Inexpensive and Flexible SERS Substrates on Adhesive Tape Based

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Inexpensive and flexible SERS substrates on adhesive tape based on biosilica plasmonic nanocomposites Aysun Korkmaz, Maya Kenton, Gulsen Aksin, Mehmet Kahraman, and Sebastian Wachsmann-Hogiu ACS Appl. Nano Mater., Just Accepted Manuscript • DOI: 10.1021/acsanm.8b01336 • Publication Date (Web): 17 Aug 2018 Downloaded from http://pubs.acs.org on August 19, 2018

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ACS Applied Nano Materials

possible as well, in particular for the chemical characterization of biological particles of nano- to micro-meter sizes.

Keywords: plasmonics, Raman, SERS, biomaterials, diatoms, adhesive tape, bacteria

*Corresponding Authors [email protected] [email protected]

Introduction Surface-enhanced Raman scattering (SERS) is an emerging analytical technique used for the identification and characterization of chemical and biological molecules or structures that uses the enhancement of the Raman signal for molecules in near vicinity of plasmonic nanostructures to provide a spectroscopic fingerprint of these molecules of interest. 1-3 For enhancement factors as high as 10ଵସ , this technique allows us to reach single molecule sensitivity. enhancement is based on two mechanisms: electromagnetic enhancement

8-9

4-7

SERS

and chemical

enhancement.10-11 Electromagnetic enhancement is due to the electric field generated on the surface of noble metal nanostructures by excited surface plasmons (SPs). When a molecule interacts with the electric field generated on the surface, an intramolecular dipole moment is induced causing an increase in the polarizability of the molecule. As a result, an enhanced Raman signal is obtained.12 The magnitude of the electromagnetic enhancement depends on the plasmonic properties (intensity of the electric field and wavelength of the SPs) of the nanostructures. Chemical enhancement is due to charge transfer between metallic nanostructures

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and adsorbed molecules on plasmonic nanostructures. Its magnitude depends on the chemical structure of the molecule. Electromagnetic enhancement is the major component, with a typical enhancement factor of 104-107, while the contribution of chemical enhancement is limited to an enhancement factor of 10 - 102.12-13 Since electromagnetic enhancement is the major contributing mechanism, research focuses on targeted engineering of novel plasmonic structures to obtain high enhancement factors while maintaining reproducibility across the substrates.14-16 Up to date, gold (Au) and silver (Ag) plasmonic nanostructures are most commonly used as SERS substrates due to their high enhancement factors and the availability of plasmonic resonances in the visible and NIR regions. 17

Since electromagnetic enhancement is the major contributing mechanism to SERS

enhancement, the fabrication of novel plasmonic structures having high enhancement factors while also maintaining reproducibility across the substrates is of great interest. Colloidal suspensions of NPs are widely used for SERS substrates due to easy preparation and wide tunability of their plasmonic resonances. The significant drawbacks of using NPs as a SERS substrate are their limited enhancement factor due to the charge properties between the NPs and analytes, as well as having poor reproducibility depending on their aggregation status. To overcome these drawbacks, researchers are focusing on developing novel fabrication approaches that allow for better control the plasmonic properties of the fabricated nanostructures resulting in a overall higher and spatially more uniform enhancement factors. These structures include nanoholes,18-21 nanovoids,22-24 nanoclusters,25 and nanodomes.26-27 The nanostructures described in these articles have good reproducibility due to the well-defined structure of the substrates, but the approaches used here are, in general, time-consuming and require sophisticated laboratory equipment for the fabrication. In addition, although these well-defined


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nanostructures are capable of enhancing the SERS signals with good reproducibility, they still lack scalability, robustness, and are typically expensive. Photonic crystals (PCs) are capable of controlling and manipulating light using submicrometer scale arrays of dielectric materials.28-29 Recently, guided-mode resonances (GMRs) in photonic crystals (PCs) have attracted significant attention in SERS-related applications due to their potential for additional signal enhancement. GMRs can substantially intensify the localized electric fields (E) generated on the structures by optical coupling of the GMRs of the PCs and the LSPR of the NPs. 30-33 Furthermore, the fabrication of these complex photonic-crystal structures can only be done using expensive and time-consuming fabrication techniques such as top-down lithographic and reactive-ion etching.34 While the fabrication of these structures may be complex for humans, nature is able to provide us with an abundance of inexpensive photonic-crystals. Diatoms are unicellular marine microalgae that are enclosed in a cell wall composed of amorphous silica, called a frustule. These frustules possess hierarchical patterned nanostructured arrays, giving rise to photonic crystal properties.35 Recently, there has been great attention on diatoms for their potential applications in next generation biomedical applications, such as drug delivery, biosensing, and membrane technology.

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Furthermore, there is also evidence

showing that diatoms can be utilized in improving SERS enhancement by optically coupling the GMRs of the diatom frustules with the LSPRs of the nanostructures resulting in a lower limit of detection. In addition to this increase in enhancement, the large surface area of the porous diatom structure increases the chance of analyte attachment, which is significant for SERS-based biosensing.38 Recently, diatoms have been combined with metallic nanoparticles in order to add plasmonic properties to their photonic crystal properties, in particular for use as SERS substrates.38-44



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The first observation of additional SERS enhancement by porous biosilica nanocomposites (diatoms) was shown through self-assembly of AgNPs on the biosilica shell. The results of this study demonstrated that attaching metallic nanoparticles to a diatom results in an increase in the optical extinction by a factor of 2, and an increase in the sensitivity by a factor of 4, compared to assembled AgNPs on a glass structure. This additional enhancement is attributed to the coupling between LSPRs of the nanoparticles and the GMR of the diatom frustule.

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These unique

structural properties of diatoms make them suitable for SERS-based biosensing, and have attracted the interest of many groups. In another article, SERS substrates were prepared by modifying diatoms with APTES in order to obtain amino groups on the surface, enabling the attachment of different sized AgNPs.44 The results of this paper demonstrate that the successful coupling of the diatoms’ GMRs with the LSPs of the AgNPs results in an increase of the SERS signal. Yet another study reported SERS substrates that were prepared by modifying diatoms with APTES in order to obtain amino groups on the surface, enabling the attachment of different sized AuNPs.40 This composite, consisting of diatoms and self-assembled AuNPs, was pressed into tablets and used as SERS substrate. Other types of composites such as two biosilicananoparticle composites were also demonstrated in the literature.38 The first composite consisted of diatoms with self-assembled AuNPs. The second composite was prepared via in-situ growth: an AgNO3 solution was incubated with diatoms and then reduced using ascorbic acid to obtain AgNPs attached to diatoms. To prepare samples for SERS measurements, the biosilica composites were first dried onto glass slides, followed by dropping the analyte onto the slide. The results showed the capability of these composites for label-free SERS-based chemical and biological sensing.


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In terms of applications, a 3-D diatom-based hybrid plasmonic-biosilica SERS substrate was used for label-free detection of TNT. 41 The diatom-AgNP composite was prepared by the in-situ growth method, using the same procedure as in the previously mentioned paper, however an inkjet printer was added in order to minimize the amount of sample solution dispensed and maximize the Raman signal by concentration via evaporation.45 Other studies reported selfassembly of AuNPs of different sizes (20, 40, 60nm) on diatom frustules.

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The preparation of

the diatom-AuNP composites involved several steps. First, a diatom suspension was dropped on a glass slide and dried. The gold nanoparticles were then added on top of the dried diatoms, resulting in a diatom-AuNP composite substrate. Finally, the analyte was dropped onto the substrate, and SERS measurements were obtained. The maximum SERS enhancement was obtained when the 40nm AuNPs were used, with an enhancement factor of 1.66x107.

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As the

fabrication method is critical in determining the optical properties as well as the cost of the biosensor, research groups compared the enhancement factors of two AgNP-diatom structures fabricated via different techniques.38 Diatom-AgNP composites were fabricated via in-situ growth and via conventional self-assembly techniques. The diatom-AgNP composites fabricated using in-situ growth were shown to have a higher enhancement factor than the self-assembled diatom-AgNP composites by a factor of 2.5, due to its higher optical field enhancement.38 Other reported uses include a red silanized diatomaceous structure that was decorated with AuNPs to use for the detection of pesticides via SERS.43 In our study we fabricated an inexpensive, robust, flexible diatom-based SERS substrate, whose simplicity enables it to be used for a wide range of global health and global development applications. Prior to the preparation of the diatom-AgNP composites, a 2 mm wide template for the nanocomposite material was fabricated. After the template was constructed, two different



approaches were employed in the fabrication of the diatom-AgNP SERS substrate. The first approach involved obtaining pure diatom strips on office-grade transparent adhesive tape using the template, followed by the dropping of different volumes of AgNP suspension. The second approach involved the preparation of diatom-AgNP composites having different AgNP concentrations, which were then dried and prepared as strips on adhesive tape using the same template as used in the first method. The fabricated diatom-AgNP tape-based SERS substrates were characterized using UV-Vis spectroscopy and scanning electron microscopy (SEM). The SERS performance of the substrates was evaluated using 4-Aminothiophenol (4-ATP) and Rhodamine-6G. The composite strips with the highest SERS performance were used for evaluating their possible applications in biosensing, such as label-free protein and bacteria detection. Results and Discussion Fabrication of cheap, simple, robust and flexible SERS substrates having higher enhancement factors with higher reproducibility is significant for the development of label-free biosensors based on SERS detection. In this study, we fabricated an inexpensive, robust, flexible diatombased SERS substrate on a regular (office-grade) transparent adhesive tape. Schematic illustration of fabrication steps and photos of SERS substrate is shown in the Figure 1.


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A

Template for strips Filling template

Sticking tape on filled the template

Peeling off the tape from the surface

Obtaining the strips on the tape

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Figure 1. Schematic illustration (A) and photos (B) of fabrication of SERS substrates on a tape.

There are five fabrication steps for the fabrication of diatom-based SERS substrate. First step was the template preparation for the strips that allows to fill in diatoms or diatom-AgNPs composites. The template was then filled with the diatoms or diatom-AgNPs. Adhesive tape was then applied on the filled template. Finally, the tape was then peeled off from the surfaces to obtain strips on the tape. Two approaches were used to fabricate the diatom-based SERS substrates. First approach (as we will call it later on) is the obtaining diatom strips on the tapes, then the dropping different volume of AgNPs suspension to obtain diatom-based SERS substrate. Second approach (as we will call in later in the manuscript) is the preparation of diatom-AgNP composites having different concentration of AgNPs, then obtaining diatom-based SERS substrates on tapes. The photos of the preparation steps for the second approach were provided in the Supporting Information (SI) as Figure S1. The obtained SERS-active strips on the tapes were characterized using visual/photographic evaluation of the color as well as quantitative UV/Vis spectroscopy. Figure 2 shows the color changes of the diatom-AgNP composites (A) and the obtained strips (B) when different



concentrations of AgNPs are mixed with diatoms, and the absorbance spectra (C) of the obtained SERS-active strips on the tapes. Diatoms

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Figure 2. Color changes of diatoms and diatom-AgNP composites prepared using different volume of AgNPs (A), obtained strips using diatoms and these composites (B), and absorbance spectra of the prepared strips on the tapes (C).

It is clearly seen that, when the concentration of AgNPs was increased, the color of the composite become darker due to the increasing of the number of the AgNPs attached to the diatoms (see Figure 2A). SERS-active strips were prepared using bare diatoms and diatomAgNP composites having different concentration of AgNPs. Similar color changes were observed compared to composite powders (see Figure 2B). It is well known that plasmonic nanoparticles exhibit color due to their absorption and scattering properties. The color change of the composites demonstrates the presence (adherence) of the AgNPs to the composites. In addition to qualitatively estimating the color changes of the powders, we performed quantitative absorption measurements of the strips to demonstrate not only the presence of AgNPs but also determine the concentration of AgNPs (see Figure 2C). While there is no absorption maximum for the strip prepared using bare diatoms, the strips containing the composite diatoms-AgNPs have absorption maxima centered around 420 nm that are characteristic for AgNPs. The absorbance increases when the concentration of AgNPs was increased. However, when the volume of the AgNPs is 480 mL, a decrease in the absorbance was observed, which could be due to the dampening of the surface plasmons by aggregation of AgNPs. In order to evaluate microscopically the structure of the composite and adherence of the AgNPs to the surface of diatoms, we recorded SEM images that are shown in Figure 3. The strips imaged here were prepared by using 120 mL of AgNPs as an example.



Figure 3. SEM images of diatom-AgNP composite strip prepared by using 120 mL of AgNPs. The image on the left shows a large field of view that comprises numerous structures present in our substrate, and the image on the right is a magnification of the area marked with a rectangle.

As seen in these images, the diatoms used in our experiments are a random assortment of diatoms of different sizes and shapes, originating likely from different species. Some of the frustules are, in addition, incomplete or broken. It is known that the pore size of the diatoms is in the tens of nanometers to micrometers is size and therefore they are often not observed with optical microscopes. On the other hand, the SEM images shown in Figure 3 were obtained after depositing the sample with a thin (approximately 10nm) gold layer that improves the conductivity and allows for such images to be obtained, but it also masks the fine pore structure of the frustules. While heterogeneous in appearance, we believe that these diatomic structures still exhibit pores that can be used for SERS, but at a much lower cost. Nevertheless, intact frustules can be observed as the one highlighted in the right panel of Figure 3. Here we can also notice the presence of protrusions on the frustules, which are likely the AgNPs that are coating the surface


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of the porous silica. The frustules are already very heterogenous and display many broken parts. Due to this we do not observe changes when the substrate is bent or stretched.

Color changes and absorption spectra of SERS active strips prepared by dropping different volume of AgNPs (by the first approach) were provided in the SI as Figure S2. The color of the strip having lowest number of AgNPs (dropped lowest volume) is close to the color of pure diatoms. As we discussed above, when the volume of AgNPs was increased on the diatom strips, the color of the strip became darker due to the increasing of the number of the AgNPs (see Figure S2 A). The absorption band at around 420 nm was obtained from all strips having AgNPs due to the surface plasmons of the AgNPs (see Figure S2 B). The presence and concentration of AgNPs on the strips were demonstrated by changes in color and absorption spectra.

In order to understand the interaction between the AgNPs and diatomic frustules and the longterm stability of the composite, suspensions of 0.2 g of diatom powder, prepared composite powders having 6 different concentrations of AgNPs, and bare AgNPs dissolved in 10 mL of water (see Figure S3A) were compared. After initial preparation, the suspensions are opaque with colors depending on the concentration of the AgNPs, as seen in Figure S3A (the higher the concentration, the darker the suspension). After a weeklong sedimentation, the pure AgNPs are still dispersed in the water, while all other particles have precipitated at the bottom of the tubes. This indicates that binding between the AgNPs and frustules takes place, with the heavier diatoms dragging the metallic particles to the bottom (Figure S3B). UV/vis absorption spectra were obtained from top of the tubes after sedimentation in order to calculate the number of AgNPs present in the supernatants of the diatoms and composites (Figure S3C).



Diatoms suspension color was white when it was suspended in water. When the diatom-AgNP composites were prepared and dissolve them in water again, color of the composites having lower number of AgNPs was close to AgNPs. However, as the concentration of AgNPs was increased, the suspension color becomes darker. This is due to the aggregation of AgNPs to generate larger aggregates on the diatoms. All suspensions were allowed to precipitate for one week to evaluate the interaction and stability of composites. As seen in the Figure S3B, diatoms and all composites (diatom-AgNP) were settled down to the bottom of the falcon tubes. However, AgNPs suspension was still stable due to the repulsive force between negative charges due to the presence of citrate ion on the AgNPs. There were no significant supernatants color differences between diatom and composites. Absorption spectra were obtained from the all supernatants to calculate detached AgNPs by using absorbance of the synthesized AgNPs. It is well known that, when the AgNPs suspension was synthesized using citrate reduction method, the concentration of AgNPs is 1x1011 NP/mL. The absorbance could not be measured from the original suspension of AgNPs due to the exceedingly large number of AgNPs. Thus, the suspension was diluted 10 times (10x). The absorption spectra of all supernatants and 10xdiluted AgNPs are seen in the Figure S3C. The supernatant having highest detached NPs was the composite prepared by mixing 1g of diatoms and 480 ml of AgNPs. When the volume of AgNPs was decreased, the absorbance of supernatant (detached AgNPs) decreased as well. The absorbance value of the supernatants obtained at around 420 nm was used for the calculation of number of AgNPs on the diatom surfaces by comparing the absorbance of the 10x-diluted AgNPs. Percentages of the detached AgNPs were in the range of 0.2-2%. When the volume of AgNPs was increased, the percentage of the detached AgNPs increased as well. The maximum percentage was 2% for the diatom-AgNP composite prepared by using 480 mL of AgNPs. It


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means that, only approximately 10 mL of AgNPs detached from the surfaces of diatoms and 470 mL of AgNPs attached and became stable on the diatom surfaces. To remove the detached AgNPs from the suspension, there times washing steps were applied for all composites. Figure S4 show the color changes of the supernatants and absorption spectra after each centrifugation cycle, as an example. The color of the supernatant of the first cycle centrifugation was more yellowish due to the presence of AgNPs, however, when the 3-cycle centrifugation was done there was no color (see Figure S4A). Absorption spectra were obtained from the supernatants after each centrifugation (see Figure S4B) and the absorbance decreased after each centrifugation until no absorbance was measured after three washing cycles that seem, therefore, to be sufficient to remove the detached AgNPs for the composites. To further understand the interaction between the AgNPs and frustules, we performed zeta potential measurements of the suspensions of diatom, composites, and AgNPs. These measurements allow us to understand how the surface charge of frustules and AgNPs is affected by the binding between the two. The results are provided in the Table 1. Table 1. Zeta potentials of suspensions of diatom, composites and AgNPs.

Zeta potentials (mV) Diatoms

20 mL AgNPs

30 mL AgNPs

60 mL AgNPs

120 ml AgNPs

240 mL AgNPs

480 mL AgNPs

AgNPs

-30.72 (±1.24)

-32.93 (±1.83)

-33.82 (±1.67)

-34.85 (±3.49)

-36.43 (±1.43)

-38.03 (±3.74)

-41.85 (±1.44)

-50.81 (±2.88)



Zeta potentials of diatoms and AgNPs were at around -30 mV and -50 mV respectively, which are consistent with the literature. When the volume of AgNPs was increased to prepare the diatom-AgNP composites, increment in zeta potentials was obtained from -30 mV to -42 mV due to the increased number of AgNPs on the diatom surface. This is expected when binding between the two components takes place.

The composites having different nanoparticle concentrations were washed to assess the changes of the number of nanoparticles on the diatom surfaces. The color changes of composites powders and strips prepared using these composites and absorption spectra of the strips prepared washed and non-washed composites are provide in the SI as Figure S5. The color changes of the washed and non-washed composites and strips prepared using these substrates are not significant due to the low number of nanoparticles detached from the diatom surface during the washing steps. Absorption spectra were obtained to evaluate the quantification of nanoparticles on the strips prepared using washed and non-washed composites. It is seen that, the changes in the absorbance profile and intensity in not significant between the strips. The results demonstrate that, washing is not significant for the composite preparation. The composites can be used as prepared, with no additional washing step being required. We used two approaches for the fabrication of diatom-AgNP composite strips. In the first approach we prepare pure diatom strips and then drop AgNPs on the strips. During the evaporation of water, the AgNPs adhere to the diatom surfaces due to adsorption. For the second approach, diatoms and AgNPs are mixed and heated until evaporation of water promotes the adherence of the AgNPs on the diatom surfaces. In this way, bonding between the AgNPs and diatoms occurs without any additional linker. The stability of the mixtures, the observed color


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changes, the UV/Vis absorption spectra of strips and zeta potential measurements of suspensions of composites demonstrate that the adsorption of AgNPs is strong.

SERS spectra were obtained from the strips prepared using the two different approaches described earlier using 4-ATP as a test molecule. 4-ATP is a known test molecule for SERS substrates as it forms a uniform monolayer on the metallic surface. Figure 4 shows the reproducibility of SERS spectra of 4-ATP assembled on the strip prepared using first approach obtained from arbitrarily chosen ten spots (A), obtained from the strips prepared in different days (B), intensity changes of SERS spectra of 4-ATP obtained from the strips having different concentration of AgNPs (C), and intensity changes graph for the 1080 cm-1 Raman peak depending on the nanoparticles concentration on the strips (D). 140000

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Figure 4. SERS spectra of 4-ATP assembled on the strip obtained from arbitrarily chosen ten spots (A), obtained from the strips prepared in different days (B), intensity changes of SERS spectra of 4-ATP obtained from the strips having different concentration of AgNPs (C), and intensity changes graph at around 1080 cm-1 depending on the nanoparticles concentration on the strips (D).

As seen Figure 4 A, the reproducibility of the SERS spectra obtained from 4-ATP assembled on the strips is not good for SERS application. This could be due to aggregation of AgNPs randomly with different sizes on the diatom surfaces resulting in irreproducible SERS signal. The reproducibility of SERS spectra obtained from different strips prepared in different days (Figure 4B) is also not satisfactory for analytical SERS measurements. The increment in the intensity of the SERS signal is observed when the nanoparticle concentration (different volume) is increased.

Figure 5 shows the reproducibility of SERS spectra of 4-ATP assembled on the strip prepared using second approach obtained from arbitrarily chosen ten spots (A), obtained from the strips prepared in different days (B), intensity changes of SERS spectra of 4-ATP obtained from the strips having different concentration of AgNPs (C) and intensity changes graph at around 1080 cm-1 depending on the nanoparticles concentration on the strips (D).


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Figure 5. SERS spectra of 4-ATP assembled on the strip obtained from arbitrarily chosen ten spots (A), obtained from the strips prepared in different days (B), intensity changes of SERS spectra of 4-ATP obtained from the strips having different concentration of AgNPs (C), and intensity changes graph at around 1080 cm-1 depending on the nanoparticles concentration on the strips (D).

As seen Figure 5A, the reproducibility of the SERS spectra obtained from 4-ATP assembled on the strips is significantly better than in Figure 4A. The calculated percent coefficient of variation was %16, which is proper for SERS experiment. The reproducibility of SERS spectra obtained from different strips prepared in different days (Figure 5B) is also proper for SERS experiments. The average SERS intensity of 4-ATP is almost same obtained from the strips prepared in different days. Different SERS intensity was obtained assembled 4-ATP on the strips prepared using different from the volume of AgNPs (see Figure 5C). The increment in SERS intensity was



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obtained up to 240 mL of AgNPs. When the volume of AgNPs was increased for the preparation of composite, a lower SERS intensity was obtained. This is due to the more aggregation of AgNPs resulting in dampening surface plasmons of aggregates of AgNPs, which generate low SERS activity. Maximum SERS intensity was obtained when the strip prepared using diatomAgNP composite with 250 mL of AgNPs was used. The intensity changes at 1080 cm-1 was clearly seen in Figure 5C. A 50x microscope objective was used for the all data presented in Figure 5. When on the other hand a 20x microscope objective was used (with a larger sample volume being interrogated but a lower laser power density for excitation) the same trend was observed (provided in the SI as Figure S6.)

When comparing the reproducibility data presented in Figure 4A and Figure 5A it is important to note that they were obtained from strips prepared via the two different methods presented earlier. Figure 4A uses the first approach where the fabrication of diatom-AgNPs strips is by dropping of different volume of AgNPs on the pre-prepared pure diatom strips.

In this approach, the

aggregation of AgNPs is not controllable due to the drying process that occurs over time and large inhomogeneities are expected and therefore the results are less reproducible. However, for the second approach we prepared the composite first, then fabricate the SERS active strips. In this approach, we mixed AgNPs and diatom and heated until they dry. During this procedure, AgNPs are homogeneously distributed on the diatom surfaces and allow us to obtained more reproducible SERS spectra.

We start our study with relatively inhomogeneous frustules that were cheaper and easier to procure. Therefore we expect that our final substrate will exhibit a relatively irregular structure.


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However, we demonstrate that this is not a significant drawback for the reproducibility of the SERS measurements. We use a laser beam of about 1-2 micrometers in diameter, which averages the SERS signal over a large number of AgNPs and even several frustules. We believe this to be another reason for the observed reproducibility of the substrates prepared in the second way, while the substrates prepared with the first method exhibit a relatively low reproducibility.

In order to verify whether the tape substrate or the glue it contains contributes to the SERS spectra, we tested the strips with and without any analytes. The results are presented in the SI material (Figure S7) and indicate that there is no significant contribution from the substrate, likely due to the relatively thick layer of the diatom/AgNP layer (approximately 30 microns). In Figure S7 we show that, while we can measure a broad background spectrum without any analyze (red curve), there are no clear SERS peaks that we otherwise see when we have an analyte such as 4-ATP (blue curve). We also performed a stability check of the strip by incubating them for one hour with the 1 mM ATP ethanolic solution and noted no significant changes in the consistence of the strips.

The next step is to determine the performance of the strips in terms of SERS enhancement. For analytical measurements the consensus is that an analytical enhancement factor needs to be used, which is an average enhancement factor over many contributing plasmonic particles. We therefore calculated analytical enhancement factor (AEF) using the formula reported in the literature

46

: AEF=ISERS/IBulkxCBulk/CSERS. The peak at 1512 cm-1 was used to calculate the

enhancement factor for the strip having highest SERS intensity. Rhodamine 6G solutions at concentrations of 1.0x10-1 M and 1.0x 10-5 M are used for the bulk Raman and SERS



experiments, respectively, using laser at 785 nm. Figure 6 shows the spectra of bulk Raman and SERS spectra of rhodamine 6G dropped on a CaF2 slide and on the strip.

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Figure 6. Comparison of Raman and SERS spectra of Rhodamine 6G (0.1 M and 1.0x10-5 M) dropped on a CaF2 and on strip having highest SERS activity.

The concentration factor (the ratio of the Rhodamine 6G concentration used for spontaneous Raman vs SERS) was 1.0x104, and the intensity ratios were around 10. From here, maximum enhancement factors were found to be approximately 1.0x105. While this enhancement factor is relatively low compared with other structured reported in the literature, we would like to emphasize the this measurement is likely not very accurate due to dispersion of the small Rhodamine molecules within the pores of the frustules, which likely biases the measurement towards a lower enhancement factor. On the other hand, we expect significantly better results for


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samples that do not freely diffuse within the pores (such as is the case for biological objects of sizes larger than approximately 100 nm). The determine whether there is an SERS advantage for diatom-AgNP strips vs pure AgNP strips, an experiment was performed and the results are presented in the SI Figure S8. A solution of Rhodamine 6G was deposited on these strips and we note that the SERS activity of the composite strip is around 9 times higher than for the strip prepared using AgNPs. This additional enhancement compared with AgNPs alone is likely due to the combined LSPR and GMR contributions. The additional peaks observed at around 1000 cm-1 are attributed to AgNPs.

Next, the SERS tape strip having highest SERS enhancement was used for label-free biosensing of bacteria and proteins. The bacteria suspensions and protein solutions were dropped on the strips and obtained SERS spectra. 20 times diluted bacteria suspensions were dropped on the strips and obtained SERS spectra for the label-free characterization of bacteria. We also dropped the bacteria suspension on a strip prepared with pure diatoms, for comparison. SERS spectra were obtained from the S. aureus dropped both diatom-AgNP composite strip and diatom strip (provided in the SI as Figure S9). When the S. aureus was dropped on a diatom strip, the Raman spectrum is too week to be measured, as there is no Raman enhancement. However, when the S. aureus was dropped on a diatom-AgNP composite strip, the Raman spectrum is significantly more intense due to the presence of surface Raman enhancement and a SERS spectrum can be measured. SEM images of the S. aureus dropped on a strip were also provided in the SI as Figure S10. SEM images demonstrate that, after drying, the bacteria cover the strip surface and make contact with the diatom-AgNP composites thus enhancing the eventual Raman signal. Since surface coverage



seems relatively different depending on the imaged area we evaluated the effect of surface coverage on the SERS reproducibility of the spectra of bacteria. Ten SERS spectra obtained from arbitrarily chosen spots on the sample dropped S. aureus (see Figure S11). As seen in the Figure S11, the reproducibility of the SERS spectra of S. aureus is proper for SERS experiments. The surface coverage does not significantly affect the reproducibility of the spectra. As the next step in our evaluation of the ability of these nanocomposite strips to identify bacteria, we tested three different types of bacteria (S. aureus, E. coli, and Bacillus spp.) and two different strains of S. aureus (ATCC6538 and ATCC29213). SERS spectra obtained from the bacteria on the strips and PCA analysis plot for the bacteria are shown in Figure 7.


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Figure 7. SERS spectra of bacteria dropped on a strip (A) and PCA plot obtained from the SERS spectra of bacteria (B).



The differences of the recorded SERS spectra of three different types of bacteria with two strains of S. aureus are noticeable even by simple eye examination. The spectra of bacteria shown in the Figure 7A are the average of eight to ten spectra obtained from arbitrarily chosen spots under the laser. To better highlight the difference between strains and species, PCA analysis was performed on eight to ten spectra obtained from each sample. As seen in Figure 7B, 2 different strains of S. aureus clearly separate from each other. The three bacteria species are also well separated based on the SERS spectral differences obtained from the bacteria. The result demonstrates that the diatom-AgNP composite strip can be used as SERS active platform for the label-free characterization and identification of bacteria. Label-free detection of proteins dropped on the strip was also performed to evaluate the capability of the characterization of biological molecules on the strip using SERS. Figure 8 show the SERS spectra of model proteins dropped on the strips. The SERS spectrum of each protein is slightly different as noted by simple visual inspection.


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BSA Hemoglobin Lysozyme Cyt c

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Figure 8. SERS spectra of BSA, hemoglobin, lysozyme, and cytochrome c, dropped on the strip. All protein concentrations are 50. 0 µg/mL. Conclusions We demonstrated a method for simple fabrication of a plasmonic composite material that uses a readily available biomaterial (diatomic frustules) and metallic nanoparticles. This method further includes the placing of the composite on a flexible and inexpensive substrate (transparent, officegrade adhesive tape) that allows for the use in label-free sensing of biological samples such as proteins and bacteria. To our knowledge, this is the first report on a composite diatom-AgNP material that is SERS active on adhesive tape as well as the first report on using these strips for protein and bacteria measurements. The noted advantages of this SERS active platform come from the fact that the strips are stable and can be used for liquid samples, large areas can be easily fabricated, the analytical SERS enhancement factor is large (~105), the spectra are



reproducible (compared with drops), and less Ag material is used for the fabrication of strips than for other types of substrates. We performed a detailed characterization of the material including the identification of plasmonic resonances, determination of diatomic surface coverage with metallic nanoparticles, stability of the nanocomposite, and calculation of the analytical SERS enhancement factor. We then performed measurements of biologically-relevant materials such as proteins and bacteria and concluded that the sensitivity is sufficient for such measurements while preserving chemical specificity for separation of different bacteria species or strains of bacteria. The applications can be expended, however, to other biological entities such as cellular organelles (exosomes and other extracellular vesicles), viruses, or cells. Applications outside the diagnostics field can be envisioned as well, such as for detection of explosives, bioterrorism agents, pollutants, or food toxins. Due to the low cost of the materials, stability of the substrate, and simplicity of the preparation, these substrates may be used for applications in low resource environments.

Materials and Methods Materials Silver Nitrate (> 99%), Sodium citrate dihydrate (> 99%), Rhodamine 6G (dye content ~95%), 4-Aminothiophenol (97%), Celite(R) 209, and proteins were purchased from Sigma-Aldrich. Methods: Synthesis of AgNPs Synthesis of silver nanoparticles, as reported by Lee and Meisel,47 was synthesized by dissolving 90 mg of silver nitrate (AgNO3) in 500ml of water. This solution was then heated and stirred at 300 rpm until boiling. Once boiling, 10 ml of 1% sodium citrate dihydrate solution was added


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slowly to the AgNO3 solution. Solution was kept boiling for an additional hour. Color change from transparent to yellowish-green was observed, demonstrating nanoparticle formation. Fabrication of diatom-AgNP composite strips on office-grade transparent adhesive tape The first step in the fabrication of the diatom-AgNP composites strips involved the fabrication of a tape-based template. The template consists of several strips whose dimensions are 5 cm x 0.2 cm. After the template was constructed, two different approaches were employed in the fabrication of the diatom-AgNP composites strips. For the first approach, the template was filled with pure diatoms. Box-sealing adhesive tape was attached to the diatom-filled template, and after applying pressure to ensure the diatoms were attached properly to the tape, the tape was pulled off the template, resulting in diatom strips on the tape. To obtain the diatom-AgNP composite strips, different volumes of AgNPs (20 µL, 40 µL, 60 µL and 80 µL) were dropped on the prepared pure diatom strips. The second approach began by the preparation of diatom-AgNP composites of different AgNP concentrations. For this purpose, 1g of diatoms were mixed with different volume of AgNPs (20 mL, 30 mL, 60 mL, 120 mL, 240 mL, 480 mL), and then heated until all suspension water had evaporated. The composites were then dried in an oven for half an hour at 120 degrees Celsius, until dried completely. Once dried, these composites were used to fill the same template used in the first approach, obtaining strips of different AgNP concentrations on tape. A schematic illustration of these fabrication processes is shown in the Figure 1. The thickness of the strips was measured to be approximately 30 micrometers. Stability Test To assess the stability of the diatom-AgNP interaction, 8 different test tubes were prepared: 6 test tubes consisted of 0.2 g of each of the 6-different diatom-AgNP composites suspended in 10 mL of water, 1 tube consisted of 0.1 g of pure diatoms dissolved in water, and 1 tube contained the



AgNP suspension. After a week, the supernatant was collected and the absorbance of the solution measured in order to calculate the percentage of attachment of the AgNPs to the diatom surfaces. Zeta potential measurements were also performed using Malvern Zetasizer Nano ZS to assess the surface charge properties of the composite, which depend on the AgNP concentration on the diatom surfaces. Characterization of the nanocomposite material UV/Vis. Spectroscopy The absorbance of the fabricated diatom-AgNP composites on tape were measured using Perkin Elmer Lambda 750 instrument by scanning the absorption of the sample using absorbance wavelengths ranging from 300-800 nm. Scanning Electron Microscopy (SEM) SEM images were obtained using a Carl Zeiss SEM instrument at high vacuum with the acceleration voltage of 10 kV to obtain information regarding the morphology of the composite strips and the aggregation status of the silver nanoparticles. SERS measurements 4-ATP was used to evaluate the SERS performance of the fabricated diatom-AgNP composite strips. For this purpose, the fabricated strips were first treated with ATP (1 mM) dissolved in ethanol for an hour and then the composite surfaces were washed with ethanol and water. The SERS measurements were performed with a Renishaw InVia Reflex Raman Microscopy System (Renishaw Plc., U.K.) equipped with 785-nm laser. When the AgNPs form aggregates, the wavelength of the surface plasmons of the AgNPs broadens providing more flexibility in the excitation wavelength. While the fractal aggregates give the maximum enhancement due to the more possible hot spots in the aggregates, when the


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AgNPs form larger aggregates, the absorption band broadens and reduces the SERS activity due to the dampening of the surface plasmons. This is observed in our strips as well, as shown in the SI, Figure S12. A laser at 785 nm and a 50x magnification objective (numerical aperture: 0.75) with a laser power of 1.5 mW was used for all experiments. The exposure time and accumulation were 1 s and 1, respectively. To calculate the SERS enhancement factor of the strip having the highest SERS performance, two spectra were obtained. First, the Raman spectra of a 5 µL of Rhodamine 6G (0.1 M) solution dropped on a CaF2 slide was obtained. Then, 5µL of Rhodamine 6G (1.0 × 10−5 M) was dropped on a diatom-AgNP composite strips, and SERS spectra were obtained using 0.3 mW excitation from a 785nm laser excitation and a 50x magnification microscope objective (NA of 0.75) for excitation and collection of the spectra. Spectra of 4-ATP and Rhodamine 6G were obtained using an acquisition time of 1s and averaged over 3 accumulations, and were averaged over 10 different (randomly selected) spots on the sample. They were later used for the calculation of analytical enhancement factor of the SERS substrate. Strips having the highest SERS activity were then used for further experiments involving labelfree sensing of proteins and bacteria. Proteins (BSA, Hemoglobin, Cytochrome c and Lysozyme) with a concentration of 50 µg/mL were prepared. Diatom-AgNP composite strips were dipped in a solution of proteins for 30 minutes and SERS spectra were obtained after drying. The laser power, exposure time and accumulation were 1.5 mW, 1 s and 3, respectively. SERS spectra of proteins were the average of ten spectra acquired from randomly selected spots on the sample. Four model bacteria (S. aureus ATCC 6538, S. aureus ATCC 29213, E. coli ATCC 25322, and Bacillus spp.) were used to test the performance of the SERS active strip for the label-free characterization of bacteria



using SERS. The bacteria were inoculated from the stock solution of bacteria at 37 ˚C for 24 hours on Mueller Hinton Agar (MHA). The bacteria grown on MHA were collected with a plastic loop and washed three times with water via centrifugation. The bacteria collected in this way was then diluted 20 times before dropping on a strip and dropped 5 µL of bacteria suspension and waited to dry. The SERS spectra were obtained from the samples using a laser at 785 nm and a ×20 objective with a laser power of 3 mW. The exposure time and accumulation were 1 s and 1, respectively. The presented SERS spectra of model bacteria were the average of eight to ten spectra acquired from randomly selected spots on the sample. Principal Component Analysis Principal component analysis (PCA) was performed for Raman spectra of different bacteria (9 individual spectra for S. aureus ATCC 6538 and E. coli ATCC 25322, 10 individual spectra for S. aureus ATCC 29213 and 8 individual spectra for Bacillus spp., respectively) using PCA function in the software MATLAB R2017b. Different clusters are obtained for each sample along with PC1, PC2 and PC3 axes.

Supporting Information Details about the preparation of composite materials, characterization of the composites and strips, comparison with AgNP substrates, SERS characterization of the plasmonic structures, SEM characterization of the samples on the plasmonic structures.

Acknowledgments All authors would like to thank McGill University, Faculty of Engineering, for generous support. MK would also like to acknowledge NSERC support towards the SURE internship. We would also like to thank Ms. Juanjuan Liu for performing the PCA analysis.


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References 1. Albrecht, M. G.; Creighton, J. A., Anomalously Intense Raman-Spectra of Pyridine at a Silver Electrode. J Am Chem Soc 1977, 99 (15), 5215-5217. 2. Fleischmann, M.; Hendra, P. J.; Mcquillan, A. J., Raman-Spectra of Pyridine Adsorbed at a Silver Electrode. Chem Phys Lett 1974, 26 (2), 163-166. 3. Jeanmaire, D. L.; Vanduyne, R. P., Surface Raman Spectroelectrochemistry .1. Heterocyclic, Aromatic, and Aliphatic-Amines Adsorbed on Anodized Silver Electrode. J Electroanal Chem 1977, 84 (1), 1-20. 4. Blackie, E. J.; Le Ru, E. C.; Etchegoin, P. G., Single-Molecule Surface-Enhanced Raman Spectroscopy of Nonresonant Molecules. J Am Chem Soc 2009, 131 (40), 1446614472. 5. Kneipp, K.; Kneipp, H.; Kneipp, J., Surface-Enhanced Raman Scattering in Local Optical Fields of Silver and Gold Nanoaggregatess - From Single-Molecule Raman Spectroscopy to Ultrasensitive Probing in Live Cells. Accounts Chem Res 2006, 39 (7), 443-450. 6. Nie, S. M.; Emery, S. R., Probing Single Molecules and Single Nanoparticles by SurfaceEnhanced Raman Scattering. Science 1997, 275 (5303), 1102-1106. 7. Prinz, J.; Heck, C.; Ellerik, L.; Merk, V.; Bald, I., DNA Origami Based Au-Ag-Core-Shell Nanoparticle Dimers with Single-Molecule SERS Sensitivity. Nanoscale 2016, 8 (10), 56125620. 8. Moskovits, M., Surface-Roughness and Enhanced Intensity of Raman-Scattering by Molecules Adsorbed on Metals. J Chem Phys 1978, 69 (9), 4159-4161. 9. Moskovits, M., Surface-Enhanced Spectroscopy. Rev Mod Phys 1985, 57 (3), 783-826. 10. Otto, A., Surface-Enhanced Raman-Scattering - Classical and Chemical Origins. Top Appl Phys 1984, 54, 289-418. 11. Persson, B. N. J., On the Theory of Surface-Enhanced Raman-Scattering. Chem Phys Lett 1981, 82 (3), 561-565. 12. Haynes, C. L.; McFarland, A. D.; Van Duyne, R. P., Surface-Enhanced Raman Spectroscopy. Anal Chem 2005, 77 (17), 338a-346a. 13. Campion, A.; Kambhampati, P., Surface-Enhanced Raman Scattering. Chem Soc Rev 1998, 27 (4), 241-250. 14. Kahraman, M.; Mullen, E. R.; Korkmaz, A.; Wachsmann-Hogiu, S., Fundamentals and Applications of SERS-Based Bioanalytical Sensing. Nanophotonics-Berlin 2017, 6 (5), 831-852. 15. Avella-Oliver, M.; Puchades, R.; Wachsmann-Hogiu, S.; Maquieira, A., Label-free SERS Analysis of Proteins and Exosomes with Large-Scale Substrates from Recordable Compact Disks. Sensor Actuat B-Chem 2017, 252, 657-662. 16. Kahraman, M.; Daggumati, P.; Kurtulus, O.; Seker, E.; Wachsmann-Hogiu, S., Fabrication and Characterization of Flexible and Tunable Plasmonic Nanostructures. Sci RepUk 2013, 3.



17. Sharma, B.; Frontiera, R. R.; Henry, A. I.; Ringe, E.; Van Duyne, R. P., SERS: Materials, Applications, and the Future. Mater Today 2012, 15 (1-2), 16-25. 18. Brolo, A. G.; Arctander, E.; Gordon, R.; Leathem, B.; Kavanagh, K. L., NanoholeEnhanced Raman Scattering. Nano Lett 2004, 4 (10), 2015-2018. 19. Lee, S. H.; Bantz, K. C.; Lindquist, N. C.; Oh, S. H.; Haynes, C. L., Self-Assembled Plasmonic Nanohole Arrays. Langmuir 2009, 25 (23), 13685-13693. 20. Yu, Q. M.; Braswell, S.; Christin, B.; Xu, J. J.; Wallace, P. M.; Gong, H.; Kaminsky, D., Surface-Enhanced Raman Scattering on Gold quasi-3D Nanostructure and 2D Nanohole Arrays. Nanotechnology 2010, 21 (35). 21. Yu, Q. M.; Guan, P.; Qin, D.; Golden, G.; Wallace, P. M., Inverted Size-Dependence of Surface-Enhanced Raman Scattering on Gold Nanohole and Nanodisk Arrays. Nano Lett 2008, 8 (7), 1923-1928. 22. Baumberg, J. J.; Kelf, T. A.; Sugawara, Y.; Cintra, S.; Abdelsalam, M. E.; Bartlett, P. N.; Russell, A. E., Angle-Resolved Surface-Enhanced Raman Scattering on Metallic Nanostructured Plasmonic Crystals. Nano Lett 2005, 5 (11), 2262-2267. 23. Cintra, S.; Abdelsalam, M. E.; Bartlett, P. N.; Baumberg, J. J.; Kelf, T. A.; Sugawara, Y.; Russell, A. E., Sculpted Substrates for SERS. Faraday Discuss 2006, 132, 191-199. 24. Lang, X. Z.; Qiu, T.; Yin, Y.; Kong, F.; Si, L. F.; Hao, Q.; Chu, P. K., Silver Nanovoid Arrays for Surface-Enhanced Raman Scattering. Langmuir 2012, 28 (23), 8799-8803. 25. Ye, J.; Wen, F. F.; Sobhani, H.; Lassiter, J. B.; Van Dorpe, P.; Nordlander, P.; Halas, N. J., Plasmonic Nanoclusters: Near Field Properties of the Fano Resonance Interrogated with SERS. Nano Lett 2012, 12 (3), 1660-1667. 26. Choi, C. J.; Xu, Z. D.; Wu, H. Y.; Liu, G. L.; Cunningham, B. T., Surface-Enhanced Raman Nanodomes. Nanotechnology 2010, 21 (41). 27. Wu, H. Y.; Choi, C. J.; Cunningham, B. T., Plasmonic Nanogap-Enhanced Raman Scattering Using a Resonant Nanodome Array. Small 2012, 8 (18), 2878-2885. 28. John, S., Strong Localization of Photons in Certain Disordered Dielectric Superlattices. Phys Rev Lett 1987, 58 (23), 2486-2489. 29. Yablonovitch, E., Inhibited Spontaneous Emission in Solid-State Physics and Electronics. Phys Rev Lett 1987, 58 (20), 2059-2062. 30. Kim, S. M.; Zhang, W.; Cunningham, B. T., Coupling Discrete Metal Nanoparticles to Photonic Crystal Surface Resonant Modes and Application to Raman Spectroscopy. Opt Express 2010, 18 (5), 4300-4309. 31. Liu, C.; Wang, Z.; Li, E. W.; Liang, Z. X.; Chakravarty, S.; Xu, X. C.; Wang, A. X.; Chen, R. T.; Fan, D. L., Electrokinetic Manipulation Integrated Plasmonic Photonic Hybrid Raman Nanosensors with Dually Enhanced Sensitivity. Acs Sensors 2017, 2 (3), 346-353. 32. Xu, X. B.; Hasan, D. H.; Wang, L.; Chakravarty, S.; Chen, R. T.; Fan, D. L.; Wang, A. X., Guided-Mode-Resonance-Coupled Plasmonic-Active SiO2 Nanotubes for Surface Enhanced Raman Spectroscopy (vol 100, 191114, 2012). Appl Phys Lett 2012, 101 (5). 33. Zhao, Y.; Zhang, X. J.; Ye, J.; Chen, L. M.; Lau, S. P.; Zhang, W. J.; Lee, S. T., MetalloDielectric Photonic Crystals for Surface-Enhanced Raman Scattering. Acs Nano 2011, 5 (4), 3027-3033. 34. Cheng, C. C.; Scherer, A., Fabrication of Photonic Band-Gap Crystals. J Vac Sci Technol B 1995, 13 (6), 2696-2700. 35. Jeffryes, C.; Solanki, R.; Rangineni, Y.; Wang, W.; Chang, C. H.; Rorrer, G. L., Electroluminescence and Photoluminescence from Nanostructured Diatom Frustules Containing Metabolically Inserted Germanium. Adv Mater 2008, 20 (13), 2633-+. 36. Ragni, R.; Cicco, S.; Vona, D.; Leone, G.; Farinola, G. M., Biosilica from Diatoms Microalgae: Smart Materials from Bio-Medicine to Photonics. J Mater Res 2017, 32 (2), 279291.


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37. Albert, K.; Huang, X. C.; Hsu, H. Y., Bio-Templated Silica Composites for NextGeneration Biomedical Applications. Adv Colloid Interface Sci 2017, 249, 272-289. 38. Kong, X. M.; Squire, K.; Li, E. W.; LeDuff, P.; Rorrer, G. L.; Tang, S. N.; Chen, B.; Mckay, C. P.; Navarro-Gonzalez, R.; Wang, A. X., Chemical and Biological Sensing Using Diatom Photonic Crystal Biosilica With In-Situ Growth Plasmonic Nanoparticles. Ieee T Nanobiosci 2016, 15 (8), 828-834. 39. Chamuah, N.; Chetia, L.; Zahan, N.; Dutta, S.; Ahmed, G. A.; Nath, P., A Naturally Occurring Diatom Frustule as a SERS Substrate for the Detection and Quantification of Chemicals. J Phys D Appl Phys 2017, 50 (17). 40. Chen, J.; Qin, G. W.; Chen, Q.; Yu, J. Y.; Li, S.; Cao, F.; Yang, B.; Ren, Y. P., A Synergistic Combination of Diatomaceous Earth with Au Nanoparticles as a Periodically Ordered, Button-Like Substrate for SERS Analysis of the Chemical Composition of Eccrine Sweat in Latent Fingerprints. J Mater Chem C 2015, 3 (19), 4933-4944. 41. Kong, X. M.; Xi, Y. T.; Le Duff, P.; Chong, X. Y.; Li, E. W.; Ren, F. H.; Rorrer, G. L.; Wang, A. X., Detecting Explosive Molecules from Nanoliter Solution: A New Paradigm of SERS Sensing on Hydrophilic Photonic Crystal Biosilica. Biosens Bioelectron 2017, 88, 63-70. 42. Ren, F. H.; Campbell, J.; Wang, X. Y.; Rorrer, G. L.; Wang, A. X., Enhancing Surface Plasmon Resonances of Metallic Nanoparticles by Diatom Biosilica. Opt Express 2013, 21 (13), 15308-15313. 43. Zhang, H.; Sun, L.; Zhang, Y.; Kang, Y.; Hu, H. H.; Tang, H. R.; Du, Y. P., Production of Stable and Sensitive SERS Substrate Based on Commercialized Porous Material of Silanized Support. Talanta 2017, 174, 301-306. 44. Ren, F.; Campbell, J.; Rorrer, G. L.; Wang, A. X., Surface-Enhanced Raman Spectroscopy Sensors From Nanobiosilica With Self-Assembled Plasmonic Nanoparticles. IEEE J Sel Top Quantum Electron 2014, 20 (3), 6900806. 45. Kong, X. M.; Xi, Y. T.; LeDuff, P.; Li, E. W.; Liu, Y.; Cheng, L. J.; Rorrer, G. L.; Tan, H.; Wang, A. X., Optofluidic Sensing from Inkjet-Printed Droplets: the Enormous Enhancement by Evaporation-Induced Spontaneous Flow on Photonic Crystal Biosilica. Nanoscale 2016, 8 (39), 17285-17294. 46. Le Ru, E. C.; Blackie, E.; Meyer, M.; Etchegoin, P. G., Surface-Enhanced Raman Scattering Enhancement Factors: a Comprehensive Study. J Phys Chem C 2007, 111 (37), 13794-13803. 47. Lee, P. C.; Meisel, D., Adsorption and Surface-Enhanced Raman of Dyes on Silver and Gold Sols. J Phys Chem-Us 1982, 86 (17), 3391-3395.


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