Name: using extraordinary optical transmission to quantify cardiac biomarkers in human serum
Uploaded: 2017-12-13T12:30:00
Description: Watch this Scientific Journal Video about Using Extraordinary Optical Transmission to Quantify Cardiac Biomarkers in Human Serum at JoVE.com

AP acknowledges the support of Prof T Venkatesan, Director, NUS Nanoscience and Nanotechnology Initiative and Office of the Deputy President (National University of Singapore) (R-398-000-084-646). CLD acknowledges the support of the Singapore Ministry of Health National Medical Research Council under its clinician scientist funding scheme, NMRC/CSA/035/2012, and the National University of Singapore. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

1. Fabrication of the Sensor and Acquisition of the Data
<ol>
	<li>Preparation of the nickel mold
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		<li>Coat a 220 nm-thick layer of negative electron beam resist onto a 600 &#181;m-thick 4-in silicon wafer. Write the designed nanohole array on this wafer using an electron beam lithography system.
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			<li>To accelerate the e-beam writing, write the patterns with a low dotmap (N) of 20k for each 300 &#181;m field size (A) (i.e., there are 0.4 billion dots mapped on each 300 &#181;m<...

<ol>
	<li>Ebbesen, T. W., Lezec, H. J., Ghaemi, H., Thio, T., Wolff, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extraordinary+optical+transmission+through+sub-wavelength+hole+arrays.">Extraordinary optical transmission through sub-wavelength hole arrays.</a> Nature. 391 (6668), 667-669 (1998).</li><li>Krishnan, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evanescently+coupled+resonance+in+surface+plasmon+enhanced+transmission.">Evanescently coupled resonance in surface plasmon enhanced transmission.</a> Optics Comm. 200 (1), 1-7 (2001).</li><li>Yang, J. -. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+optical+transmission+mediated+by+localized+plasmons+in+anisotropic,+three-dimensional+nanohole+arrays.">Enhanced optical transmission mediated by localized plasmons in anisotropic, three-dimensional nanohole arrays.</a> Nano letters. 10 (8), 3173-3178 (2010).</li><li>Kim, J. H., Moyer, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transmission+characteristics+of+metallic+equilateral+triangular+nanohole+arrays.">Transmission characteristics of metallic equilateral triangular nanohole arrays.</a> Appl Phys Lett. 89 (12), 121106 (2006).</li><li>Liu, H., Lalanne, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscopic+theory+of+the+extraordinary+optical+transmission.">Microscopic theory of the extraordinary optical transmission.</a> Nature. 452 (7188), 728-731 (2008).</li><li>Shon, Y. -. S., Choi, H. Y., Guerrero, M. 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P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Designing+surface+plasmon+resonance+of+subwavelength+hole+arrays+by+studying+absorption.">Designing surface plasmon resonance of subwavelength hole arrays by studying absorption.</a> JOSA B. 29 (4), 521-528 (2012).</li><li>Ding, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+a+cardiac+biomarker+in+human+serum+using+extraordinary+optical+transmission+(EOT).">Quantification of a cardiac biomarker in human serum using extraordinary optical transmission (EOT).</a> PloS one. 10 (3), 0120974 (2015).</li><li>Im, H., Sutherland, J. N., Maynard, J. A., Oh, S. -. 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Simulating the interaction between incident light and the nanostructures makes it possible to identify the appropriate peak (in the transmission spectrum), whose shift can be recorded as a function of the concentration of the analyte. It is important to note that the localization of the bands with respect to the structure of the sensor is crucial to the choice of the right band, whose shift can be tracked to sense the analyte. The visualization can be achieved through simulations. This is also critical to the design of a...

Chemical sensors based on nanohole arrays have been a subject of numerous investigations since the first report on extraordinary optical transmission (EOT) was published by Ebbesen et al. in 19981. When light impinges on periodic arrays of nanohole structures of sub-wavelength dimensions, enhanced transmission occurs at specific wavelengths. This occurs when the incident light couples with Bloch-wave surface polariton (BW-SPP) and/or localized surface plasmons (LSP)2.
The underlying physical principle exploited when biosensing with such periodic arrays is simple. Adsorption of molecules onto or near the interface of metal changes the dielectric constant of the medium in contact with the metal, in turn shifting the location of the transmission bands in the spectrum. The spectrum itself can be adjusted by nano-engineering the shape, size, and separation distance3,4,5. By design, sensors based on EOT have characteristic bands in their spectra that facilitate specific assignments6,7,8 during the investigation of molecular binding events. This is a crucial advantage over commercially available surface plasmon resonance (SPR) platforms.
Sensors using EOT typically involve a light source optically aligned such that a collimated beam is incident on the sensing surface. Techniques to generate large nanohole surfaces, such as co-polymer templates and interference and nanosphere lithography, have poor reproducibility9. Due to these limitations in accurately fabricating large surfaces that show the EOT phenomenon, an optical microscope was required to correctly position the light source and detector. To simplify the technique, high-quality nanoimprinting lithography (NIL)10 was employed. This enabled the production of large sensor surface areas11 (mm-scale), removing the need for a microscope to look for the sensing surface on a chip. Instead, this sensor could be easily interfaced with a standard fiber optic cable.
Since the transmission peaks for this nanohole array are contained in the visible to near-infrared region (NIR), it is perfectly suited to sensing binding events for biomolecules in an aqueous environment. The expected optical behavior of the nanohole array was simulated. The result was then verified through studies with liquids of standard refractive indexes (RI). This array was then used to measure the concentration of cardiac troponin I (cTnI) in the complex background of human serum. cTnI is the clinical gold standard for the diagnosis of acute myocardial infarction.
Using this sensor, it is possible to detect and quantify cTnI in human serum at a limit of detection (LOD) of 0.55 ng/mL, which is clinically relevant. The detection is much quicker than the most commonly used technology in this domain, enzyme-linked immunosorbent assay (ELISA). Furthermore, the sensing surface can easily be regenerated and therefore reused. Hence, this work demonstrates the promise of nanohole arrays as a viable point-of-care (POC) technology for biosensing within complex biofluids.

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			<td height="43" style="height:43px;width:199px;">Electron Beam Lithography setup</td>
			<td style="width:220px;">Elionix ELS 700</td>
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			<td height="22" style="height:22px;width:199px;">o-Xylene</td>
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			<td height="22" style="height:22px;width:199px;">Photo-curable NIL resist</td>
			<td style="width:220px;">micro resist technology GmbH</td>
			<td style="width:199px;">mr-UVCur21-300nm</td>
			<td style="width:91px;"></td>
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			<td height="22" style="height:22px;width:199px;">Light Curing System</td>
			<td style="width:220px;">Dymax&#160;</td>
			<td style="width:199px;">Model 2000 Flood</td>
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			<td height="22" style="height:22px;width:199px;">E-beam deposition machine</td>
			<td style="width:220px;">Denton Explorer</td>
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			<td height="43" style="height:43px;width:199px;">UV-visible spectrometer&#160;</td>
			<td style="width:220px;">Ocean optic HR2000+ (Dunedin, FL, USA)</td>
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			<td height="43" style="height:43px;width:199px;">Standard refractive index liquids&#160;</td>
			<td style="width:220px;">Cargill Inc (Cedar Grove, USA)</td>
			<td style="width:199px;">18032</td>
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			<td height="22" style="height:22px;width:199px;">Plotting software</td>
			<td style="width:220px;">Origin</td>
			<td style="width:199px;">Origin Pro 9</td>
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			<td height="22" style="height:22px;width:199px;">10-carboxy-1-decanethiol&#160;</td>
			<td style="width:220px;">Dojindo Laboratories (Japan)</td>
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			<td height="22" style="height:22px;width:199px;">1-octanethiol&#160;</td>
			<td style="width:220px;">Sigma-Aldrich, MO, USA</td>
			<td style="width:199px;">471386</td>
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			<td style="width:220px;">BioRad (CA, USA)</td>
			<td style="width:199px;">1762410</td>
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		<tr height="22">
			<td height="22" style="height:22px;width:199px;">Anti-troponin antibody 560</td>
			<td style="width:220px;">Hytest (Finland)</td>
			<td style="width:199px;">4T21</td>
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			<td height="22" style="height:22px;width:199px;">Ethanolamine-HCl solution</td>
			<td style="width:220px;">BioRad (CA, USA)</td>
			<td style="width:199px;">1762450</td>
			<td style="width:91px;"></td>
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			<td height="43" style="height:43px;width:199px;">Surface Plasmon Resonance setup</td>
			<td style="width:220px;">BioRad XPR36 (Haifa, Israel)</td>
			<td style="width:199px;"></td>
			<td style="width:91px;"></td>
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			<td height="22" style="height:22px;width:199px;">Multiplexed SPR chip</td>
			<td style="width:220px;">BioRad</td>
			<td style="width:199px;">GLC</td>
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			<td height="22" style="height:22px;width:199px;">Human cTnI standard</td>
			<td style="width:220px;">Phoenix Pharmaceuticals</td>
			<td style="width:199px;">EK -311-05</td>
			<td style="width:91px;"></td>
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			<td height="22" style="height:22px;width:199px;">Glycine-HCl</td>
			<td style="width:220px;">BioRad (CA, USA)</td>
			<td style="width:199px;">1762221</td>
			<td style="width:91px;"></td>
		</tr>
	</tbody>
</table>

using extraordinary optical transmission to quantify cardiac biomarkers in human serum

For a biosensing platform to have clinical relevance in point-of-care (POC) settings, assay sensitivity, reproducibility, and ability to reliably monitor analytes against the background of human serum are crucial.
Nanoimprinting lithography (NIL) was used to fabricate, at a low cost, sensing areas as large as 1.5 mm x 1.5 mm. The sensing surface was made of high-fidelity arrays of nanoholes, each with an area of about 140 nm2. The great reproducibility of NIL made it possible to employ a one-chip, one-measurement strategy on 12 individually manufactured surfaces, with minimal chip-to-chip variation. These nanoimprinted localized surface plasmon resonance (LSPR) chips were extensively tested on their ability to reliably measure a bioanalyte at concentrations varying from 2.5 to 75 ng/mL amidst the background of a complex biofluid-in this case, human serum. The high fidelity of NIL enables the generation of large sensing areas, which in turn eliminates the need for a microscope, as this biosensor can be easily interfaced with a commonly available laboratory light source. These biosensors can detect cardiac troponin in serum with a high sensitivity, at a limit of detection (LOD) of 0.55 ng/mL, which is clinically relevant. They also show low chip-to-chip variance (due to the high quality of the fabrication process). The results are commensurable with widely used enzyme-linked immunosorbent assay (ELISA)-based assays, but the technique retains the advantages of an LSPR-based sensing platform (i.e., amenability to miniaturization and multiplexing, making it more feasible for POC applications).

The optical setup for taking measurements is shown in Figure 1A. An image of the actual nanohole array is given in Figure 1B. To understand the physics driving the sensing process, the COMSOL simulation software was used to simulate the distribution of the plasmonic field in an aqueous environment. The results from the simulation were then related to the actual measurement. A previously published study contains details of the ass...

Watch this Scientific Journal Video about Using Extraordinary Optical Transmission to Quantify Cardiac Biomarkers in Human Serum at JoVE.com

Using Extraordinary Optical Transmission to Quantify Cardiac Biomarkers in Human Serum

This work describes a nanoimprinting lithography method to fabricate high-quality sensing arrays that work on the principle of extraordinary optical transmission. The biosensor is low-cost, robust, easy to use, and can detect cardiac troponin I in serum at clinically relevant concentrations (99th percentile cutoff &#8764;10-400 pg/mL, depending on the assay).

For a biosensing platform to have clinical relevance in point-of-care (POC) settings, assay sensitivity, reproducibility, and ability to reliably monitor ...

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