Multicolor microwave-accelerated metal-enhanced fluorescence (M-MAMEF)

摘要

The present invention relates to the use of multiple different light emitting molecules that emit different and detectable emission signals to provide systems and methods to detect different target products in a single assay sample, wherein the different light emitting molecules are positioned an optimal distance from metallic particles thereby enhancing emissions. Preferably, the systems and methods further comprise use of either microwave or sonic energy to increase binding reactions, timing of such reactions within the assay sample and reduce background non-specific biological absorption.

主权项

1. A method of decreasing the detection time of a metal-enhanced fluorescence assay used for detecting a single nucleotide sequence or multiple and different target nucleotides, the method comprising: applying a multiplicity of metallic particles to a surface of a quartz substrate used in an assay system, wherein the metallic particles are silver or a combination of silver and gold or a combination of silver and aluminum and wherein the metallic particles are metallic islands, colloids, or nanostructures;
connecting a capture nucleotide to the metallic particles selected from two to seven different capture nucleotides, wherein each of the two to seven different capture nucleotides has binding affinity for a different target nucleotide; introducing a solution suspected of including the two to seven different target nucleotides, wherein any of the two to seven different target nucleotides present in the sample bind to the corresponding capture nucleotide; introducing from two to seven different detector nucleotides, wherein each of the detector nucleotides has binding affinity for a different target nucleotide and wherein each of the two to seven detector nucleotides includes a different fluorescence molecule, wherein the different fluorescence molecule is positioned from about 6 nm to about 30 nm from the metallic particles when the detector nucleotide binds to the corresponding target nucleotide that is bound to the capture nucleotide, and wherein the metallic particles are positioned about 6 to 9 nm from each other to avoid inter-molecular energy transfer between fluorescent molecules upon excitation; applying microwave energy to the assay system for a time period sufficient to increase binding reactions between the two to seven capture nucleotides and/or detector nucleotides with the two to seven different target nucleotides; applying electromagnetic energy at a different frequency type than the microwave energy to excite the corresponding and different fluorescence molecules, using either one photon or multiphoton excitation; and detecting different fluorescence signals from the different fluorescence molecules by either visual discrimination of emissions having emission wavelengths of different colors or using a variety of filter and diffraction grating to detect the desired emission wavelengths for independent emission detection.