Article Cite This: J. Am. Chem. Soc. XXXX, XXX, XXX−XXX
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Smart SERS Hot Spots: Single Molecules Can Be Positioned in a Plasmonic Nanojunction Using Host−Guest Chemistry Nam Hoon Kim,†,¶ Wooseup Hwang,†,‡,¶ Kangkyun Baek,*,† Md. Rumum Rohman,† Jeehong Kim,†,‡ Hyun Woo Kim,† Jungho Mun,§ So Young Lee,∥ Gyeongwon Yun,† James Murray,† Ji Won Ha,∥ Junsuk Rho,§,⊥ Martin Moskovits,*,# and Kimoon Kim*,†,‡ †
Center for Self-assembly and Complexity, Institute for Basic Science, Pohang 37673, Republic of Korea Department of Chemistry, §Department of Chemical Engineering, and ⊥Department of Mechanical Engineering, Pohang University of Science and Technology, Pohang 37673, Republic of Korea ∥ Department of Chemistry, University of Ulsan, Ulsan 44610, Republic of Korea # Department of Chemistry and Biochemistry, University of California, Santa Barbara, California 93106, United States ‡
S Supporting Information *
ABSTRACT: Single-molecule surface-enhanced Raman spectroscopy (SERS) offers new opportunities for exploring the complex chemical and biological processes that cannot be easily probed using ensemble techniques. However, the ability to place the single molecule of interest reliably within a hot spot, to enable its analysis at the single-molecule level, remains challenging. Here we describe a novel strategy for locating and securing a single target analyte in a SERS hot spot at a plasmonic nanojunction. The “smart” hot spot was generated by employing a thiol-functionalized cucurbit[6]uril (CB[6]) as a molecular spacer linking a silver nanoparticle to a metal substrate. This approach also permits one to study molecules chemically reluctant to enter the hot spot, by conjugating them to a moiety, such as spermine, that has a high affinity for CB[6]. The hot spot can accommodate at most a few, and often only a single, analyte molecule. Bianalyte experiments revealed that one can reproducibly treat the SERS substrate such that 96% of the hot spots contain a single analyte molecule. Furthermore, by utilizing a series of molecules each consisting of spermine bound to perylene bisimide, a bright SERS molecule, with polymethylene linkers of varying lengths, the SERS intensity as a function of distance from the center of the hot spot could be measured. The SERS enhancement was found to decrease as 1 over the square of the distance from the center of the hot spot, and the single-molecule SERS cross sections were found to increase with AgNP diameter.
1. INTRODUCTION Measurements at the single-molecule level are seminal goals in molecular sciences, since they can provide valuable information regarding the dynamics of individual molecules as a function of time and space.1 Such insights augment our understanding of complex chemical2 and biological processes.3 Multiple spectroscopic and microscopic techniques have been shown4 capable of reporting molecular information at the single-molecule limit. Among them, surface-enhanced Raman spectroscopy (SERS) can yield vibrational spectra of single molecules on account of the large optical enhancements at SERS hot spots, which, oftentimes, are junctions between plasmonic nanostructures. Several strategies have been reported for single-molecule detection by SERS, including concentration control,5 bianalyte/isotopologue measurements,6 tip-enhanced Raman scattering,7 and nanogap-enhanced Raman scattering.8 Great emphasis has been placed on hot spots in producing intense SERS, and efforts have been made to develop techniques that locate target analytes predominantly at SERS hot spots.9,10 Despite this, few studies have been devoted to understanding the enhancement © XXXX American Chemical Society
distribution within an active hot spot and the location of a molecule resident within it, presumably due to the lack of methods for precise positioning of analytes.10 Recently, Willets mapped the position of a single molecule in a SERS hot spot using super-resolution microscopy,11 and Hou and co-workers determined the location and orientation of porphyrin molecules adsorbed on a silver substrate.12 However, these approaches still require special instruments and/or conditions such as ultrahigh vacuum and low temperature. Cucurbit[n]uril (CB[n], n = 5−8, 10, 14), a family of rigid, hollow pumpkin-shaped macrocyclic molecules, can form very stable and specific host−guest complexes with various guest molecules in aqueous solution.13 Recently, Mahajan, Scherman, and Baumberg,14 and Li15 reported the self-assembly of CB[n]bridged gold nanoparticles, and gold nanoparticles on a gold film separated by CB[n]. The CB[n] acts as a molecular spacer to give a well-defined nanojunction with a 0.9 nm gap, so that Received: February 7, 2018 Published: February 27, 2018 A
DOI: 10.1021/jacs.8b01501 J. Am. Chem. Soc. XXXX, XXX, XXX−XXX
Article
Journal of the American Chemical Society
on this approach, we are able to map the relative magnitude of the SERS enhancement with subnanometer resolution at various locations within the hot spots.
guest molecules inside the CB[n] cavity produce intense SERS signals. However, the nanoparticles blocked the carbonyl portals of CB[n], thereby limiting entry to the cavity of CB[n], which may consequently hinder the utilization of the host− guest chemistry of CB[n]. A simple method to immobilize a CB[n] molecule with an accessible cavity at the nanojunction between metal nanoparticles and a metal surface is, therefore, highly desirable. Such a method would allow a wider range of guest molecules to be placed within the nanojunction, and hence interrogated by SERS at the single-molecule level, and likely greater reproducibility. Here, we report a novel strategy for localizing and securing a single target molecule within a plasmonic nanojunction utilizing mercaptopropyloxy-CB[6] (thiol-CB[6])16 as a dual-function linker. Thiol-CB[6], a version of CB[n] in which its periphery is fully thiolated, is used edgewise to link a single silver nanoparticle (AgNP) to a Ag substrate through strong S−Ag bonds, thereby allowing the open CB[6] carbonyl portal to capture a single molecule of interest, automatically locating it in a hot spot (Figure 1). Consequently, an analyte of interest
2. EXPERIMENTAL SECTION 2.1. General Procedures and Materials. Octanoic acid and tris(2-carboxyethyl)phosphine hydrochloride (TCEP) were purchased from Aldrich and used as received. Thioacetate-functionalized cucurbit[6]uril (precursor of thiol-CB[6], PTE-CB[6]),16 Ag nanoparticles (20, 40, 60 nm),18 spermine-C(n)-perylene bisimide (spmC(n)-PBI) (n = 0, 2, 5, 7),19 and spermine-conjugated fluorescein isocyanate (spm-FITC)20 were synthesized as described in the literature. Other chemicals, unless specified, were reagent grade and used as supplied without further purification. Highly purified water with a resistivity greater than 18.0 MΩcm−1 (Millipore Milli-Q System) was used to prepare the aqueous solutions. A macroscopically smooth Ag substrate was prepared by e-beam evaporation of titanium (5 nm) and silver (300 nm) onto a freshly cleaved Si wafer. AFM images were collected using a Park System NX10 and VEECO Dimension 3100. FT-IR spectra were recorded on an Agilent Technologies Cary 600 series FT-IR spectrophotometer. SEM images were obtained using a Hitachi cold SEM microscope. 2.2. Fabrication of SERS Hot Spot. A solution of PTE-CB[6] (5 nM, 0.2 mL) in ethanol and a solution of octanoic acid (20 mM, 0.05 mL) in ethanol were combined and diluted to 1 mL with ethanol. An aqueous solution of sodium hydroxide (40 wt %, 0.01 mL) was added to the mixture for in situ generation of thiol-CB[6]. After the resulting solution was incubated for 10 min at room temperature, an aqueous solution of TCEP (1 mM, 0.03 mL) was added to the solution to prevent the formation of disulfide bonds between the thiol-CB[6] molecules. After 10 min of incubation, a Ag substrate (5 × 5 mm2) was immersed in the solution for 10 min, washed with water and ethanol, and finally immersed in a solution of AgNP for 10 min. The AgNPimmobilized Ag substrate was washed with water and then soaked in a solution of octanoic acid (1 mM) in ethanol (1 mL) to passivate the remaining surface of AgNP. After incubation for 6 h, the resulting substrate was washed with ethanol and water and dried with N2 gas. 2.3. Raman Measurements. Raman measurements were performed with a Renishaw inVia microRaman system. Samples were excited by a 632.8 nm He−Ne laser through a 50× (NA = 0.75) objective. The Raman band of a silicon wafer at 520 cm−1 was used to calibrate the spectrometer. The laser power was kept under 1 mW for the experiments investigating the accessibility of molecules to CB[6]; for the single-molecule experiments,
