recipe on the organoleptic flavor showed extract content of 0.86 g, sugar content of 6.04 g and citric acid content of 0.215 g. The optimum recipe on the organoleptic taste showed extract content of 0.92 g, sugar content of 6.39 g and citric acid content of 0.251 g. The optimum recipe on the overall palatability showed extract content of 0.86 g, sugar content of 6.65 g and citric acid content of 0.272 g. The response surface of the Hunter's color b value was similar to the response of the overall palatability; therefore, the optimum conditions accepted by the consumers were 0.8 g Bokbunja extract content and 0.6 g sugar content in the Hunter's color a value of 6.0.

Changes of Glucose-6-Phosphate Dehydrogenase on Alloxan-induced Diabetic Rat (Alloxan으로 유발된 당뇨쥐의 간조직 Glucose-6-Phosphate Dehydrogenase 효소의 변화)

  • 박수영;조경혜
    • Biomedical Science Letters
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    • v.3 no.2
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    • pp.169-175
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    • 1997
  • The hypoglycemic and metabolic effects of Commelina communis L. extract were investigated in alloxan-induced diabetic rats. The increased blood glucose level in the diabetic rats was sinificantly lowered with the treatments of the plant protein extract. Administration of the plant extract ellicited the significant increase of glucose-6-phosphate dehydrogenase (G6PD) activity in liver of alloxan-induced rats. Three isozyme patterns(band I, II & III : in order decreasing mobility) of G6PD were found when normal rat liver extract were subjected to electrophoresis on native polyacrylamide gel. On the other hand, G6PD band patterns of alloxan-induced rat liver extract were found band II isozyme missing. By treatment of plant extract in alloxan-induced rats has been showed pattern the recovery of missing band patterns. This indicates that changes of the G6PD isozyme might be related to the cellular process of diabetes.

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Output Enhancement of Rhodamine 6G Dye Laser by Rhodamine 560 Energy Transfer Dye (Rhodamine 560을 이용한 rhodamine 6G 색소 레이저의 출력 증가)

  • 장원권;이민희
    • Korean Journal of Optics and Photonics
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    • v.5 no.2
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    • pp.266-271
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    • 1994
  • The output power and the energy of Rh-6G dye laser were enhanced by the mixture of Rh-560 dye whose fluorescence spectrum was coincident with the absorption spectrum of Rh-6G. The argon filled coaxial flashlamp used for pulsed pumping and argon laser for CW pumping. The concentration of Rh-6G dye was optimized in each pumping method before Rh-560 dye was mixed in Rh-6G dye solution. In the coaxial flash lamp pumped Rh-6G laser the output energy was increased about 30% when Rh-560 was mixed at 1% of Rh-6G concentration. In the case of argon laser pumping with multiline, the output power was increased 18% at the concentration of 2.5%. In the single line laser pumping, the output power was enhanced more efficiently. The power enhancements were 72% and 88% when the pumping wavelengths were 488 nm and 514.5 nm respectively. ively.

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