The emergence of microarray-based technologies has revolutionized the analysis of gene expression and DNA sequence. In advanced mircroarray applications, nucleic acids are labelled with fluorescent dyes and hybridized to cDNAs or oligonucleotides configured on a solid surface. Fluorescent signals are detected and interpreted using sophisticated instrumentation that relies on scanning confocal fluorescence microscopy and fluorescent image analysis software. In practice, the performance and accessibility of many microarray systems is encumbered by limited sensitivity of the fluorescent label(s), poor dynamic range, fluorescence quenching, photobleaching and expensive instrumentation. To address these microarray system performance issues, we have applied resonance light-scattering (RLS) particle technology, a novel, ultra-sensitive and relatively simple signal generation and detection technology, to microarray systems. Submicroscopic RLS gold and silver particles of uniform dimension (20–120 nm diameter range) scatter incident white light to generate coloured light that appears as highly intense fluorescence. The intensity of coloured light signal generated by a single RLS particle is 104–106 times greater than the signal from a single molecule of the most sensitive fluorophore. The intense RLS signal does not photobleach and the colour of scattered light generated by RLS particles depends on particle composition, size, shape and the refractive index of the scattering medium. The intensity and colour of individual RLS particles can be accurately predicted to develop multiplex detection systems by practical application of mathematical calculations derived from electromagnetic radiation theory1. The high level of signal intensity allows detection at extremely low particle concentration by the unaided eye (10−15–10−16 M) and single RLS particles can be detected under appropriate illumination conditions using low-power magnification2. On solid surfaces, the RLS signal can be detected by the unaided eye at particle densities as low as 0.01–0.05 particles per square micron. RLS particles can be derivatized with a variety of molecules for analytical bioassays including proteins, antibodies, small molecule ligands, nucleic acids and oligonucleotide probes. Once derivatized, RLS particles are highly selective for detection of specific biomolecular targets. Simple detection instrumentation consisting of an illumination source, configured light microscope, video camera and image analysis system has been developed and successfully applied to microarrays. The optical properties and practical application of RLS particles for microarray-based gene expression and DNA sequence analysis will be discussed.
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