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Nanoarrays: Self-assembled Hotspots for Enhanced Analyte Detection

Periodic Reporting for period 1 - N-SHEAD (Nanoarrays: Self-assembled Hotspots for Enhanced Analyte Detection)

Reporting period: 2015-04-08 to 2017-04-07

The three most significant challenges facing biosensing are inaccuracy, insensitivity, and low-throughput detection. One technique that is capable of facing these challenges is SERS which has demonstrated potential for extreme sensitivity and rapid, multiple-analyte detection within complex mixtures. Early stage diagnosis of disease requires the detection of trace amounts of analyte in multi-component biological samples (blood, urine, saliva). It is therefore particularly important for sensors to be reach low detection limits.

The objective of the action was to exploit the self-assembly of nanoparticles (NPs) at liquid-liquid interfaces (LLI) to produce a 2D homogeneous array of plasmonic NPs for ultrasensitive surface enhanced Raman (SERS) sensing. To achieve this we had several goals; to assemble spherical and shaped plasmonic NPs at the LLI and to test their Raman response and potential for sensing of colorectal cancer.

We investigated the assembly of spherical and shaped NPs at the LLI, working towards more control over their assembly and understanding their structure. Through in situ manipulation of the NP structure and spacing within the film we were able to observe the direct effects of plasmon coupling on the Raman and optical response of the arrays. Through a collaborative project, NPs were driven to the LLI through electrochemical means. The process was reversible. These NPs were only 16 nm in diameter but there is potential to apply this research to larger NPs more suitable for SERS experiments. A NP immunoassay was constructed for the detection of carcinoembryonic antigen (CEA), a biomarker for the presence of colorectal cancer. We were able to detect down to 10 ng/mL (a concentration above 10 ng/mL indicates that colorectal cancer may be present). The detection was specific, with no signal seen in the control sample. The duration of the test was also faster than conventional immunoassays (requires two incubation steps of 24 h). We were able to detect CEA after only 2 h incubation on both steps.
We assembled small gold NPs at the LLI. The films were analysed with x-ray reflectivity and GISAXS to determine the coverage and interparticle spacing of the NPs at the interface and correlated with optical reflectance measurements in order to produce a “plasmon ruler”.
Large (42 nm) gold NPs were assembled at the LLI. The citrate concentration in the aqueous phase was varied to control the assembly of the NPs. I studied the optical properties of the films made at different citrate concentration with transmission spectroscopy. The spacing between the NPs could be increased or decreased in situ depending on whether the amount of citrate was increased or decreased respectively. The reversibility of this process was studied.
The 42 nm gold NPs were then tagged with a Raman reporter (4-MBA) and similar modulation of the spacing with citrate concentration was carried out while simultaneously monitoring the SERS and optical response.
I performed a study on the assembly of gold nanorods at the LLI where I decreased the spacing between the rods in situ by adding acid to the organic phase. During this transition to a close packed GNR array, I monitored the optical and SERS response of 4-MBA which was adsorbed onto the GNRs. The Raman signal increased 40 fold.
Shaped NPs were assembled at the liquid-air interface (LAI). The arrays formed at the LAI were extracted and dried onto glass substrates to form a device. Their Raman and optical response was measured, determining the optimum geometry and composition of NPs which presented enhanced Raman signals. Spherical gold NPs, nanorods, stars, triangles and spherical silver NPs were studied.
We investigated the detection of CEA, a biomarker for colorectal cancer. A NP sandwich immunoassay approach was used using crumpled gold substrate with silver NPs tagged with a Raman reporter. To form the sandwich immunoassay, the gold substrates were incubated for 2h in serum or buffer samples containing CEA down to concentrations of 10 ng/mL. A concentration above 10 ng/mL indicates that colorectal cancer may be present. The CEA binds to the CEA antibody on the gold substrate. The substrate is them incubated in the silver NP solution. Due to the CEA antibody on the surface of the silver NPs they can bind with the CEA protein attached to the gold surface. Only samples containing CEA presented a signal for the 4-MBA tagged NPs.

Exploitation and dissemination of results:
D1.2 “Tuneable 2D self-assembly of plasmonic nanoparticles at liquid | liquid interfaces. Nanoscale 2016, 8, 19229-19241.”
D1.3 Manuscript under preparation “The effect of citrate on the assembly of gold nanoparticles at liquid-liquid interfaces. 2017.”
D1.4 Manuscript under preparation: “Controlling the SERS response of 2D gold nanoparticle films at the liquid-liquid interface. 2017.”
D1.5 Manuscript (submitted to Nature Materials): “Electrotuneable Nanoplasmonic Liquid Mirror, 2017.”
D2.1 Manuscript under preparation. “A simple, fast, upscalable method to assemble plasmonic NPs into arrays at a liquid interface for application as SERS substrates, 2017”
D 2.2 Publication (accepted), “Monitoring plasmon coupling and SERS enhancement through in situ nanoparticle spacing modulation, Faraday Discussions, 2017. Article ID: FD-ART-05-2017-000162.R1”
Seminars and conferences:
Oral presentation at the SERS Faraday Discussions, 2017 (Glasgow, UK). “Monitoring plasmon coupling and SERS enhancement through in situ nanoparticle spacing modulation”
London Centre of Nanotechnology seminar series, Imperial College London, 2017. “Tuneable 2D self-assembly of plasmonic nanoparticles at liquid interfaces”
King’s College London, Physics department, invited seminar, 2016. “Precise control on the self-assembly of plasmonic nanoparticles at liquid interfaces tailored for SERS applications”
Through this project we have created novel 2D NP architectures which are of great interest to a variety of fields spanning, but not limited to; optics (mirrors and filters), plasmonics, sensing (SERS and optical based), and catalysis. The assembly mechanisms were thoroughly investigated in order to provide utmost control over NP assembly, allowing the design of homogeneous and reproducible structures that could form the basis of a device. Through fine tuning the spacing between the NPs we were able to correlate optical features (plasmon resonance bands) with the spacing as determined from GISAX. Thereby forming a plasmon ruler where the researcher merely has to measure the reflectance or absorbance of the array to determine the interparticle spacing. Through monitoring the optical properties of the arrays as the interparticle spacing was changed, we were able to reveal new aspects on plasmon coupling processes between NPs spanning large arrays.
Through preliminary research on the detection of colorectal cancer biomarkers we were able to detect CEA within the limits where CEA becomes elevated due to the possible presence of colorectal cancer. The immunoassay was specific, only presenting a signal when CEA was present. The test was faster than typical immunoassays (24h each incubation). We incubated for 2 h. Of particular significance is that the test takes a blood sample, patient compliance with stool sampling tends to be low and so more cases of colorectal cancer will be caught early by providing a blood test.