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Lee, Seungah,Park, Guenyoung,Chakkarapani, Suresh Kumar,Kang, Seong Ho Elsevier 2015 Biosensors & bioelectronics Vol.63 No.-
<P><B>Abstract</B></P> <P>Novel, fluorescence-free detection of biomolecules on nanobiochips was investigated based on plasmonic nanometal scattering in the evanescent field layer (EFL) using total internal reflection scattering (TIRS) microscopy. The plasmonic scattering of nanometals bonded to biomolecules was observed at different wavelengths by an electromagnetic field in the EFL. The changes in the scattering of nanometals on the gold-nanopatterned chip in response to the immunoreaction between silver nanoparticles and antibodies allowed fluorescence-free detection of biomolecules on the nanobiochips. Under optimized conditions, the TIRS immunoassay chip detected different amounts of immobilized antigen, i.e., human cardiac troponin I. The sandwich immuno-reaction was quantitatively analyzed in the dynamic range of 720zM–167fM. The limit of detection (<I>S</I>/<I>N</I>=4) was 600zM, which was ~140 times lower than limits obtained by previous total internal reflection fluorescence and dark field methods. These results demonstrate the possibility for a fluorescence-free biochip nanoimmunoassay based on the scattering of nanometals in the EFL.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Fluorescence-free detection of biomolecules on nanobiochips by total internal reflection scattering microscopy. </LI> <LI> Observation of the scattering shape and intensity of native nanometals by controlling optical components. </LI> <LI> Highly sensitive fluorescent-free detection of cTnI with a detection limit of 600zM. </LI> </UL> </P>
Lee, Seungah,Cho, Nam-Pyo,Kim, Jung Dong,Jung, Hyungil,Kang, Seong Ho Royal Society of Chemistry 2009 The Analyst Vol.134 No.5
<P>This paper describes a single-molecule sandwich immunoassay method that utilizes total internal reflection fluorescence microscopy (TIRFM) at the single-molecule level for nanoarray protein chip applications. Nanoarray patterning of a biotin-probe with a spot diameter of 179 ± 1 nm was performed successfully on a (3-mercaptopropyl)trimethoxysilane (MPTMS)-coated glass substrate by atomic force microscopy (AFM). The formation of biotin patterns was confirmed directly by observing the heights of bound streptavidin and biotin-antibody on glass substrates using an AFM in contact mode. Target protein molecules (or antigen) at the zepto-molar (zM) concentration level (× 10<SUP>−21</SUP> M) were detected on MPTMS-coated glass nanoarray protein chips by TIRFM. Finally, cytokine clinical samples (<I>i.e.</I> TNF-α and IL-1α) as cancer marker protein molecules were applied to nanoarray protein chips, and detection limits were at 600 zM.</P> <P>Graphic Abstract</P><P>The single-molecule sandwich immunoassay for the ultra-sensitive detection of TNF-α on an MPTMS-coated glass substrate is feasible for the detection of the target protein molecules at the zM concentration level (× 10<SUP>−21</SUP> M). <IMG SRC='http://pubs.rsc.org/services/images/RSCpubs.ePlatform.Service.FreeContent.ImageService.svc/ImageService/image/GA?id=b822094h'> </P>
Lee, Kyuseo,Lee, Seungah,Yu, Hyunung,Kang, Seong Ho American Scientific Publishers 2010 Journal of Nanoscience and Nanotechnology Vol.10 No.5
<P>We detected 'tumor necrosis factor-alpha (TNF-alpha)' on a gold nanoarray protein chip at the attomolar (aM) concentration level (x10(-18)) by total internal reflection fluorescence microscopy (TIRFM). The 4 x 5 nanoarray patterning of gold with a spot diameter of 500 nm was successfully achieved on 10-mm square glass substrates using an electron beam evaporator. The dithiobis(succinimidyl propionate) as a protein linker and the Protein A/G to help oriented immobilization of antibody were used. The interactions of individual protein molecules were detected based on the sandwich fluorescence immunoassay by TIRFM. The linear regression equation for TNF-alpha in the range of 13 aM-130 fM was determined to be y = 6687.8x +126133 (R = 0.9938). The detection limit was 1.3 aM (S/N = 3). These results show that TNF-alpha as a tumor marker protein molecule was applied to gold nano-patterned protein chip.</P>
Dual-color prism-type TIRFM system for direct detection of single-biomolecules on nanoarray biochips
Seungah Lee,강성호,정봉현 한국물리학회 2008 Current Applied Physics Vol.8 No.6
This study examined the applicability of a prism-type simultaneous dual-color total internal reflection fluorescence microscopy (TIRFM) system for the simultaneous detection of nano biomolecules on nanoarray biochips at the single-molecule level. The dual-color TIRFM system with two individual laser beams and a high-sensitivity camera was used for the simultaneous dual-color detection of two different nano beads (i.e. 20 nm yellow–green and crimson fluorescent FluoSpheres beads), and single-protein molecules labeled with different fluorescent dyes (i.e. actin from rabbit muscle conjugated with Alexa Fluor 488 and Alexa Fluor 633 goat anti-rabbit IgG) without a time-delay and the need to move the sample. When this system was applied to two different single-protein molecules labeled with different fluorescent dyes on the GPTS/CHI/GA-modified glass nanoarray chip, the full images of the biomolecules at the single-molecule level were obtained simultaneously in two different colors using a Dual-ViewTM. The dual-color TIRFM system is quite suitable for the biological imaging at the single-molecule level on nanoarray biochips. This study provides a benchmark for directly monitoring the interactions and detecting the colocalization of two different nano biomolecules, and can be applied to the development of a nanoarray biochip at the single-molecule level.
Lee, Seungah,Yu, Hyunung,Kang, Seong Ho The Royal Society of Chemistry 2013 Chemical communications Vol.49 No.75
<P>Individual silver nanoparticle-conjugated target protein (cTnI) molecules on gold-nanopatterned chip were selectively detected by wavelength dependent-enhanced dark field illumination. Using specific nanoparticles with unique sizes and materials, the immunotargeted nanoparticle on the chips was detected at the single-molecule level by monitoring changes in the plasmonic resonance based on wavelength dependence.</P> <P>Graphic Abstract</P><P>Individual silver nanoparticle-conjugated target protein molecules (human cardiac troponin I) on a nanobiochip were selectively detected by wavelength dependent-enhanced dark field illumination. <IMG SRC='http://pubs.rsc.org/services/images/RSCpubs.ePlatform.Service.FreeContent.ImageService.svc/ImageService/image/GA?id=c3cc44153a'> </P>
Lee, Seungah,Yu, Hyunung,Kang, Seong Ho The Royal Society of Chemistry 2015 Chemical communications Vol.51 No.5
<P>Immunoassays on nanopatterned chips through TIRS detection based on reconstructing the three dimensional position provided a nanoscale accuracy of the lateral resolution by using the <I>z</I>-stage controller in the spatial range up to 10 nm. This method offers highly accurate and sensitive quantification with the zeptomolar (∼10<SUP>−21</SUP> M) detection of proteins.</P> <P>Graphic Abstract</P><P>The total internal reflection scattering system incorporating a <I>z</I>-nanopositioner is introduced to explore the precise immunoassay on gold-nanopattemed chips by lateral resolution improvement. <IMG SRC='http://pubs.rsc.org/services/images/RSCpubs.ePlatform.Service.FreeContent.ImageService.svc/ImageService/image/GA?id=c4cc07665f'> </P>
( Seungah Yoo ),( Hyojung Kim ),( Juhee Han ),( Chulhwan Bang ),( Youngmin Park ),( Junyoung Lee ),( Jihyun Lee ) 대한피부과학회 2019 대한피부과학회 학술발표대회집 Vol.71 No.2
Background: Actinic keratosis (AK) has been considered as a precancerous lesion which can progress to squamous cell carcinoma, yet no gold standard treatment has been established. There have been few studies comparing therapeutic efficacy of treatment modalities for AK. Objectives: We tried to investigate the effective management of AK by comparing the clearance and recurrence rates of treatment modalities. Methods: We reviewed medical records of 410 patients diagnosed with AK in Seoul St. Mary’s Hospital from February 2015 to February 2019. Patients treated with two or more treatment methods, or with no more than 6 months of follow-up were excluded. Finally total 296 patients were included. Results: Average treatment period was the shortest in ingeonl mebutate (IM), followed by photodynamic therapy (PDT), imiquimod (IMQ), and cryotherapy (3 days, 3.5, 6.1, 9.0 weeks respectively). The number of visits was the lowest in PDT, followed by IMQ, cryotherapy, and IM (1.75, 1.82, 2.27, 3.00). IMQ and PDT showed better therapeutic efficacy (same as 80.0%), than cryotherapy and IM (79.7, 77.8%). In recurrence rates, cryotherapy was the lowest, followed by IMQ, PDT, and IM (5.4, 5.6, 7.7, 15.2%). Conclusion: Cryotherapy and IMQ appeared to be effective and less-recurring treatments. PDT seemed to be a good choice, despite a slightly higher recurrence rate, it had a higher clearance rate, shorter treatment period, and fewer visits. IM was less effective especially due to a higher recurrence rate.