Short communicationLabel-free antibody–antigen binding detection by optical sensor array based on surface-synthesized gold nanoparticles
Introduction
Nanoparticle-based biosensors have been suggested as a possible alternative to continuous-gold surface plasmon resonance sensors (McFarland and Duyne, 2003, Stuart et al., 2004). The strong optical extinction in the visible region of the electromagnetic spectrum exhibited by noble metal nanoparticles arises when the incident photon frequency is in resonance with a localised surface plasmon mode in the conduction band of the nanoparticles. The optical extinction has a contribution from both scatter and absorption with scattering dominating when spherical particles have a diameter greater than approximately 25 nm. The optical properties of the nanoparticles depend on their composition, shape, and size (Noguez, 2007) and therefore they may be tuned and optimised for a given application by nanoparticle-fabrication methods. Biosensor platforms based on the optical properties of nanoparticle arrays and even single particles have been used to monitor biological binding events without the need for a fluorescent marker (Haes et al., 2004, Heaton et al., 2001).
One evolving application of nanoparticle arrays and in particular label-free binding studies is in the field of high-throughput screening. The conventional instrumentation for high-throughput screening relies on fluorescently labelled markers and is capable of screening perhaps 104 compounds per day. However, although the fluorescent assay intensity information can be quantitatively correlated to concentrations, which is routinely realised in single-target techniques such as ELISA assays, the high-throughput screening is often limited only to “yes-no”—presence or absence of the biological molecule species, typically a protein or a gene. There is a large overhead in preparing the 104 different molecules for screening simply from the logistics of preparation of the plates and printing out the molecules and a more information-rich array screening technology is required.
Label-free surface plasmon detection techniques open a number of exciting possibilities for monitoring not just the presence or absence of a particular biomolecule but also to determine its absolute concentration. Further, an array allows the absolute concentrations of a number of molecules to be detected simultaneously to produce a pattern of molecular expression such as disease biomarkers, pointing towards a patient-specific treatment regimen. In contrast to the large body of work on nanoparticle-based platforms for single-target molecule analysis (Haes and Duyne, 2004), the techniques for simultaneous monitoring multiple targets are less well developed. Most of the progress is achieved in surface plasmon resonance imaging, where protein concentration detection limit of ca. 1 nM was reported (Lee et al., 2006, Phillips et al., 2006) To our knowledge pure light-scattering techniques were not used in microarray imaging applications.
This work presents the fabrication of multiple-target sensor arrays based on gold nanoparticles and demonstrates its application in detecting two antibodies in whole antisera by using a light-scattering sensor array reader.
Section snippets
Experimental
Arrays of gold nanoparticle surfaces are fabricated on glass slides and functionalized with target molecules. The array is then imaged in a near-field configuration and the scattered light is collected by a camera. The spatial arrangement of gold nanoparticles on the array is achieved by printing 4 nm gold seed nanoparticles from a colloidal solution using an inkjet printer into the array configuration on the surface of a glass substrate. Larger gold nanoparticles (∼100 nm) are then grown on the
Results and discussion
A measure of the sensitivity of array spots may be obtained by monitoring the change in the scattered intensity recorded in each of the R or G channels when the refractive index of the bulk medium is changed in the flow cell. The smallest detectable change in the bulk refractive index (RI) is termed here the refractive index sensitivity (RIS) of the sensor and is defined aswhere I is the scattered light intensity and σI is its standard deviation. Typically the manufactured
Conclusions
The array reader has demonstrated the potential for nanoparticle light scattering to form the basis of a small-form array reader which can interrogate a biomarker array printed on an array spot forming a biophotonic surface. Critically, there is a clear specific versus non-specific binding discrimination for the antibodies contained in the whole antisera. The short-range nature of the nanoparticle plasmon field, typically penetrating one radius into the medium above the particle, is more
Acknowledgement
We would like to thank RCUK for a Basic Technology Grant “2D Attogram Surface Plasmon Imaging”, grant number: EP/C52389X/1.
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