IMPSENS · Improving the detection sensitivity for protein micro-arrays: wave-guide SPR imaging and nano-particle enhanced imaging
FP6 — Marie Curie Actions (Human Resources and Mobility)
- Duration
- 2006-06-01 → 2008-05-31
- EU contribution
- €229,327
- Participants
- 1
- Scheme
- IIF
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Results in brief
Final Activity Report Summary - IMPSENS (Improving the detection sensitivity for protein microarrays: waveguide SPR imaging and nanoparticle enhanced imaging)
Surface plasmon resonance microscopy (SPRM) is an emerging technique for the observation of surface phenomena on the molecular scale with unrivalled sensitivity. A surface plasmon is a collective oscillation of surface bound electrons in a nobel metal, such as gold. Surface plasmons are excited by photons, under conservation of momentum and energy. Technically, p-polarised light is passed through a coupling arrangement onto a semitransparent thin gold layer and the reflected light, i.e. the light that is not coupled into surface plasmons is analysed with a digital camera. The coupling conditions, such as angle of incidence and wavelength of the light can be used to describe the medium adherent to the metal film. In contrast to fluorescence microscopy SPRM does not require labelling with flurophores, the observed contrast is based on the refractive index only. This is a tremendous advantage, since it allows the observations of artificial lipid membranes, proteins and DNA in the native, unaltered state. A particular application is the observation of protein-protein interactions to determine, for example the affinity between antibodies and antigens. One partner in the complex is immobilised on the metal surface, by covalent linkage and the ligand is flushed across the surface. Upon binding between the partners, the total surface bound mass is increased and this signal can easily be measured in real time. This allows it to describe the interaction strength and dynamic between a virtually limitless numbers of biomolecules and provides a platform for studying complex biological systems in the laboratory. Utilising wide field microscopy it is possible to deposit hundredths of different proteins or DNA samples in microspots and test complex mixtures of analytes. This allows high throughput screening for the potential diagnosis of diseases and genetic defects.
Data: CORDIS, © European Union
Project objective
Surface plasmon resonance (SPR) spectroscopy has proven its utility and versatility in many studies, with a focal point in biochemical investigations. SPR is regarded as the most sensitive method in label-free analysis. Surface plasmon resonance imaging (S PRi) improves on the single spot interrogation of a sample by imaging an area of the sample, which can consist of a diverse sample matrix. Instead of monitoring the angular shift of the reflection minimum, a fixed angle measurement near the SPR angle is performed.A very high detection sensitivity comparable to that of fluorescence spectroscopy is often demanded in various measurements to probe low concentration binding/interactions or weak affinities. Research efforts will focus on improvement of the detect ion sensitivity by developing wave-guide SPR imaging (WSPRi) and using nanoparticles for signal enhancement. The use of WSPRi allows for the excitation of plasmon resonance with both p and s polarized light. The resulting reflectivity spectra posses a much reduced half-width which translates to a higher sensitivity in imaging mode. Nanoparticle functionalised antibodies in combination with silver staining will be applied and the protocol optimised, aiming at a signal improvement by two to three orders of magnitude compared to existing SPR imaging assays.
Original text from CORDIS.
Participants
- QUEEN MARY & WESTFIELD COLLEGE, UNIVERSITY OF LONDON · LONDONCoordinatorCity levelUnited Kingdom
Links
Data: CORDIS, © European Union