Nanoclusters and nanofilms have the potential to amplify fluorescence and thus to enhance the signal of labeled biomolecules on biochip surfaces. Fluorescent molecules are bound at a certain distance to a resonant layer of a metal or a semiconductor or both, resulting in enhanced absorption and emission of the fluorophore within the electromagnetic near-field. This property makes the system highly useful for interaction studies, including those of DNA and proteins. Due to the amount of data, derived from various sequencing projects and from Proteomic interaction studies within the next years, microarrays (or biochips) will represent a central technology in every lab facilitating high-throughput screening and being easily interfaced with computer databases. However, most chips suffer from the disadvantage of insufficient signal-to-noise (background) ratio and are thus limited to molecules of medium-to-high abundance. Novel approaches are needed for identification of, e.g., low copy RNAs or regulatory proteins. Here we present a study, using novel surface enhanced chips in the standard glass-slide-formats. Applying surface-enhanced fluorescence (SEF), the chips turned out to be useful for interaction studies, such as DNA hybridization, thereby strongly enhancing the on-chip-signals. Compared to standard glass-slide-DNA chips, both the fluorescent signals as well as signal-to-noise ratio were considerably higher.

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The most fundamental properties of metal nanoclusters, namely the high local-field enhancement and nanoscale resonance behavior of the cluster electron plasma when exited by electromagnetic radiation, have been used to set up a variety of sensors transducing biorecognitive interactions into optical signals. This paper focuses on applications of resonant-cluster technology, which enabled us to monitor biorecognitive binding of a variety of proteins on a chip, thus constructing high-throughput interaction-screening devices. Decisive for this type of sensor is the nanometric distance from the local field surrounding a cluster to other parts of the sensor interacting with this field. In particular, the cluster-mirror or cluster-fluorophore distance gives rise to a variety of enhancement phenomena. Depending on the desired application this "resonance"- distance is approximately 5-400 nm. All types of sensor can be set up on photolithographically constructed microchips, but microscopic glass slides can also be employed; this also enables the use of standard devices for dotting and read out. Using slide based chips a standard format of 3,200 microdots (125 microm in diameter) was the basis of either microassays applying direct optical transduction via surfaceenhanced absorption or striking for more sensitivity via surface-enhanced fluorescence.

The combination of phage display antibody arrays with a novel nanotransducer technique based on resonant nanoparticles in a nanosandwiched film enables the sensitive parallel screening of proteins. Using the resonance of nanoparticles with their induced mirror dipoles in a thin-film structure, limitations of fluorophores, such as unspecific background and nonvisibility to the eye, can be overcome, thereby leading to an optical signal significantly more sensitive than that of standard colloid techniques. The signal can be both directly observed as a color change of a microdot at the sensor surface and tuned throughout the visible range of the spectrum. Here we report the application of an optical chip using scFv-antibody-antigen interactions. Artificial scFv-antibodies against a variety of proteins, including yeast enzymes and bovine serum albumin (as a standard), were constructed via Phage Display. These scFv-antibodies were then coated onto metal nanoclusters and bound to their antigens that were arrayed as nanodroplets at the resonance layer of the chip. ScFv-Antibody-antigen interaction resulted in a visible array of microdots. Using resonance-enhanced absorption, the absorption signal of the spots was amplified by one to two orders of magnitude (compared to colloid-based techniques). For quantitative analysis, either an 8-micron scanner or a CCD camera (resolution 4 microns) was employed to gain direct-reflection spectra rather than unspecific scatter data (prone to dust and unspecific interaction). Our results demonstrate that this device enables high-throughput proteomics to overcome some limitations of fluorescence, enzyme labels, and colloid techniques.

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