• KIEE International Transactions on Electrophysics and Applications
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• v.11C no.3
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• pp.85-90
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• 2001

The determination of DNA hybridization reaction can apply the molecular biology research, clinic diagnostics, bioengineering, environment monitoring, food science and other application area. So, the improvement of DNA detection system is very important for the determination of this hybridization reaction. In this study, we report the characterization of the probe and target oligonucleotide hybridization reaction using the evanescent field microscopy. First, we have fabricated DNA chip microarray. The particles which were immobilized oligonucleotides were arranged by the random fluidic self-assembly on the pattern chips, using hydrophobic interaction. Second, we have detected DNA hybridization reaction using evanescent field microscopy. The 5'-biotinylated probe oligonucleotides were immobilized on the surface of DNA chip microarray and the hybridization reaction with the Rhodamine conjugated target oligonucleotide was excited fluorescence generated on the evanescent field microscopy. In the foundation of this result, we could be employed as the basis of a probe olidonucleotide, capable of detecting the target oligonucleotide and monitoring it in a large analyte concentration range and various mismatching condition.

• Proceedings of the KIEE Conference
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• 2003.10a
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• pp.89-91
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• 2003

The determination of DNA hybridization reaction can apply the molecular biology research, clinic diagnostics, bioengineering, environment monitoring, food science and other application area. So, the improvement of DNA detection system is very important for the determination of this hybridization reaction. In this study, we report the characterization of the probe and target oligonucleotide hybridization reaction using the evanescent field microscopy. First, we have fabricated DNA chip microarray. The particles which were immobilized oligonucleotides were arranged by the random fluidic self-assembly on the pattern chips, using hydrophobic interaction. Second, we have detected DNA hybridization reaction using evanescent field microscopy. The 5'-biotinylated probe oligonucleotides were immobilized on the surface of DNA chip microarray and the hybridization reaction with the Rhodamine conjugated target oligonucleotide was excited fluorescence generated on the evanescent field microscopy. In the foundation of this result, we could be employed as the basis of a probe olidonucleotide, capable of detecting the target oligonucleotide and monitoring it in a large analyte concentration range and various mismatching condition.

• Journal of the Korean Society of Visualization
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• v.3 no.1
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• pp.3-19
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• 2005

Novel optical techniques are presented for three-dimensional tracking of nanoparticles; Optical Serial Sectioning Microscopy (OSSM) and Ratiometric Total Internal Reflection Fluorescent Microscopy (R-TIRFM). OSSM measures optically diffracted particle images, the so-called Point Spread Function (PSF), and dotermines the defocusing or line-of-sight location of the imaged particle measured from the focal plane. The line-of-sight Brownian motion detection using the OSSM technique is proposed in lieu of the more cumbersome two-dimensional Brownian motion tracking on the imaging plane as a potentially more effective tool to nonintrusively map the temperature fields for nanoparticle suspension fluids. On the other hand, R-TIRFM is presented to experimentally examine the classic theory on the near-wall hindered Brownian diffusive motion. An evanescent wave field from the total internal reflection of a 488-nm bandwidth of an argon-ion laser is used to provide a thin illumination field of an order of a few hundred nanometers from the wall. The experimental results show good agreement with the lateral hindrance theory, but show discrepancies from the normal hindrance theory. It is conjectured that the discrepancies can be attributed to the additional hindering effects, including electrostatic and electro-osmotic interactions between the negatively charged tracer particles and the glass surface.

• Journal of the Korean Society of Visualization
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• v.21 no.3
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• pp.65-74
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• 2023

Compared to epifluorescence(EPI) microscopy which captures fluorescence from the entire depth of sample, total internal reflection fluorescence(TIRF) can selectively visualize only a single surface of it. TIRF uses a thin evanescent field generated by the total internal reflection of laser light on surface. However, conventional TIRF system are designed for total internal reflection to occur at the upper surface of sample, making them unsuitable for sessile droplet imaging. We designed a TIRF system suitable for a sessile droplet imaging by utilizing slide glass as a lightguide. We presented the details for constructing the TIRF system using a prism, slide glass, air slit, and optical trap. Then, we compared the TIRF with EPI by imaging the droplet with fluorescent particles during its drying process. As a result, TIRF allows us to distinctly visualize the drying pattern on the bottom surface of droplet.

• Bulletin of the Korean Chemical Society
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• v.34 no.9
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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.