Surface plasmon resonance (SPR) enhanced biosensors
i Abstract A biosensor is a device that employs biomolecules or biomimetic components as capturing elements to detect analyte, record the information, which is translated mostly into electric signals. Different kinds of biosensors are based on different detection principles. Optical biosensors utilize the incident light as source to excite the molecules so that the signals, related to changes in refractive index, extinction/absorption, fluorescent emission and Raman scattering etc can be
... ed. A plasmon is a collective oscillation of the free electrons in a noble metal. At the surface of a metal, plasmons take the form of surface plasmon polaritons (SPPs). When a surface plasmon is confined to a particle, free electrons participate in the collective oscillation and it is termed as localized surface plasmon (LSP), while plasmons propagating along the metal surface are defined as propagating surface plasmon (PSP). Surface plasmons are commonly excited by external electromagnetic radiation and a common feature of plasmons are that they concentrate the field to the near surface region, and can thus be used to enhance extinction, fluorescent, Raman and other signals. For example, binding of molecules in the vicinity of the surface induces the refractive index changes, which causes variations in the extinction spectra, eg. shift in peak position at a particular wavelength and/or line shape. The sensitivity, specificity, selectivity, robustness, portability, speed and costs are the significant evaluation factors of a biosensor. The sensitivity and specificity are mainly related to the sensing mechanism and binding characteristics. The sensing principle in most of my works are SPR and SPR enhanced fluorescence. In the first part of my work, I designed two kinds of Au nanohole array, with (PAuNH array) and without (AuNH array) photoresist between the Au layer and the glass slides for the purpose of fluorescent enhancement using a model dye Alexa 647. This work is undertaken in close collaboration with researchers from IMRE, Singapore. The fabrication steps of the arrays were composed of electron-beam lithography and nano-imprint lithography. In comparison with the PAuNH array, Abstract ii the AuNH array offered higher field enhancement for fluorescence when only LSP was excited. The detections operated on AuNH array using the collective effects of LSP and PSP turned out to be a promising combination in that it theoretically by simulation and experimentally revealed that the fluorescent enhancement was stronger in 2 orders of magnitude as compared with flat Au thin film substrate irradiated off-resonance. In spite of intrinsic affinity of the receptor to the analyte, the external electric field provides extra force to increase the interaction frequency and probability, thus augment the signals of charged targets at increased speed. Non-specific binding is also reduced or even eliminated when the non-specific species possess different charge in relation to the analyte. I built an electro-focusing enhanced LSPR biosensor for the detection of human troponin I (cTnI). The limit of detection (LOD) in buffer can be improved by 2 orders of magnitude upon changing from no bias, 0 V, to -0.2 V bias applied on the sensor surface. The nonspecific adsorption of serum albumin (i.e. BSA) and rabbit serum is also significantly reduced at -0.2 V. The point-of care device based on grating Au film for the detection of endotoxin (lipopolysaccharide, LPS) was demonstrated, which displayed excellent portability and good sensitivity. Relying only on the built-in LED light and CCD camera alleviated the size and the light 3D-printing sample holder furnished the device with control the light path and portability. Based on monitoring the resonant wavelength shift, the results showed the feasibility for the detection of LPS at detection limit of 32.5ng/ml.