Effect of Alkylamines on Morphology Control of Silver Nanoshells for

Subscriber access provided by University of Massachusetts Amherst Libraries

Functional Nanostructured Materials (including low-D carbon)

Effect of Alkylamines on Morphology Control of Silver Nanoshells for Highly Enhanced Raman Scattering Myeong Geun Cha, Homan Kang, Yun-Sik Choi, Yoojin Cho, Minwoo Lee, Ho-Young Lee, Yoon-Sik Lee, and Dae Hong Jeong ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.8b15674 • Publication Date (Web): 04 Feb 2019 Downloaded from http://pubs.acs.org on February 4, 2019

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ACS Applied Materials & Interfaces

Effect of Alkylamines on Morphology Control of Silver Nanoshells for Highly Enhanced Raman Scattering Myeong Geun Cha§, †, Homan Kang+, †, Yun-Sik Choi§, Yoojin Cho§, Minwoo Lee§, Ho-Young Lee± , Yoon-Sik Lee⊥, * and Dae Hong Jeong§, ¶, * §

+

Department of Chemistry Education, Seoul National University, Seoul 08826, Korea.

Department of Radiology, Harvard Medical School and Gordon Center for Medical Imaging, Massachusetts General Hospital, Boston, MA 02129, USA

±

Department of Nuclear Medicine, Seoul National University Bundang Hospital, Seongnam 13620, Korea.

⊥

School of Chemical and Biological Engineering, Seoul National University, Seoul 08826, Korea. ¶

Center for Educational Research, Seoul National University, Seoul 08826, Korea.

KEYWORDS: silver nanoshell, morphology, alkylamine, surface-enhanced Raman scattering (SERS), in vivo detection

ACS Paragon Plus Environment

ACS Applied Materials & Interfaces 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 2 of 32

ABSTRACT: Morphology control of the surface of a nanostructure is a key issue in modulating its surface plasmon resonance and scattering properties. Here, we studied the effect of alkylamines on morphology control during one-step fabrication of silver nanoshells (NS) for highly enhanced Raman scattering. Various types of alkylamine were used to study the effects of chain length, existence of hydroxyl group and degree of alkyl chain on the surface morphology of silver NS. The alkylamines influenced the silver ion reduction and the growth of silver domains, resulting in distinctive morphology changes. The optical properties of the silver NSs of different surface morphologies were characterized by surface-enhanced Raman spectra. Especially, when long alkylamines were used, intense and uniform SERS signals were obtained at visible and NIR region and their enhancement factor (EF) were ~10 7. To detect cancer biomarkers in vivo, as a feasibility test, silver NSs were modified to highly NIR-active nanoprobes and successfully applied to detect colon cancer without causing non-specific interactions. Our one-step fabrication method of silver NSs is simple and can overcome various hurdles of morphology control, and can be extended to other metal nanostructures of controlled surface morphologies or shape.


Page 3 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


1. Introduction

Metal nanostructures are used in various fields such as surface-enhanced Raman scattering (SERS),1-4 bioimaging,5-8 disease diagnosis and therapy,9 and catalysis.10 To achieve the desired functions in these applications, numerous methods for controlling the shape and morphology of metal nanostructures have been developed. So far, various shapes of nanostructures have been reported, including sphere,11-12 rod,13-14 star,15-16 shell,17-18 flower,19-20 cube21-22 and polyhedral types23-24 with unique optical properties depending on their size, shape, and composition.18, 25-28 Recently, metal nanoshells (NSs) have received much attention because their surface plasmon resonance (SPR) can be easily tuned in a wide range from the visible to the near IR (NIR) region by modifying their core size, shell thickness, surface morphology, and atomic composition.29-34 Various types of NSs with controllable surface morphologies such as a multi-branched30, 35-38 or bumpy structure have been developed.39-42 These NSs have highly localized electron density at their surface and the scattering properties in the NIR region is much better tunable than nanospheres and nanorods. However, most of the reported methods require multistep synthesis, such as seed-mediated growth18, 30 and layer-by-layer growth43-45 to obtain the desired nanoshell structure by using excess amount of bulky surfactants such as cetyltrimethylammonium bromide (CTAB) and their derivatives to change the surface morphology.31, 46 Such an excessive use of surfactants limits the biomedical applications of NSs due to the toxicity of cationic surfactants,47 and the removing process of the surfactants can change the surface morphology and induce the aggregation of nanoparticles.48 In an effort to solve this problem, we have reported a seedless and surfactant-free synthetic method for bumpy silver NSs (AgNS) in the previous study.39-41 In the presence of


3


Page 4 of 32

alkylamine AgNSs were fabricated by reduction of silver ion on the silica surface without deposition of seed metals. Considering that the metal shell structures have their distinctive plasmon resonance in the visible and NIR region and the surface morphologies to a local hot spot for highly enhanced Raman scattering, controlling the local roughness and shell morphology is essential for active SERS substrates. Herein, we investigated the effects of alkylamines on the control local roughness of silver NSs as both a reducing and a capping agent. We found that just changing alkylamines in the synthesis of silver nanoshells alters morphologies and scattering properties of silver nanoshells altered easily with control of seeding and growth speed of silver domains without any additional synthetic steps. Especially, alkylamines with different chain lengths were employed to study the effects of chain length because alkyl group can influence the reduction of silver ion and the growth of silver domains. We also studied the effects of hydroxyl group and number of alkyl chains on the thickness of silver and the domain heterogeneity. The optical properties of the resulting silver NSs with different surface morphologies were characterized by surface-enhanced Raman spectra at the single-particle level. They were successfully applied as NIR-active SERS nanoprobes to target colon cancer biomarkers in vivo. These results indicate that the silver NSs have a great potential as a highly sensitive NIR-active SERS nanoprobes for use as a biological target detection tool.

2. Result and Discussion

For the one-step, surfactant-free synthesis of silver NSs, previously reported method was followed by using different kinds of alkylamines (Figure 1a). Briefly, silica nanoparticles (NPs) were synthesized as a dielectric core using the modified Stöber method,49 and their average diameter was 170 ± 15 nm. Silica surfaces were functionalized with thiol group to introduce the


4

Page 5 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


silver domain on the silica surface. Then, silver nitrate in ethylene glycol was reduced by alkylamine, resulting in the formation of silver domains on the silica surface. Because the reduction potential of silver ion in ethylene glycol is decreased by creating a silver ion/diol chelate, the silver shelling formation can be completed within 1 h at room temperature (Figure 1a). Alkylamine plays a crucial role in this reaction as both a capping ligand and reducing agent during the formation of silver NSs. Therefore, we hypothesized that using different kinds of alkylamines could control the reduction rate of silver ion on the silica surfaces. Figure 1b shows the seven kinds of alkylamines used to synthesize the silver NSs: Butylamine (C4), octylamine (C8), dodecylamine (C12), hexadecylamine (C16), ethanolamine (C2OH), propanolamine (C3OH), and tributylamine (tri-C4). These alkylamines were classified according to the following categories; alkyl chain length, existence of hydroxyl group, and number of alkyl chain. C4 was selected as a reference alkylamine for comparison. Firstly, we focused on the effect of the alkyl chain length by varying the number of carbons in the alkyl chain from 4 to 16 (C4, C8, C12, and C16 were used). The structure and surface morphology of silver NSs synthesized with C4, C8, C12, and C16 (named as ‘amine abbreviation-NS’, e.g. C4-NS for butylamine-silver NS) were observed by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) (Figure 2a-d). The overall surface morphology of C4-NS was round, bumpy structures in a complete silver shell. Interestingly, as the alkyl chain length became longer, the silver NS was formed with spiky shapes on the bumpy shell structure. The C16-NSs, in particular, had many spiky shapes on their surfaces. As shown in Figure 2e, C16-NS exhibited strong SPR from visible to near-infrared region. This strong SPR properties of C16-NS was due to their unique silver domain structures which strongly increase


5


Page 6 of 32

surface roughness and consequently lead to higher extinction than C4-NS. As a result, overall extinction of C16-NS is higher than C4-NS in all visible and NIR region. Next, we investigated the effect of reaction time on surface morphologies of silver NSs and the mechanism of their formation. A time-dependent silver shell formation of C4-NS and C16-NS was observed by TEM at each time intervals (1, 2, 5, 10, 15, 20, 30, and 60 min). Interestingly, in the case of C4-NS, a lot of silver domains covered the silica surface even after 1 min (Figure 3a). Silver shell formation was completed within 5 min, after which the shell thickness slightly increased. After 20 min, there was little change in the size. As shown in Figures 3b and c, the color of the C4NS solution and the UV-VIS-NIR extinction spectra were also monitored to evaluate the optical property change of C4-NS with reaction time. Along with the growth of C4-NS observed by TEM, a significant change in the color and extinction spectrum was observed and a strong SPR band began to appear in the NIR region after 5 min. In contrast, the silver shell growing process was slow in the case of C16-NS. After 10 min only a few silver NPs were attached to the silica surface and no silver shell had formed. The solution color was changed from transparent to dark and there was a distinct increase in NIR extinction after 20 min. Thereafter, heterogeneous spiky-bumpy shaped silver domains were formed and the silver shell forming process was completed (Figure 3d). These data indicate that there was a relatively slow nucleation and growth of the silver domain during C16-NS synthesis compared with C4-NS, which is attributable to the weak reducing power of the C16-amine compared with the C4-amine. To explain the different results of the silver shell formation of C4-NS and C16-NS, we propose two possible mechanisms related to the nucleation and growth speed of the silver domains (Figure 4). In the case of C16-NS, where the nucleation of silver domains occurred slowly, the grain of the silver domains grew less randomly and formed heterogeneous spiky-bumpy type structures rather


6

Page 7 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


than homogeneous bumpy structures. In the case of C4-NS, where the nucleation of silver domains occurred rapidly, the grain of the silver domains grew more randomly, causing the silver shell formation to occur all over the surface, so that the silver domain became relatively round shaped. In short, when a long alkylamine chain is used, the rate of nucleation and growth is slow and relatively heterogeneous spiky-bumpy shaped surface morphologies are formed. Conversely, when a short alkylamine chain is used, the rate of nucleation and growth is fast and their morphologies become bumpy rather than spiky-bumpy shaped. For verifying our hypothesis, additional experiment under increased temperature (50 ℃) was performed to verify temperature dependence of silver NSs formation. Figure S2 and S3 shows the TEM images and UV-Vis spectra of 4 different silver NSs with time. All data shows that silver NSs formation was faster in the higher temperature compared with room temperature. However, overall morphologies of silver NSs did not change compared with those at room temperature. This observation can be interpreted as that reduction kinetics from Ag+ to Ag0 are fastened by increasing temperature while nucleation and growth speed on the silica surface are not affected by increasing temperature since still different types of alkylamines regulate nucleation and growth speed of silver domains on the silica surface. For further verification of this hypothesis, we measured XRD patterns of the silver NSs to identify the facet information of the different silver NSs. Figure S4 shows the same pattern and indexes for the different silver NSs regardless of the alkylamines, indicating that morphology formation of silver NSs with different alkylamines is not affected by facets. In addition, we studied the effect of hydroxyl group containing alkylamines on morphology control of silver NSs by using C2OH and C3OH (C2OH-NS and C3OH-NS) and the results were compared with the structure of silver NSs (C4-NS). SEM images of those silver NSs showed that


7


Page 8 of 32

they all have bumpy surface morphologies (Figure S5). However, the overall size of each silver NS was different, indicating that the thicknesses of their silver shells are different, because they had the same sized silica core (Figure S6a-c). To calculate the silver shell thickness, we first measured the diameter of the silver NSs in the TEM images (n=20) and then subtracted the core diameter from the NS’s size (d=170±15 nm). According to the calculation data from Figure S6d, the silver shell became thinner when hydroxyl group containing alkylamines was used, probably because the hydroxyl group is more electronegative than the amine group and thus has a stronger inductive effect on alkylamine. The hydroxyl group containing alkylamine has lower basicity than the non-hydroxyl amine (the pKb of C4 is 3.22 and that of C2OH is 4.5)50 and the reduction power of the hydroxyl amine became weaker. As a result, the hydroxyl group slightly decreased the nucleation and growth process of silver domains compared with C4 amine, which caused the silver shell thickness become thinner than that of C4-NS (Figure S6f). We also investigated the effect of the number of alkyl chains in alkylamines on morphology control of silver NSs by using tri-C4. The SEM images revealed an overall bumpy surface morphology, but with some incomplete shell surfaces. In addition, the domain size of tri-C4-NS is more heterogeneous than those of other silver NSs (Figure S7a). To verify the heterogeneous nature of silver domain of tri-C4-NS, we measured the size of each 150 silver domain on the silver NSs from SEM images. Figure S7c represents the average domain sizes and standard deviations of C4-NS and tri-C4-NS, respectively. The data show that their average domain sizes are similar, but the size distribution of tri-C4-NS is much larger than that of C4-NS. This difference is attributable to the steric hindrance caused by three alkyl chains of tri-C4-NS. The amine group of tri-C4 amine is less exposed to the bulk phase than that of other single chain amines, which in turn reduces its reduction power. If the reduction process is fast, initial nucleation is followed by


8

Page 9 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


domain growth, resulting in a relatively homogeneous particle size distribution; if the reduction process is very slow, nucleation occurs along with the domain growth, resulting in a relatively heterogeneous particle size distribution.51 Therefore, various size of silver domains are observed in the case of tri-C4-NS and relatively homogeneous silver domains are observed in the case of C4NS (Figure S7d). To evaluate the SERS activity of each silver NS in the visible and NIR window, we conducted SERS measurements on seven kinds of silver NSs. After 4-fluorobenzenethiol (4-FBT) was adsorbed on the surface of each silver NS as a Raman labeling compound, the SERS intensity of the 4-FBT Raman band at 386 cm-1 was measured in solution phase. We first tested stability of each silver NSs by measuring SERS intensities with long term interval from fabricated date. Figure S8 shows SERS intensity profile of each Silver NSs with fabricated time. SERS intensities of each silver NSs did not change significantly although some decrease were obtained compared with original fabricated state. This result means our fabricated silver NSs have long-term stability because of well capping properties of alkylamines and PVP. Next, we measured SERS intensities of each silver NSs at three laser excitation levels (532, 660, and 785 nm). As shown in Figure 5a, C16-NS and C12-NS, with a spiky-bumpy type morphology, exhibited stronger SERS intensity at all laser excitation levels than the other types of silver NSs (Figure S9 and S10). This result is attributable to the structure of C16-NS and C12-NS, which consist of spiky-bumpy shaped silver domains or silver domains with some edges at the corners. Noble metal nanostructures with spiky-bumpy and edge structures can have a strong electric field with high SERS enhancement because the electromagnetic field at the end of the nanostructures’ spikes or edges is greatly enhanced, leading to strong SERS intensity of the Raman label compound.52-53


9


Page 10 of 32

Another notable property of the SERS activity of the silver NSs is the intense SERS signal produced when irradiated with a NIR rather than a visible laser-line. To further verify the enhanced scattering properties of each silver NS in the NIR region, we measured the SERS intensities of single silver NSs in solution phase (Figure 5a) and calculated their signal enhancement factor (EF). As shown in Figures 5b, S11, and S12, when the average SERS signal EFs of seven kinds of silver NSs (n=20) were compared, the SERS signal EFs of C16-NS and C12-NS were 10 times higher than those of the other silver NSs. The SERS signal EFs of C16-NS and C12-NS turned out to be very large considering that the average EF of single gold and silver nanospheres is ca. 104, and that of single nanostars and nanorods is up to 105 in the NIR region.16, 54 In addition, SERS signal EFs of C16-NS and C12-NS were much higher than single gold nanospheres or Au/Ag hollow structures which is measured by 4-FBT as Raman label molecules.55 Figure 5c shows the representative Raman mapping data of a single C16-NS for calculating the EF. SERS was measured using point-by-point mapping with a step size of 0.5 μm, an integration time of 1s, and 785 nm excitation. Then, SEM images were obtained with the same area of Raman mapping to confirm the presence of single silver NSs. The image of the SERS intensity map was overlaid with the SEM image of a single C16-NS, which identified that the SERS mapping corresponded to the site of each C16-NS, and the SERS EFs were calculated to be as greater than 107 in all cases. Other silver NSs, including C12-NS, C2OH-NS, and tri-C4-NS, also showed that the SERS mapping corresponded to the position of each silver NS and their SERS EFs were calculated as ranging 106~107 (Figures S13a, S14a, and S15a). The single C16-NS SERS spectra obtained from the Raman mapping data in Figure 5c are shown in Figure 5d. The data indicated that the SERS signals from C16-NSs are strong enough to be detected for further application in the NIR window (average SERS intensity of about 2000 counts per second), and verified that the


10

Page 11 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Raman mapping data corresponded to the SERS signal of 4-FBT molecules adsorbed on C16-NS. Data from other silver NSs also indicated that the Raman mapping data corresponded to a single silver NS SERS signal from 4-FBT molecules (Figures S13b, S14b, and S15b). As a feasibility test for bio-application of the silver NSs, C16-NS which has strong SERS properties in the NIR window was modified to detect biomarkers in vivo by using tatraspanin-8 (TSPAN8) antibodies against the human colon carcinoma (HCT8) cell in a xenograft mouse model. Before in vivo test, we already studied that the developed PEGylated AgNS did not cause any in vivo and in vitro toxicity as was verified by cytotoxicity tests.40 Figure 6a presents a schematic illustration of the surface modification on C16-NS. First, C16-NS nanoprobes which contain three different Raman label compound (RLC)s (4-FBT, 4-BBT, and 4-CBT) were prepared by adsorbing each RLC onto the C16-NS surfaces. To improve the biocompatibility and colloidal stability of the C16-NS surfaces, the C16-NSs were encapsulated with methoxypoly (ethylene glycol) sulfhydryl (mPEG-SH). PEG is a well-known surface coating material for preventing NP’s aggregation and degradation in biological organs or cells, and is used to evaluate their circulation time in vivo. Thus, 4-FBT treated C16-NS was encapsulated with mPEG-SH and mPEG(SH)-COOH mixtures, where the carboxyl group was utilized for conjugating C2 antibodies, one of the TSPAN8 antibody candidates, using the NHS/EDC coupling method. Prior to in vivo active targeting using C16-NS nanoprobes, we measured the SERS spectra of three kinds of C16-NS nanoprobes in PBS solution using a micro Raman system with 785 nm excitation. Figure 6b shows the spectra of the three C16-NS nanoprobes. Their distinctive Raman band positions (C16-NS4-FBT: 386 cm-1, C16-NS4-BBT: 488cm-1, C16-NS4-CBT: 541 cm-1) are proper for multiplex detection of biological samples. Next, we intravenously injected the three kinds of C16NS nanoprobes as a 1:1:1 mixture into the tail vein of a human colon cancer xenograft mouse and


11


Page 12 of 32

detected the SERS signal using a portable Raman system with 785-nm excitation. Thirty minutes after the injection, we obtained the 4-FBT SERS signal only from the tumor position and the three kinds of signal mixture spectra from the liver. These results indicate that all three C 16-NS nanoprobes were circulated properly in the blood vessels of the mouse, and specifically targeted the tumor position without any non-specific interaction in vivo. In addition, these data demonstrate that the silver NSs, especially C16-NS, can be used as an effective multiplex biomarker detection probe with strong NIR SERS or other optical properties in in vivo or in vitro analyses (Figure 6c). After in vivo detection, we extracted the tumor and liver from the xenograft mouse model and measured the SERS signals using the micro-Raman system with 785-nm excitation. Figure 6d shows the SERS spectra of the extracted tumor and liver and the data indicate that C16-NS nanoprobes were successfully injected into the blood circulation system of the xenograft mouse model and specifically targeted the tumor without any non-specific interaction in the tumor region, which was verified by the SERS signals of 4-FBT from the C2 antibody-conjugated C16-NS4-FBT probes.

3. Conclusion

We studied the effect of alkylamines on morphology control of silver NSs and their enhanced Raman scattering properties. We have proved that the surface morphology of silver NS particles can be easily controlled by varying the type of alkylamines, as well as the reduction speed of silver ion and the growth of the silver domains. Among them, C16-NSs have spiky-bumpy type morphologies and their scattering properties were high in the visible and NIR regions, when compared with the previously reported short alkyl chain treated silver NSs, which have bumpytype morphologies. Moreover, the SERS EF of the spiky-bumpy type silver NSs was calculated to


12

Page 13 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


be over 107, which is 10 times higher than the bumpy-type silver NSs under NIR excitation. For bio-application, we modified C16-NSs to highly NIR-active nanoprobes to detect cancer biomarkers in vivo, and successfully identified the targeted colon cancer without non-specific interactions. Our method of morphology control can be applied to other types of nanoparticles to enhance their optical properties, which can be utilized in the basic studies of surface chemistry of various nanostructures and for sensitive biological detection.

4. Experimental Section

Materials. Tetraethylorthosilicate (TEOS), ammonium hydroxide (NH4OH, 28-30%) 3mercaptopropyl trimethoxysilane (MPTS), ethylene glycol (EG), poly(vinyl pyrrolidone) (PVP, Mw ~40,000), silver nitrate (AgNO3, 99.999%), octylamine, butylamine, dodecylamine, hexadecylamine, tributylamine, ethanolamine and 3-amino-1-propanol, 4-fluorothiophenol (4FBT), 4-bromothiophenol (4-BBT), 4-chlorothiophenol (4-CBT), N-hydroxysuccinimide (NHS) and N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride (EDC) were purchased from Sigma-Aldrich (St. Louis, MO, USA) and used without further purification. Methoxypoly (ethylene glycol) sulfhydryl (m PEG-SH, Mw ≈ 5,000) was purchased from Sunbio (Anyang, Korea). Poly (ethylene glycol)-2-mercaptoethyl ether acetic acid (HOOC-PEG-SH, Mw 5,000) was purchased from Creative PEG Works (Winston Salem, NC, USA). Ethanol (99.9%) was bought from Daejung (Busan, Korea). Deionized (DI) water was used for the whole experiment. Synthesis of silver NS using different species of alkylamine. Tetraethylorthosilicate (TEOS, 1.6 mL) was dissolved in 40 mL of absolute ethanol (99.9%), and 3 mL of ammonium hydroxide (28-30%) was added to the reaction mixture. The resulting mixture was then stirred vigorously for 20 h at 25 ℃. The synthesized silica nanoparticles (silica NPs) were then centrifuged and washed


13


Page 14 of 32

with ethanol several times to remove excess reagents and dissolved in ethanol with 50 mg/mL concentration. Next, the silica NP surfaces were functionalized with a thiol group. Five milliliter of 50 mg/mL silica NPs (250 mg) was added to a reaction flask containing 250 μL of MPTS and 50 μL of aqueous ammonium hydroxide (28-30%). Then, this reaction mixture was stirred for 9 h at 25 ℃, and the resulting MPTS-treated silica NPs were centrifuged and washed with ethanol several times. A small quantity of the resulting MPTS-treated silica NPs was mixed with 25 mL of ethylene glycol, followed by the addition of 5 mg of PVP. Then, 25 mL of AgNO3 solution (in ethylene glycol) was mixed with silica NP solution. A varying amount of alkylamine (5 mM) was quickly added to the reaction mixture and the mixture was stirred for 1 h at 25 ℃. All silver NS were synthesized in round-bottomed glass flask. During the synthesis, reactions were carried out under room-light and air condition without further degassing process. Finally, synthesized silver NSs were centrifuged and washed several times with ethanol for purification. Instruments. The absorption spectra of the silver NSs were measured using a UV-Visible spectrometer (Optizen 2120UV, Mecasys). The morphology and size of the silver NSs were observed using a TEM (JEM1010, JEOL) and SEM (FE-SEM, JSM-6701F, JEOL). The SERS spectra were obtained using a confocal micro-Raman system (LabRam 300, JY-Horiba) equipped with an optical microscope (BX41, Olympus). In the micro-Raman system, back-scattering geometry was used for collecting the Raman scattering signal, and the signal was detected using a thermo-electrically cooled (-70 ℃) CCD detector. The excitation laser was focused and the Raman signals were collected using a ×10 and ×100 objective lens (Olympus, 0.90 NA). SERS Measurement. Seven kinds of Raman label-adsorbed silver NS suspensions were injected into capillary tube. Then, the SERS spectrum of each silver NS was measured three times by the Raman microscope system. This experiment was carried out using a ×10 objective lens with


14

Page 15 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


785-nm photoexcitation at 2.3 mW laser power and 10 s acquisition time. For single particle measurement, seven kinds of silver NS suspensions, pretreated with Raman label compounds onto their NS surface (0.1 mg/mL in ethanol), were dropped on a slide glass. Then, the SERS spectra were measured by point-by-point mapping with 0.5 μm step size using a ×100 objective lens (Olympus, 0.90 NA) with the same excitation source as used in the above experiment and using 1.1 mW laser power and 1 s acquisition time. After the experiment, the same mapping area was observed using field emission-scanning electron microscopy (FE-SEM). Intensities for calculating enhancement factor were determined by using average intensities of detecting pixels on mapping image around each particle. Calculation of the SERS Enhancement Factor: SERS enhancement factors (EF) for the silver NSs were estimated using the following equation: EF = (ISERS × Nnormal)/(Inormal × NSERS ), where ISERS and Inormal are the intensity of the bands from SERS and normal Raman scattering, respectively, and Nnormal and NSERS are the number of 4-FBT molecules in pure form and selfassembled on the silver NSs. The peak at 1075 cm− 1 (for 4-FBT) was used to estimate the EF. Raman signal intensity was measured for both single particles and neat 4-FBT using identical laser power for the EF calculation. Probing volume (18.8 μm3) was approximated as a cylinder form with a diameter of 2 μm and a height of 6 μm for the normal Raman measurements. NSERS was calculated by geometrical estimation of the particle’s surface area (silver NS is assumed to be a complete spherical shape, r = 125 nm) and a molecular footprint of 4-FBT (0.383 nm2/molecule), assuming the 4-FBT molecules formed a complete monolayer. Actual surface area of each silver NS is larger than the assumed surface area of spherical silver NS since silver NS has local bumpy structures. In order to estimate the variation of real surface area to the assumed spherical one, a model of many small particles on a spherical silver NS was considered. The number of the small


15


Page 16 of 32

particles and their average size was obtained from the TEM images of the local structure of silver NS (Figure 2). For the cases of C4-NS and Tri-C4-NS, the estimated variation of the surface area increased by ca. 25% and 35%, respectively (Table S1). This rough estimation implies that the EF value based on simple spherical model is over-estimated by the extent of surface area variation even though the variation is not significant. Preparation of PEGylated C16-NSs. A 1 mL portion of Raman label compound (4-FBT, 4BBT, 4-CBT, 2 mM in ethanol) was added to 1 mg of each silver NS and was incubated for 1 h at 25 ℃. The resulting silver NSs, which had adsorbed the Raman label compound on the silver domain surface, were centrifuged and washed with ethanol several times. To improve the application and biocompatibility of the silver NSs, their surfaces were coated with PEG. A 1-mL portion of COOH-PEG-SH (2 mM in ethanol) was mixed with 1 mg of 4-FBT-labeled silver NS and for 1 h. Finally, the coated silver NSs were centrifuged several times and redispersed in 0.1 M of phosphate buffered saline (PBS, 7.0). Preparation of conjugated C2 antibody onto PEGylated C16-NSs. Carboxylic acid– functionalized PEGylated silver NSs were activated with 2 mM of EDC and 5 mM of NHS in 0.1 M PBS (pH 6.0). Then, 20 μg of the antibody was added to the activated PEGylated silver NS dispersion. The mixture was shaken for 2 h at room temperature. The C2 antibody conjugated silver NSs were washed with 0.1 M of PBS (pH 7.0) and TPBS (PBS containing 0.1 wt of Tween20) several times. In vivo biomarker detection using an C16-NS nanoprobe in a human colon cancer xenograft mouse model. All animal studies were performed according to the protocols approved by the Institutional Animal Care and Use Committee of the Seoul National University. The HCT8 cells (0.1 mL in DMEM/matrigel 50% v/v) were harvested and injected into a nude mouse (OrientBio,


16

Page 17 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Sungnamro, Korea). Once the tumor diameter reached 1–1.5 mL, the mixture of C16-NS4-FBT, C16NS4-BBT, and C16-NS4-CBT was injected through the tail vein. The SERS spectra were obtained using an optical fiber-coupled portable-Raman system (B&W TEK, i-Raman) equipped with a 785 nm diode laser 30 min after injection of the C16-NS nanoprobes. Working distance between the fiber optic probe and the sample was adjusted to 6 mm. ASSOCIATED CONTENT Supporting Information. The Supporting Information is available free of charge on the ACS Publication Website. Additional information includes magnified SEM images of C16-NS, C12-NS, C8-NS and C4-NS, characterization data on the effect of alkylamine with hydroxyl group and length of alkyl chain, SERS spectra of silver NSs with different laser excitation, EF calculation data for C 2OH-NS, C3OH -NS and tri-C4-NS, and Raman mapping data for C12-NS, C2OH -NS, and tri-C4-NS. (PDF) AUTHOR INFORMATION Corresponding Author *E-mail: [email protected] *E-mail: [email protected] Author Contributions †

These authors contributed equally

Notes The authors declare no competing financial interest. ACKNOWLEDGMENT This work was supported by research funds from the Korea Health Technology R&D project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number: HI17C1264). REFERENCES


17


1.

Page 18 of 32

Kang, H.; Jeong, S.; Koh, Y.; Geun Cha, M.; Yang, J. K.; Kyeong, S.; Kim, J.; Kwak, S.

Y.; Chang, H. J.; Lee, H.; Jeong, C.; Kim, J. H.; Jun, B. H.; Kim, Y. K.; Hong Jeong, D.; Lee, Y. S., Direct Identification of On-bead Peptides Using Surface-enhanced Raman Spectroscopic Barcoding System for High-throughput Bioanalysis. Sci Rep 2015, 5, 10144. 2.

Zhang, H.; Liu, M.; Zhou, F.; Liu, D.; Liu, G.; Duan, G.; Cai, W.; Li, Y., Physical

Deposition Improved SERS Stability of Morphology Controlled Periodic Micro/Nanostructured Arrays Based on Colloidal Templates. 2015, 11 (7), 844-853. 3.

Zhang, T.; Sun, Y.; Hang, L.; Li, H.; Liu, G.; Zhang, X.; Lyu, X.; Cai, W.; Li, Y., Periodic

Porous Alloyed Au-Ag Nanosphere Arrays and Their Highly Sensitive SERS Performance with Good Reproducibility and High Density of Hotspots. ACS Appl Mater Interfaces 2018, 10 (11), 9792-9801. 4.

Zhang, T.; Zhou, F.; Hang, L.; Sun, Y.; Liu, D.; Li, H.; Liu, G.; Lyu, X.; Li, C.; Cai, W.;

Li, Y., Controlled Synthesis of Sponge-like Porous Au–Ag alloy Nanocubes for Surface-enhanced Raman Scattering Properties. Journal of Materials Chemistry C 2017, 5 (42), 11039-11045. 5.

Jeong, S.; Kim, Y. I.; Kang, H.; Kim, G.; Cha, M. G.; Chang, H.; Jung, K. O.; Kim, Y. H.;

Jun, B. H.; Hwang, D. W.; Lee, Y. S.; Youn, H.; Lee, Y. S.; Kang, K. W.; Lee, D. S.; Jeong, D. H., Fluorescence-Raman Dual Modal Endoscopic System for Multiplexed Molecular Diagnostics. Sci Rep 2015, 5, 9455. 6.

Kim, C.; Song, K. H.; Gao, F.; Wang, L. V., Sentinel Lymph Nodes and Lymphatic Vessels:

Noninvasive Dual-modality In vivo Mapping by Using Iindocyanine Green in Rats-Volumetric Spectroscopic Photoacoustic Imaging and Planar Fluorescence Imaging. Radiology 2010, 255 (2), 442-450.


18

Page 19 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


7.

Chen, Y.-S.; Frey, W.; Kim, S.; Kruizinga, P.; Homan, K.; Emelianov, S., Silica-Coated

Gold Nanorods as Photoacoustic Signal Nanoamplifiers. Nano Lett. 2011, 11 (2), 348-354. 8.

Cha, M. G.; Lee, S.; Park, S.; Kang, H.; Lee, S. G.; Jeong, C.; Lee, Y. S.; Kim, C.; Jeong,

D. H., A Dual Modal Silver Bumpy Nanoprobe for Photoacoustic Imaging and SERS Multiplexed Identification of In vivo Lymph Nodes. Nanoscale 2017, 9 (34), 12556-12564. 9.

Kang, B.; Afifi, M. M.; Austin, L. A.; El-Sayed, M. A., Exploiting the Nanoparticle

Plasmon Effect: Observing Drug Delivery Dynamics in Single Cells via Raman/Fluorescence Imaging Spectroscopy. ACS Nano 2013, 7 (8), 7420-7427. 10. Lee, J.; Jang, D.-J., Highly Efficient Catalytic Performances of Eco-Friendly Grown Silver Nanoshells. The Journal of Physical Chemistry C 2016, 120 (7), 4130-4138. 11. Bastús, N. G.; Comenge, J.; Puntes, V., Kinetically Controlled Seeded Growth Synthesis of Citrate-Stabilized Gold Nanoparticles of up to 200 nm: Size Focusing versus Ostwald Ripening. Langmuir 2011, 27 (17), 11098-11105. 12. Bastús, N. G.; Merkoçi, F.; Piella, J.; Puntes, V., Synthesis of Highly Monodisperse Citrate-Stabilized Silver Nanoparticles of up to 200 nm: Kinetic Control and Catalytic Properties. Chemistry of Materials 2014, 26 (9), 2836-2846. 13. Ye, X.; Jin, L.; Caglayan, H.; Chen, J.; Xing, G.; Zheng, C.; Doan-Nguyen, V.; Kang, Y.; Engheta, N.; Kagan, C. R.; Murray, C. B., Improved Size-Tunable Synthesis of Monodisperse Gold Nanorods through the Use of Aromatic Additives. ACS Nano 2012, 6 (3), 2804-2817. 14. John, C. L.; Strating, S. L.; Shephard, K. A.; Zhao, J. X., Reproducibly Synthesize Gold Nanorods and Maintain Their Stability. RSC Adv. 2013, 3 (27), 10909-10918.


19


Page 20 of 32

15. Cheng, L.-C.; Huang, J.-H.; Chen, H. M.; Lai, T.-C.; Yang, K.-Y.; Liu, R.-S.; Hsiao, M.; Chen, C.-H.; Her, L.-J.; Tsai, D. P., Seedless, Silver-induced Synthesis of Star-shaped Gold/silver Bimetallic Nanoparticles as High Efficiency Photothermal Therapy Reagent. J Mater Chem 2012, 22 (5), 2244-2253. 16. Khoury, C. G.; Vo-Dinh, T., Gold Nanostars For Surface-Enhanced Raman Scattering: Synthesis, Characterization and Optimization. The Journal of Physical Chemistry C 2008, 112 (48), 18849-18859. 17. Sauerbeck, C.; Haderlein, M.; Schürer, B.; Braunschweig, B.; Peukert, W.; Klupp Taylor, R. N., Shedding Light on the Growth of Gold Nanoshells. ACS Nano 2014, 8 (3), 3088-3096. 18. Sanchez-Gaytan, B. L.; Park, S.-J., Spiky Gold Nanoshells. Langmuir 2010, 26 (24), 19170-19174. 19. Xie, J.; Zhang, Q.; Lee, J. Y.; Wang, D. I. C., The Synthesis of SERS-Active Gold Nanoflower Tags for In Vivo Applications. ACS Nano 2008, 2 (12), 2473-2480. 20. Jena, B. K.; Raj, C. R., Seedless, Surfactantless Room Temperature Synthesis of Single Crystalline Fluorescent Gold Nanoflowers with Pronounced SERS and Electrocatalytic Activity. Chemistry of Materials 2008, 20 (11), 3546-3548. 21. Wang, Y.; Zheng, Y.; Huang, C. Z.; Xia, Y., Synthesis of Ag nanocubes 18-32 nm in Edge Length: The Effects of Polyol on Reduction Kkinetics, Size Control, and Reproducibility. J Am Chem Soc 2013, 135 (5), 1941-51.


20

Page 21 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


22. Klinkova, A.; Therien-Aubin, H.; Ahmed, A.; Nykypanchuk, D.; Choueiri, R. M.; Gagnon, B.; Muntyanu, A.; Gang, O.; Walker, G. C.; Kumacheva, E., Structural and Optical Properties of Self-assembled Chains of Plasmonic Nanocubes. Nano Lett 2014, 14 (11), 6314-21. 23. Ahn, J.; Wang, D.; Ding, Y.; Zhang, J.; Qin, D., Site-Selective Carving and Co-Deposition: Transformation of Ag Nanocubes into Concave Nanocrystals Encased by Au-Ag Alloy Frames. ACS Nano 2017. 24. Xia, X.; Zeng, J.; McDearmon, B.; Zheng, Y.; Li, Q.; Xia, Y., Silver Nanocrystals with Concave Surfaces and Their Optical and Surface-enhanced Raman Scattering Properties. Angew Chem Int Ed Engl 2011, 50 (52), 12542-6. 25. Liz-Marzán, L. M., Tailoring Surface Plasmons through the Morphology and Assembly of Metal Nanoparticles. Langmuir 2005, 22 (1), 32-41. 26. Mulvihill, M. J.; Ling, X. Y.; Henzie, J.; Yang, P., Anisotropic Etching of Silver Nanoparticles for Plasmonic Structures Capable of Single-Particle SERS. J. Am. Chem. Soc. 2009, 132 (1), 268-274. 27. Bardhan, M.; Satpati, B.; Ghosh, T.; Senapati, D., Synergistically Controlled Nanotemplated Growth of Tunable Gold Bud-to-blossom Nanostructures: a Pragmatic Growth Mechanism. Journal of Materials Chemistry C 2014, 2 (19), 3795-3804. 28. Guo, I. W.; Pekcevik, I. C.; Wang, M. C. P.; Pilapil, B. K.; Gates, B. D., Colloidal Coreshell Materials with 'Spiky' Surfaces Assembled from Gold Nanorods. Chem. Commun. 2014, 50 (60), 8157-8160.


21


Page 22 of 32

29. Chang, H.; Ko, E.; Kang, H.; Cha, M. G.; Lee, Y.-S.; Jeong, D. H., Synthesis of Optically Tunable Bumpy Silver Nanoshells by Changing The Silica Core Size and Their SERS Activities. RSC Advances 2017, 7 (64), 40255-40261. 30. Sanchez-Gaytan, B. L.; Swanglap, P.; Lamkin, T. J.; Hickey, R. J.; Fakhraai, Z.; Link, S.; Park, S.-J., Spiky Gold Nanoshells: Synthesis and Enhanced Scattering Properties. The Journal of Physical Chemistry C 2012, 116 (18), 10318-10324. 31. Sanchez-Gaytan, B. L.; Qian, Z.; Hastings, S. P.; Reca, M. L.; Fakhraai, Z.; Park, S.-J., Controlling the Topography and Surface Plasmon Resonance of Gold Nanoshells by a Templated Surfactant-Assisted Seed Growth Method. The Journal of Physical Chemistry C 2013, 117 (17), 8916-8923. 32. Oldenburg, S. J.; Averitt, R. D.; Westcott, S. L.; Halas, N. J., Nanoengineering of Optical Resonances. Chemical Physics Letters 1998, 288 (2), 243-247. 33. Wang, L.-X.; Jiang, X.-E., Bioanalytical Applications of Surface-enhanced Infrared Absorption Spectroscopy. Chinese Journal of Analytical Chemistry 2012, 40 (7), 975-982. 34. Jackson, J. B.; Halas, N. J., Silver Nanoshells: Variations in Morphologies and Optical Properties. The Journal of Physical Chemistry B 2001, 105 (14), 2743-2746. 35. Pedireddy, S.; Li, A.; Bosman, M.; Phang, I. Y.; Li, S.; Ling, X. Y., Synthesis of Spiky Ag–Au Octahedral Nanoparticles and Their Tunable Optical Properties. The Journal of Physical Chemistry C 2013, 117 (32), 16640-16649. 36. Zhou, N.; Li, D.; Yang, D., Morphology and Composition Controlled Synthesis of Flowerlike Silver Nanostructures. Nanoscale Research Letters 2014, 9 (1), 302-302.


22

Page 23 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


37. Wang, H.; Halas, N. J., Mesoscopic Au “Meatball” Particles. Advanced Materials 2008, 20 (4), 820-825. 38. Liu, Z.; Yang, Z.; Peng, B.; Cao, C.; Zhang, C.; You, H.; Xiong, Q.; Li, Z.; Fang, J., Highly Sensitive, Uniform, and Reproducible Surface-enhanced Raman Spectroscopy from Hollow AuAg alloy Nanourchins. Adv Mater 2014, 26 (15), 2431-9. 39. Chang, H.; Kang, H.; Yang, J.-K.; Jo, A.; Lee, H.-Y.; Lee, Y.-S.; Jeong, D. H., Ag Shell– Au Satellite Hetero-Nanostructure for Ultra-Sensitive, Reproducible, and Homogeneous NIR SERS Activity. ACS Appl. Mater. Interfaces 2014, 6 (15), 11859-11863. 40. Kang, H.; Yang, J.-K.; Noh, M. S.; Jo, A.; Jeong, S.; Lee, M.; Lee, S.; Chang, H.; Lee, H.; Jeon, S.-J.; Kim, H.-I.; Cho, M.-H.; Lee, H.-Y.; Kim, J.-H.; Jeong, D. H.; Lee, Y.-S., One-step synthesis of silver nanoshells with bumps for highly sensitive near-IR SERS nanoprobes. J. Mater. Chem. B 2014, 2 (28), 4415-4421. 41. Yang, J.-K.; Kang, H.; Lee, H.; Jo, A.; Jeong, S.; Jeon, S.-J.; Kim, H.-I.; Lee, H.-Y.; Jeong, D. H.; Kim, J.-H.; Lee, Y.-S., Single-Step and Rapid Growth of Silver Nanoshells as SERS-Active Nanostructures for Label-Free Detection of Pesticides. ACS Appl. Mater. Interfaces 2014, 6 (15), 12541-12549. 42. Liu, T.; Li, D.; Yang, D.; Jiang, M., An Improved Seed-mediated Growth Method to Coat Complete Silver Shells onto Silica Spheres for Surface-enhanced Raman Scattering. Colloids and Surfaces A: Physicochemical and Engineering Aspects 2011, 387 (1), 17-22.


23


Page 24 of 32

43. Phonthammachai, N.; Kah, J. C. Y.; Jun, G.; Sheppard, C. J. R.; Olivo, M. C.; Mhaisalkar, S. G.; White, T. J., Synthesis of Contiguous Silica−Gold Core−Shell Structures: Critical Parameters and Processes. Langmuir 2008, 24 (9), 5109-5112. 44. Liang, Z.; Susha, A. S.; Caruso, F., Metallodielectric Opals of Layer-by-Layer Processed Coated Colloids. Advanced Materials 2002, 14 (16), 1160-1164. 45. Dong, A. G.; Wang, Y. J.; Tang, Y.; Ren, N.; Yang, W. L.; Gao, Z., Fabrication of Compact Silver Nanoshells on Polystyrene Spheres Through Electrostatic Attraction. Chem. Commun. 2002, (4), 350-351. 46. Topete, A.; Alatorre-Meda, M.; Villar-Á lvarez, E. M.; Cambón, A.; Barbosa, S.; Taboada, P.; Mosquera, V., Simple Control of Surface Topography of Gold Nanoshells by a Surfactant-less Seeded-Growth Method. ACS Appl. Mater. Interfaces 2014, 6 (14), 11142-11157. 47. Alkilany, A. M.; Nagaria, P. K.; Hexel, C. R.; Shaw, T. J.; Murphy, C. J.; Wyatt, M. D., Cellular Uptake and Cytotoxicity of Gold Nanorods: molecular origin of cytotoxicity and surface effects. Small 2009, 5 (6), 701-8. 48. Iqbal, M.; Tae, G., Unstable Reshaping of Gold Nanorods Prepared by a Wet Chemical Method in the Presence of Silver Nitrate. Journal of Nanoscience and Nanotechnology 2006, 6 (11), 3355-3359. 49. Stöber, W.; Fink, A.; Bohn, E., Controlled Growth of Monodisperse Silica Spheres in the Micron Size Range. Journal of Colloid and Interface Science 1968, 26 (1), 62-69. 50. The Merck Index: An Encyclopedia of Chemicals, Drugs, and Biologicals. Wiley: 2006.


24

Page 25 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


51. Cao, G.; Wang, Y., Nanostructures and Nanomaterials: Synthesis, Properties, and Applications. World Scientific: 2011. 52. Lin, C.; Liu, Y.; Yan, H., Self-Assembled Combinatorial Encoding Nanoarrays for Multiplexed Biosensing. Nano Letters 2007, 7 (2), 507-512. 53. Baffou, G.; Quidant, R.; Girard, C., Heat Generation in Plasmonic Nanostructures: Influence of Morphology. Applied Physics Letters 2009, 94 (15). 54. Smitha, S. L.; Gopchandran, K. G.; Smijesh, N.; Philip, R., Size-dependent Optical Properties of Au Nanorods. Progress in Natural Science: Materials International 2013, 23 (1), 3643. 55. Kang, H.; Jeong, S.; Park, Y.; Yim, J.; Jun, B.-H.; Kyeong, S.; Yang, J.-K.; Kim, G.; Hong, S.; Lee, L. P.; Kim, J.-H.; Lee, H.-Y.; Jeong, D. H.; Lee, Y.-S., Near-Infrared SERS Nanoprobes with Plasmonic Au/Ag Hollow-Shell Assemblies for In Vivo Multiplex Detection. Advanced Functional Materials 2013, 23 (30), 3719-3727.


25


Page 26 of 32

Figure 1. Schematic illustration of a silver NS synthesis using various types of alkylamines. (a) Scheme of a seedless and one-step silver NS synthesis with different kinds of alkylamines. (b) Classification of each alkylamine used for synthesizing silver NSs and their chemical structure.


26

Page 27 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 2. Effect of alkyl chain length on the formation of silver NSs. TEM (upper panel) and SEM (lower panel) images of (a) C4-NS, (b) C8-NS, (c) C12-NS, and (d) C16-NS. (e) UV-Visible extinction spectra for four kinds of silver NSs.


27


Page 28 of 32

Figure 3. Time-dependent effect of the formation of silver NSs with different alkyl chains. (a) TEM images showing the growth of C4-NS as a function of reaction time. (b) UV-Visible spectra showing the growth of C4-NS as a function of reaction time. (c) Photograph of C4-NS solution at various reaction times from 1 min to 60 min showing the changes in each particle’s color. (d) TEM images showing the growth of C16-NS as a function of reaction time. (e) UV-Visible spectra showing the growth of C16-NS as a function of reaction time. (f) Photograph of C16-NS solution at various reaction times from 1 min to 60 min showing the changes in each particle’s color.


28

Page 29 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 4. Proposed reaction mechanisms of silver NS formation using long-alkyl chain and shortalkyl chain alkylamines.


29


Page 30 of 32

Figure 5. Analysis of enhanced scattering properties of silver NSs. (a) Comparison of the SERS intensities of the 4-FBT band at 386 cm-1 for SERS spectra obtained at different excitation wavelengths on four kinds of silver NSs with different alkyl chain lengths in ethanol solution. Intensities were normalized by the Raman intensity of the ethanol peak at 882 cm-1. SERS spectra were obtained using a micro-Raman system with 10-mW laser power and light acquisition time of 3 s. (b) Average enhancement factors of four kinds of silver NSs with different alkyl chain lengths. Twenty particles of each silver nanoshell were selected for enhancement factor calculation. (c) SERS intensity map of single C16-NSs at the 1075 cm-1 peak of 4-FBT. Each particle was dispersed in slide glass and SERS intensity map was overlaid with their corresponding SEM image. (d) SERS spectra of each 6 single C16-NSs which were obtained from SERS intensity map of (c). SERS spectra of (b) - (d) were obtained using a micro-Raman system by the 785-nm photoexcitation with 10-mW laser power and 1-s light acquisition time.


30

Page 31 of 32 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 6. In vivo multiplexing SERS detection and tumor targeting using C16-NSs. (a) Schematic illustration of surface modification of C16-NSs for in vivo multiplexing SERS detection and tumor targeting. (b) Three kinds of SERS spectra obtained from PBS solution containing C16-NSRLC, (c) SERS spectra of in vivo mouse injected with C16-NS using a tail-vein injection for tumor targeting and in vivo multiplexing. (d) SERS spectra of ex vivo tumor and liver of a mouse with C16-NS. (b) and (d) spectra obtained using a micro-Raman system with 785 nm photoexcitation at 33 mW laser power and light acquisition time of 1 s (in case of (b)) and 5 s (in case of (d)). (c) spectra obtained using a portable-Raman system with 785 nm photoexcitation at 90-mW laser power and light acquisition time of 30 s.


31


Table of Contents Graphic 490x295mm (150 x 150 DPI)


Page 32 of 32

Effect of Alkylamines on Morphology Control of Silver Nanoshells for

Recommend Documents