• Journal of Photoscience
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• v.12 no.2
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• pp.87-93
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• 2005

A new assay technique to distinguish between pure compounds and the isomeric mixtures has been suggested using single molecule (SM) fluorescence detection technique. Since the number of emission spots in a fluorophorespread film prepared from a genuine dye solution was determined by experimental condition, the deviation of spot numbers from the expected values could be considered to be an indication of lower purity of the sample solution. The lower limit of sample concentration for this assay was determined to be $5{\times}10^{-10}$ M to show uniform number of expected spots within 10% uncertainties in our experimental condition. An individual fluorescence intensity distribution for a mixture of isomers having doubly different emissivities was simulated by adding distributions obtained from Cy3 and nile red (NR) independently. The result indicated that the mixture could be identified from the pure compounds through the difference in the number of Gaussian functions to fit the distribution. This new assay technique can be applied to the purity test for synthetic biofluorophores which are usually prepared in small quantities not enough for classical ensemble assays.

• Journal of the Korean Chemical Society
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• v.55 no.5
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• pp.746-750
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• 2011

Fluorescence correlation spectroscopy (FCS) is an emerging fluorescence technique used to study the dynamics of proteins on a millisecond to microsecond time scale at the single-molecule level. Solution pH-modulated protein conformational changes can be manifested by binding rate, fluorescence lifetime, and binding specificity of a probe molecule. The fluorescence lifetime of Nile red (NR) bound to apomyoglobin (apoMb) was measured to be $6{\pm}0.3$ ns, much longer than that in water solution ($2.9{\pm}0.2$ ns). As the unfolding population of apoMb increased by lowering pH of solution, the fraction for the longer lifetime of NR decreased with an increasing fraction for the shorter lifetime of NR in water. Unlike 1-anilino-8-naphthalene sulfonic acid, which has many lifetimes due to nonspecific binding to the unfolded apoMb, NR bound to apoMb possesses only a single lifetime. These results suggest that NR binds specifically to native apoMb and thus can be utilized to probe the folding/unfolding dynamics of apoMb using FCS.

• Korea-Australia Rheology Journal
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• v.20 no.3
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• pp.127-142
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• 2008

The development of fluorescence microscopy of single-molecule DNA in the last decade has fostered a bold jump in the understanding of polymer physics. With the recent advance of nanotechnology, devices with well-defined dimensions that are smaller than typical DNA molecules can be readily manufactured. The combination of these techniques has provided an unprecedented opportunity for researchers to examine confined polymer behavior, a topic far less understood than its counterpart. Here, we review the progress reported in recent studies that investigate confined polymer dynamics by means of single-molecule DNA experiments.

• BMB Reports
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• v.54 no.3
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• pp.157-163
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• 2021

The transient interactions between cellular components, particularly on membrane surfaces, are critical in the proper function of many biochemical reactions. For example, many signaling pathways involve dimerization, oligomerization, or other types of clustering of signaling proteins as a key step in the signaling cascade. However, it is often experimentally challenging to directly observe and characterize the molecular mechanisms such interactions-the greatest difficulty lies in the fact that living cells have an unknown number of background processes that may or may not participate in the molecular process of interest, and as a consequence, it is usually impossible to definitively correlate an observation to a well-defined cellular mechanism. One of the experimental methods that can quantitatively capture these interactions is through membrane reconstitution, whereby a lipid bilayer is fabricated to mimic the membrane environment, and the biological components of interest are systematically introduced, without unknown background processes. This configuration allows the extensive use of fluorescence techniques, particularly fluorescence fluctuation spectroscopy and single-molecule fluorescence microscopy. In this review, we describe how the equilibrium diffusion of two proteins, K-Ras4B and the PH domain of Bruton's tyrosine kinase (Btk), on fluid lipid membranes can be used to determine the kinetics of homodimerization reactions.

• Korean Journal of Optics and Photonics
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• v.10 no.1
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• pp.1-4
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• 1999

A confocal microscope system was used to study the bursts of fluorescence photons from single dye molecules excited at 638 nm by a short-pulsed diode laser with a repetition rate of 17 MHz. A red dye, JA22, in ethylene glycol solution was used as a sample. The fluorescence decay curves of single molecules were acquired using a time-correlated single photon counting and analyzed by a maximum likelihood estimator. It was possible to measure the fluorescence decay times with an error probability of 21% at photon number of more than 40 per dye molecule.

• Korean Journal of Optics and Photonics
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• v.8 no.4
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• pp.353-355
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• 1997

A single molecule detection system, which consists of confocal fluorescence microscope and single photon counter, has been used to observe the dye concentration dependence of photon counting rate. With increasing concentration, a saturation effect of counting is observed and demonstrated on the basis of the dead time of photon detector. The equations presented here show the relations between the counting rate and some parameters such as probe volume, quantum efficiency of detector, and fluorescence photon number entered onto detector. The signal-to-noise ratio is also discussed briefly.

• Proceedings of the Optical Society of Korea Conference
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• 2009.02a
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• pp.431-432
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• 2009

• Proceedings of the Optical Society of Korea Conference
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• 2004.02a
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• pp.180-181
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• 2004

• Proceedings of the Korean Biophysical Society Conference
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• 1998.06a
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• pp.24-24
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• 1998

The effect of stabilizing single ammo acid replacements at the sites of Phe 51, Ala 70, and Met 374 on the folding of $\alpha$$_1$-antitrypsin was investigated by fluorescence spectroscopy. The residues Phe 51 and Met 374 are located in the hydrophobic core of the molecule, B $\beta$-sheet.(omitted)

• Korean Journal of Optics and Photonics
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• v.8 no.1
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• pp.42-46
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• 1997

In order to acquire the fluorescence burst signals emitted from single dye molecules that pass through a detection space defined by a confocal microscope, a computer-interfaced photon counter has been fabricated. The maximum count rate is about 80 MHz, which is limited by the counter devices used. Using both the operating computer program written by BASIC and the 486 PC computer, the minimum bin-width of 25 $mutextrm{s}$ has been achieved. The characteristics of fluorescence burst signals emitted from JA22 molecules at about 1$\times$$10^{-11}$m mol/L in ethylene glycol are discussed briefly.