• Title/Summary/Keyword: Fluorescent resonance energy transfer (FRET)

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A Possible Merge of FRET and SPR Sensing System for Highly Accurate and Selective Immunosensing

  • Lee, Jae-Beom;Chen, Hongxia;Lee, Jae-Wook;Sun, Fangfang;Kim, Cheol-Min;Chang, Chul-Hun L.;Koh, Kwang-Nak
    • Bulletin of the Korean Chemical Society
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    • v.30 no.12
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    • pp.2905-2908
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    • 2009
  • Immuno-sensing for high accurate and selective sensing was performed by fluorescence spectroscopy and surface plasmon resonance (SPR), respectively. Engineered assembly of two fluorescent quantum dots (QDs) with bovine serum albumin (BSA) and anti-BSA was fabricated in PBS buffer for fluorescence analysis of fluorescence resonance energy transfer (FRET). Furthermore, the same bio-moieties were immobilized on Au plates for SPR analysis. Naturally-driven binding affinity of immuno-moieties induced FRET and plasmon resonance angle shift in the nanoscale sensing system. Interestingly, the sensing ranges were uniquely different in two systems: e.g., SPR spectroscopy was suitable for highly accurate analysis to measure in the range of 10$^{-15{\sim}-10$ng/mL while the QD fluorescent sensing system was relatively lower sensing ranges in 10$^{-10{\sim}-6$ng/mL. However, the QD sensing system was larger than the SPR sensing system in terms of sensing capacity per one specimen. It is, therefore, suggested that a mutual assistance of FRET and SPR combined sensing system would be a potentially promising candidate for high accuracy and reliable in situ sensing system of immune-related diseases.

Development of a FRET-based High-Throughput Screening System for the Discovery of Hsp90 Inhibitors

  • Oh, Sang-Mi;Ko, Yeon-Jin;Lee, Han-Jae;Kim, Jong-Hoon;Chung, Young-Sun;Park, Seung-Bum
    • Bulletin of the Korean Chemical Society
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    • v.32 no.9
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    • pp.3229-3232
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    • 2011
  • A FRET-based high-throughput screening system was developed for the discovery of competitive smallmolecule Hsp90 inhibitors. The biarsenical fluorescein derivative FlAsH and dabcyl-conjugated Hsp90 inhibitor GM were employed as the FRET donor and quencher, respectively. The spatial proximity perturbation between FlAsH-labeled Hsp90N and GM-dabcyl upon treatment of a small molecule led to changes in the FRET-induced fluorescence, monitored in a high-throughput fashion.

DNA Dynamics: a Fluorescence Resonance Energy Transfer Study Using a Long-Lifetime Metal-Ligand Complex

  • Kang, Jung-Sook;Lakowicz, Joseph-R.;Piszczek, Grzegorz
    • Archives of Pharmacal Research
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    • v.25 no.2
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    • pp.143-150
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    • 2002
  • Fluorescent probes bound to DNA typically display nanosecond decay times and reveal only nanosecond motions. We extend the time range of measurable DNA dynamics using $[Ru(pby)_2(dppz)]^{2+}$ (bpy=2.2'-bipyridine, dppz=dipyrido[3,2-a2',3'-c]phenazine) (RuBD) which displays a mean lifetime near 90 ns. To test the usefulness of RuBD as a probe for diffusive processes in calf thymus DNA, we compared the efficiencies of fluorescence resonance energy transfer (FRET) using three donors which display lifetimes near 5 ns for acridine orange (AO), 22 ns for ethidum bromide (EB) and 92 ns for RuBD, with nile blue (NB) as the acceptor. The F rster distances for AO-NB, EB-NB and RuBD-NB donor-acceptor pairs were 42.3, 52.3, and $30.6{\;}{\AA}$, respectively. All three donors showed dramatic decreases in fluorescence intensities and more rapid intensity decays with increasing NB concentrations. The intensity decays of AO and EB in the presence of varying concentrations of NB were satisfactorily described by the one-dimensional FRET model without diffusion (Blumen and Manz, 1979). In the case of the long-lifetime donor RuBD, the experimental phase and modulation somewhat deviated from the recovered values computed from this model. The recovered NB concentrations and FRET efficiencies from the model were slightly larger than the expected values, however, the recovered and expected values did not show a significant difference. Thus, it is suggested that the lifetime of RuBD is too short to measure diffusive processes in calf thymus DNA.

Research Method of Fatty Acids Transfer between Phospholipid Model Membranes (인지질 모델막에서의 지방산 이동에 관한 연구 방법)

  • 임병순;김혜경;김을상
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.26 no.4
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    • pp.743-750
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    • 1997
  • Direct measurement of the kinetics of free fatty acid transfer between phospholipid model membrane is technically limited by the rapid nature of the transfer process. Separation of membrane-bound fatty acid by centrifugation has shown that although the equilibrium distribution of free fatty acid is determined by this method, fatty acid transfer occurs too rapidly for accurate kinetic measurements. Recently fluorescence resonance energy transfer(FRET) assay has been developed to examine transfer of fatty acids between membranes. Donor membranes which has fluorescent fatty acid, anthroyloxy fatty acid(AOFA), is mixed with acceptor membranes which has non-interchangeable fluorescent quencher, nitrobenzo-xadiazol(NBD), using stopped flow apparatus. As the fluorescent fatty acids transfer from donor membrane to acceptor membrane, fluorescence intensity would be decreased and the rate and degree of fatty acid transfer can be analyzed. Fatty acid transfer between micelles is more complicated because of bile salt. Therefore in experiments with micelles, fluorescence self quenching assay is used. At high concentrations, a fluorophore tends to quench its own fluorescence causing a reduction in fluorescence intensity. Donor micelles contained self quenching concentrations of fluorophore and acceptor micelles had no fluorophore. Upon mixing of donor and acceptor micelles, the rate of transfer of the fluorophore from the donor to the acceptor was measured by monitoring the release in self quenching when its concentration in donor decreased over time.

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Plasmid-DNAgram : Anagram Solving by Molecular Computing Based on GFP-Expressing Plasmid DNA (Plasmid-DNAgram : 녹색형광단백질 발현 Plasmid DNA 기반 분자컴퓨팅에 의한 언어 퍼즐 문제 해결)

  • Kim, Su-Dong;Lee, Eun-Seok;Zhang, Byoung-Tak
    • Annual Conference on Human and Language Technology
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    • 2003.10d
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    • pp.293-299
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    • 2003
  • 인간 게놈 프로젝트가 완료됨에 다라 생체서열과 언어 사이의 대응 관계가 부각되고 있다. 본고에서는 Lewis Carroll의 언어 유희 사례를 컴퓨터생물학의 측면에서 재조명하고, Carroll이 제시한 문제 중에서 간단한 anagram 문제의 해결을 다루고자 한다. 우선 DNA 컴퓨팅의 방법론을 적용한 DNAgram의 개념을 확장하여 plasmid-DNAgram의 개념을 새롭게 도입하였다. 이 개념을 형광단백질에 대한 DNAgram의 개념을 확장하여 plasmid-DNAgram의 개념을 새롭게 도입하였다. 이 개념을 형광단백질에 대한 FRET(fluorescent resonance energy transfer)분석기법의 응용 사례인 cameleon 형광단백질에 대한 FRET 분석기법에 적용함으로써 anagram 문제의 어휘론적, 구문론적, 의미론적, 화용론적 측면에 대응하는 바이오분자 컴퓨팅 방법론을 제안하였다.

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Techniques for Evaluation of LAMP Amplicons and their Applications in Molecular Biology

  • Esmatabadi, Mohammad javad Dehghan;Bozorgmehr, Ali;zadeh, Hesam Motaleb;Bodaghabadi, Narges;Farhangi, Baharak;Babashah, Sadegh;Sadeghizadeh, Majid
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.17
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    • pp.7409-7414
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    • 2015
  • Loop-mediated isothermal amplification (LAMP) developed by Notomi et al. (2000) has made it possible to amplify DNA with high specificity, efficiency and rapidity under isothermal conditions. The ultimate products of LAMP are stem-loop structures with several inverted repeats of the target sequence and cauliflower-like patterns with multiple loops shaped by annealing between every other inverted repeats of the amplified target in the similar strand. Because the amplification process in LAMP is achieved by using four to six distinct primers, it is expected to amplify the target region with high selectivity. However, evaluation of reaction accuracy or quantitative inspection make it necessary to append other procedures to scrutinize the amplified products. Hitherto, various techniques such as turbidity assessment in the reaction vessel, post-reaction agarose gel electrophoresis, use of intercalating fluorescent dyes, real-time turbidimetry, addition of cationic polymers to the reaction mixture, polyacrylamide gel-based microchambers, lateral flow dipsticks, fluorescence resonance energy transfer (FRET), enzyme-linked immunosorbent assays and nanoparticle-based colorimetric tests have been utilized for this purpose. In this paper, we reviewed the best-known techniques for evaluation of LAMP amplicons and their applications in molecular biology beside their advantages and deficiencies. Regarding the properties of each technique, the development of innovative prompt, cost-effective and precise molecular detection methods for application in the broad field of cancer research may be feasible.