title: Quantitative nanoimmunosensor based on dark-field illumination with enhanced sensitivity and on-off switching using scattering signals.
abstract: A nanoimmunosensor based on wavelength-dependent dark-field illumination with enhanced sensitivity was used to detect a disease-related protein molecule at zeptomolar (zM) concentrations. The assay platform of 100-nm gold nanospots could be selectively acquired using the wavelength-dependence of enhanced scattering signals from antibody-conjugated plasmonic silver nanoparticles (NPs) with on-off switching using optical filters. Detection of human thyroid-stimulating hormone (hTSH) at a sensitivity of 100 zM, which corresponds to 1-2 molecules per gold spot, was possible within a linear range of 100 zM-100 fM (R=0.9968). A significantly enhanced sensitivity (~4-fold) was achieved with enhanced dark-field illumination compared to using a total internal reflection fluorescence immunosensor. Immunoreactions were confirmed via optical axial-slicing based on the spectral characteristics of two plasmonic NPs. This method of using wavelength-dependent dark-field illumination had an enhanced sensitivity and a wide, linear dynamic range of 100 zM-100 fM, and was an effective tool for quantitatively detecting a single molecule on a nanobiochip for molecular diagnostics.
authors.affiliation: Department of Applied Chemistry and Institute of Natural Sciences, College of Applied Science, Kyung Hee University, Yongin-si, Gyeonggi-do 17104, Republic of Korea.
authors.forename: Seungah
authors.initials: S
authors.lastname: Lee
authors.affiliation: Department of Chemistry, Graduate School, Kyung Hee University, Yongin-si, Gyeonggi-do 17104, Republic of Korea.
authors.forename: He
authors.initials: H
authors.lastname: Nan
authors.affiliation: Nanobio Fusion Research Center, Korea Research Institute of Standards and Science, Daejeon 34113, Republic of Korea.
authors.forename: Hyunung
authors.initials: H
authors.lastname: Yu
authors.affiliation: Department of Applied Chemistry and Institute of Natural Sciences, College of Applied Science, Kyung Hee University, Yongin-si, Gyeonggi-do 17104, Republic of Korea; Department of Chemistry, Graduate School, Kyung Hee University, Yongin-si, Gyeonggi-do 17104, Republic of Korea. Electronic address: shkang@khu.ac.kr.
authors.forename: Seong Ho
authors.initials: SH
authors.lastname: Kang
mesh_terms: D055910:Antibodies, Immobilized / Q000737:chemistry; D015374:Biosensing Techniques / Q000295:instrumentation*; D004867:Equipment Design; D006046:Gold / Q000737:chemistry; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation*; D008029:Lighting / Q000295:instrumentation; D057230:Limit of Detection; D053768:Metal Nanoparticles / Q000737:chemistry; D049329:Nanostructures / Q000737:chemistry; D012834:Silver / Q000737:chemistry; D013972:Thyrotropin / Q000032:analysis / Q000097:blood*
chemical_list: D055910:Antibodies, Immobilized; D012834:Silver; D006046:Gold; D013972:Thyrotropin
keywords: Enhanced dark-field illumination; Quantification; Switchable nanoimmunosensor; Ultra-high sensitivity