• Bulletin of the Korean Chemical Society
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• 제26권3호
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• pp.413-417
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• 2005

Fluorescence of green fluorescent protein mutant, 2-5 GFP is observed during denaturation by guanidine. The fluorescence intensity decreases exponentially but the fluorescence lifetime does not change during denaturation. The fluorescence lifetime of the denatured protein is shorter than that of native form. As the protein structure is modified by guanidine, solvent water molecules penetrate into the protein barrel and protonate the chromophore to quench fluorescence. Most fluorescence quenchers do not affect the fluorescence of native form but accelerate the fluorescence intensity decay during denaturation. Based on the observations, a simple model is suggested for the structural change of the protein molecule during denaturation.

• Journal of Photoscience
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• 제2권1호
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• pp.19-25
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• 1995

Picosecond time-resolved and static protein fluorescence spectra and absorption spectra of octopus rhodopsin, a photorecepting protein, are measured and compared with those of bacteriorhodopsin, a photon-induced proton pumping protein, to understand the protein conformations and functions of octopus rhodopsin and its deprotonated photocycle intermediate. The bluer and weaker absorption of retinal indicates that octopus rhodopsin is better in thermal noise suppression but less efficient in light harvesting than bacteriorhodopsin. The protein fluorescence of octopus rhodopsin shows the characteristic of Trp only and the uantum efficiency and lifetime variations may result primarily from variations in the coupling strength with the retinal. The stronger intensity by four times and larger red shift by 12 nm of fluorescence suggest that octopus rhodopsin has more open and looser structure compared with bacteriorhodopsin. Fluorescence decay profiles reveal two decay components of 300 ps (60%) and 2 ns (40%). The deprotonation of protonated Schiff's base increases the shorter decay time to 500 ps and enhances the fluorescence intensity by 20%. The fluorescence and its decay time from Trp residues near retinal are influenced more by the deprotonation. The increase of fluorescence intimates that protein structure becomes loosened and relaxed further by the deprotonation of protonated Schiff's base. The driving force of sequential changes initiated by absorption of a photon is too exhausted after the deprotonation to return the intermediate to the ground state of the begun rhodopsin form.

• 한국광학회지
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• 제35권1호
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• pp.30-34
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• 2024

실크 단백질과 녹색 형광 단백질을 결합한 순수 단백질 기반 녹색 형광체의 구현과, 산화티타늄 나노 입자에 의한 형광 증폭 현상을 보고한다. 실크 단백질은 인간에게 다양한 이점을 제공하는 소재로서, 다양한 마이크로/나노 구조 형성 가능성과 투명한 성질을 가지고 있어 광학소재로서의 활용 가능성 또한 높다. 본 연구에서 제작한 소재는 강한 녹색 형광을 띠고 있으며, 산화티타늄 나노 입자에 의해 녹색 형광이 7.5배 증폭되는 것을 확인하였다.

• BMB Reports
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• 제28권6호
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• pp.546-551
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• 1995

The conformation studies of myelin P2 protein and two of its major peptides were carried out using circular dichroism and fluorescence spectroscopy in water and in lipid environments. Significant conformational changes occur when the protein or peptides were bound to gangliosides. Similar effects were also found in trifluoroethanol solutions. The conformational features of P2 protein and its major peptides were discussed in relation to the environmental changes and the disease-inducing effects.

• 대한의용생체공학회:학술대회논문집
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• 대한의용생체공학회 1997년도 추계학술대회
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• pp.475-477
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• 1997

We have analyzed the fluorescence image obtaining from green fluorescence protein (GFP). In order to monitor the fluorescence of specific gene, we used the amyloid precursor protein promoter which has been known to act as a major role in the development of Alzheimer's disease. The promoter from - 3.0 kb to + 100 base pair was inserted into the gene expression monitoring GFP vector purchased from Clontech. This construct was transfected into the PC 12 and fibroblast cells and the fluorescence image was captured by two kinds of methods. One is using cheaper CCD camera and other is SIT-CCD camera. or the higher sensitivity of the fluorescence image, we developed the multiple image grabbing program. As a results, the fluorescence image by conventional CCD camera have the similar sensitivity compared with that of the SIT-camera by applying the multiple image grabbing programs. By this system. it will be possible to construct the fluorescence monitoring system with lower cost. And gene expression in real time by fluorescence image will be possible without changing the fluorescence images.

• 대한전기학회:학술대회논문집
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• 대한전기학회 2004년도 하계학술대회 논문집 C
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• pp.2040-2042
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• 2004

This paper presents a miniature optical system for the fluorescence detection of the patterned protein chip. The patterned protein chip was fabricated using MEMS process. The fluorescence from the patterned protein chip was measured while varying the concentration of the BSA. The fluorescence light is separated spatially from the excitation beam using mini-size prism to increase SNR (Signal-to-Noise Ratio). The combination of prism and mirrors can convert the excitation light from the laser diode to uniform illumination on the specimen. We believe that the proposed system for fluorescence detection can be applied to rea1ization of point-of-care.

• Journal of Microbiology and Biotechnology
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• 제11권6호
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• pp.979-985
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• 2001

A fluorometric detection technique for HDL (High Density Lipoprotein) and LDL (Low Density Lipoprotein) was developed for application in a fiber-optic immunosensor using a protein A Langmuir-Blodgget (LB) film. For the fluorescence immunoassay, antibodies specific to HDL or LDL were imobilied on the protein A LB film, and a fluorescence amplification method was developed to overcome their weak fluorescence. The deposition of protein A using the LB technique was monitored using a surface pressure-are $({\pi}-A)$ curve, and the antibody immobilization of the protein A LB film was experimentally verified. The immobilized antibody was used to separate only HDL and LDL from a sample, then the fluorescence of he separated HDL or LDL was amplified. The amount of LDL or HDL was measured using the developed fiber optic fluorescence detection system. The optical properties resulting from the reaction of HDL or LDL with o-phtaldialdehyde, detection range, response time, and stability of the immunoassay were all investigated. The respective detection ranges for HDL and LDL were sufficient to diagnose the risk of coronary heart disease. The amplification step increased the sensitivity, while selective separation using the immobilized antibody led to linearity in the sensor signal. The regeneration of the antibody-immobilized substrate could produce a stable and reproducible immunosensor.

• KSBB Journal
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• 제22권1호
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• pp.48-52
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• 2007

본 연구에서는 sonificator를 장착하여 세포막을 파쇄하고 현장에서 형광을 이용하여 조작이 간편하고 단시간에 DNA와 단백질을 동시에 측정할 수 있는 자동화된 형광기를 개발하기 위하여 단백질을 측정하는데 최적의 파쇄조건을 확립하고자 하였다. 용액 중에 녹아 있는 공기 중의 oxygen은 collisional quenching을 일으키는데 sonification이나 가열처리를 시키면 oxygen이 제거되어 quenching 효과가 크게 감소되어 높은 형광값을 나타내었다. 0.7 X 이상의 형광시약 농도에서는 반응시작 후 1분 이내에 측정해야 하며, 0.3 X 이하의 형광시약 농도에서는 반응시작 후 2$\sim$3분 사이에 반응을 시킨 후 측정하는 것이 바람직한 것으로 나타났다. $100{\mu}g/m{\ell}$ 이상의 BSA 농도에서는 형광시약이 포화되었으며, 시료를 sonification시키면 단백질이 변성되어 눈에 보일 정도로 불투명해져서 시료 용액의 불투명도로 인해 형광 값이 감소되는 경향을 나타내었으며, $1{\mu}g/m{\ell}$ 이하의 BSA 시료에서는 sonification을 시키지 않은 시료보다 sonification을 시켰을 때 $0.125{\mu}g/m{\ell}$의 BSA를 훨씬 더 구분이 잘 되어 낮은 단백질 농도에서는 sonification시키는 것이 훨씬 유리한 것으로 나타났다.

• Bulletin of the Korean Chemical Society
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• 제34권9호
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• pp.2725-2730
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• 2013

Single C-reactive protein (CRP) molecules, which are non-specific acute phase markers and products of the innate immune system, were quantitatively detected on a gold-nanopatterned biochip using evanescent field-enhanced fluorescence imaging. The $4{\times}5$ gold-nanopatterned biochip (spot diameter of 500 nm) was fabricated by electron beam nanolithography. Unlabeled CRP molecules in human serum were identified with single-molecule sandwich immunoassay by detecting secondary fluorescence generated by total internal reflection fluorescence (TIRF) microscopy. With decreased standard CRP concentrations, relative fluorescence intensities reduced in the range of 33.3 zM-800 pM. To enhance fluorescence intensities in TIRF images, the distance between biochip surface and CRP molecules was optimally adjusted by considering the quenching effect of gold and the evanescent field intensity. As a result, TIRF only detected one single-CRP molecule on the biochip the first time.

• 대한전기학회논문지:전기물성ㆍ응용부문C
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• 제54권8호
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• pp.378-383
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• 2005

We report the development of miniature fluorescence detection systems that employ miniature prism, mirrors and low coat CCD camera to detect the fluorescence emitted from 40 fluorescently-labeled protein patterns without scanner. This kind of miniature fluorescence detection system can be used in point of care. We introduce two systems, one uses prism+mirror block and the other uses prism and two mirrors. A large NA microscope eyepiece and low cost CCD camera are used. We fabricated protein chip containing multi-pattern BSA labeled with Cy5, using MEMS technology and modified the surface chemically to clean and to immobilize proteins. The measurements show that the combination of prism and mirrors can homogenize elliptical excitation light over the sample with higher optical efficiency, and increase the separation between excitation and fluorescence light at the CCD to give higher signal intensity and higher signal to noise ratio. The measurements also show that protein concentrations ranging from 10 ng/ml to 1000 ng/ml can be assayed with very small error. We believe that the proposed fluorescence detection system can be refined to build a commercially valuable hand-held or miniature detection device.