2017 results about "Fluorophore" patented technology

The present invention provides labeled nucleic acid amplification oligonucleotides, which can be linear or hairpin primers or blocking oligonucleotides. The oligonucleotides of the invention are labeled with donor and / or acceptor moieties of molecular energy transfer pairs. The moieties can be fluorophores, such that fluorescent energy emitted by the donor is absorbed by the acceptor. The acceptor may be a fluorophore that fluoresces at a wavelength different from the donor moiety, or it may be a quencher. The oligonucleotides of the invention are configured so that a donor moiety and an acceptor moiety are incorporated into the amplification product. The invention also provides methods and kits for directly detecting amplification products employing the nucleic acid amplification primers. When labeled linear primers are used, treatment with exonuclease or by using specific temperature eliminates the need for separation of unincorporated primers. This "closed-tube" format greatly reduces the possibility of carryover contamination with amplification products, provides for high throughput of samples, and may be totally automated.

The present invention relates to methods and apparatuses for analyzing molecules, particularly polymers, and molecular complexes with extended or rod-like conformations. In particular, the methods and apparatuses are used to identify repetitive information in molecules or molecular ensembles, which is interpreted using an autocorrelation function in order to determine structural information about the molecules. The methods and apparatuses of the invention are used for, inter alia, determining the sequence of a nucleic acid, determining the degree of identity of two polymers, determining the spatial separation of specific sites within a polymer, determining the length of a polymer, and determining the velocity with which a molecule penetrates a biological membrane.

An optical detection and orientation device is provided comprising housing having an excitation light source, an optical element for reflecting the excitation light to an aspherical lens and transmitting light emitted by a fluorophore excited by said excitation light, a focussing lens for focusing the emitted light onto the entry of an optical fiber, which serves as a confocal aperture, and means for accurately moving said housing over a small area in relation to a channel in a microfluidic device. The optical detection and orientation device finds use in identifying the center of the channel and detecting fluorophores in the channel during operations involving fluorescent signals.

A method and device is disclosed for high speed, automated sequencing of nucleic acid molecules. A nucleic acid molecule to be sequenced is exposed to a polymerase in the presence of nucleotides which are to be incorporated into a complementary nucleic acid strand. The polymerase carries a donor fluorophore, and each type of nucleotide (e.g. A, T / U, C and G) carries a distinguishable acceptor fluorophore characteristic of the particular type of nucleotide. As the polymerase incorporates individual nucleic acid molecules into a complementary strand, a laser continuously irradiates the donor fluorophore, at a wavelength that causes it to emit an emission signal (but the laser wavelength does not stimulate the acceptor fluorophore). In particular embodiments, no laser is needed if the donor fluorophore is a luminescent molecule or is stimulated by one. The emission signal from the polymerase is capable of stimulating any of the donor fluorophores (but not acceptor fluorophores), so that as a nucleotide is added by the polymerase, the acceptor fluorophore emits a signal associated with the type of nucleotide added to the complementary strand. The series of emission signals from the acceptor fluorophores is detected, and correlated with a sequence of nucleotides that correspond to the sequence of emission signals.

A spectral imaging method for simultaneous detection of multiple fluorophores aimed at detecting and analyzing fluorescent in situ hybridizations employing numerous chromosome paints and / or loci specific probes each labeled with a different fluorophore or a combination of fluorophores for color karyotyping, and at multicolor chromosome banding, wherein each chromosome acquires a specifying banding pattern, which pattern is established using groups of chromosome fragments labeled with various fluorophore or combinations of fluorophores.

A method for fluorophore bias removal in microarray experiments in which the fluorophores used in microarray experiment pairs are reversed. Further, a method for calculating the individual errors associated with each measurement made in nominally repeated microarray experiments. This error measurement is optionally coupled with rank based methods in order to determine a probability that a cellular constituent is up or down regulated in response to a perturbation. Finally, a method for determining the confidence in the weighted average of the expression level of a cellular constituent in nominally repeated microarray experiments.

The present invention provides labeled nucleic acid amplification oligonucleotides, which can be linear or hairpin primers or blocking oligonucleotides. The oligonucleotides of the invention are labeled with donor and / or acceptor moieties of molecular energy transfer pairs. The moieties can be fluorophores, such that fluorescent energy emitted by the donor is absorbed by the acceptor. The acceptor may be a fluorophore that fluoresces at a wavelength different from the donor moiety, or it may be a quencher. The oligonucleotides of the invention are configured so that a donor moiety and an acceptor moiety are incorporated into the amplification product. The invention also provides methods and kits for directly detecting amplification products employing the nucleic acid amplification primers. When labeled linear primers are used, treatment with exonuclease or by using specific temperature eliminates the need for separation of unincorporated primers. This "closed-tube" format greatly reduces the possibility of carryover contamination with amplification products, provides for high throughput of samples, and may be totally automated.

Fluorescent biosensor molecules, fluorescent biosensors and systems, as well as methods of making and using these biosensor molecules and systems are described. These biosensor molecules address the problem of obtaining fluorescence emission at wavelengths greater than about 500 nm. Biosensor molecules generally include an (1) an acridine-based fluorophore, (2) a linker moiety and (3) a boronate substrate recognition / binding moiety, which binds polyhydroxylate analytes, such as glucose. These biosensor molecules further include a “switch” element that is drawn from the electronic interactions among these submolecular components. This fluorescent switch is generally “off” in the absence of bound polyhydroxylate analyte and is generally “on” in the presence of bound polyhydroxylate analyte. Thus, the reversible binding of a polyhydroxylate analyte essentially turns the fluorescent switch “on” and “off”. This property of the biosensor molecules, as well as their ability to emit fluorescent light at greater than about 500 nm, renders these biosensor molecules particularly well-suited for detecting and measuring in-vivo glucose concentrations.

An optical detection and orientation device is provided comprising housing having an excitation light source. an optical element for reflecting the excitation light to an aspherical lens and transmitting light emitted by a fluorophore excited by said excitation light, a focussing lens for focusing the emitted light onto the entry of an optical fiber, which serves as a confocal aperture, and means for accurately moving said housing over a small area in relation to a channel in a microfluidic device. The optical detection and orientation device finds use in identifying the center of the channel and detecting fluorophores in the channel during operations involving fluorescent signals.

The present invention provides plasmonic crystals comprising three-dimensional and quasi comprising three-dimensional distributions of metallic or semiconducting films, including multi-layered crystal structures comprising nanostructured films and film arrays. Plasmonic crystals of the present invention include precisely registered and deterministically selected nonplanar crystal geometries and spatial distributions providing highly coupled, localized plasmonic responses in thin film elements and / or nanostructures of the crystal. Coupling of plasmonic responses provided by three-dimensional and quasi-three dimensional plasmonic crystal geometries and structures of the present invention generates enhanced local plasmonic field distributions useful for detecting small changes in the composition of an external dielectric environment proximate to a sensing surface of the plasmonic crystal. Plasmonic crystal structures of the present invention are also useful for providing highly localized excitation and / or imaging of fluorophores proximate to the crystal surface.

An optical detection and orientation device is provided comprising housing having an excitation light source, an optical element for reflecting the excitation light to an aspherical lens and transmitting light emitted by a fluorophore excited by said excitation light, a focussing lens for focusing the emitted light onto the entry of an optical fiber, which serves as a confocal aperture, and means for accurately moving said housing over a small area in relation to a channel in a microfluidic device. The optical detection and orientation device finds use in identifying the center of the channel and detecting fluorophores in the channel during operations involving fluorescent signals.

A method and device is disclosed for high speed, automated sequencing of nucleic acid molecules. A nucleic acid molecule to be sequenced is exposed to a polymerase in the presence of nucleotides which are to be incorporated into a complementary nucleic acid strand. The polymerase carries a donor fluorophore, and each type of nucleotide (e.g. A, T / U, C and G) carries a distinguishable acceptor fluorophore characteristic of the particular type of nucleotide. As the polymerase incorporates individual nucleic acid molecules into a complementary strand, a laser continuously irradiates the donor fluorophore, at a wavelength that causes it to emit an emission signal (but the laser wavelength does not stimulate the acceptor fluorophore). In particular embodiments, no laser is needed if the donor fluorophore is a luminescent molecule or is stimulated by one. The emission signal from the polymerase is capable of stimulating any of the donor fluorophores (but not acceptor fluorophores), so that as a nucleotide is added by the polymerase, the acceptor fluorophore emits a signal associated with the type of nucleotide added to the complementary strand. The series of emission signals from the acceptor fluorophores is detected, and correlated with a sequence of nucleotides that correspond to the sequence of emission signals.

A laser scanning microscope produces molecular excitation in a target material by simultaneous absorption of three or more photons to thereby provide intrinsic three-dimensional resolution. Fluorophores having single photon absorption in the short (ultraviolet or visible) wavelength range are excited by a beam of strongly focused subpicosecond pulses of laser light of relatively long (red or infrared) wavelength range. The fluorophores absorb at about one third, one fourth or even smaller fraction of the laser wavelength to produce fluorescent images of living cells and other microscopic objects. The fluorescent emission from the fluorophores increases cubicly, quarticly or even higher power law with the excitation intensity so that by focusing the laser light, fluorescence as well as photobleaching are confined to the vicinity of the focal plane. This feature provides depth of field resolution comparable to that produced by confocal laser scanning microscopes, and in addition reduces photobleaching and phototoxicity. Scanning of the laser beam by a laser scanning microscope, allows construction of images by collecting multi-photon excited fluorescence from each point in the scanned object while still satisfying the requirement for very high excitation intensity obtained by focusing the laser beam and by pulse time compressing the beam. The focused pulses also provide three-dimensional spatially resolved photochemistry which is particularly useful in photolytic release of caged effector molecules, marking a recording medium or in laser ablation or microsurgery. This invention refers explicitly to extensions of two-photon excitation where more than two photons are absorbed per excitation in this nonlinear microscopy.

The present invention provides clostridial toxin substrates useful in assaying for the protease activity of any clostridial toxin, including botulinum toxins of all serotypes as well as tetanus toxins. A clostridial toxin substrate of the invention contains a donor fluorophore; an acceptor having an absorbance spectrum overlapping the emission spectrum of the donor fluorophore; and a clostridial toxin recognition sequence that includes a cleavage site, where the cleavage site intervenes between the donor fluorophore and the acceptor and where, under the appropriate conditions, resonance energy transfer is exhibited between the donor fluorophore and the acceptor.

In a minimally invasive surgical system, an illuminator includes a visible color component illumination source and a hardware non-visible fluorescence emission illumination source. Thus, the illuminator outputs target image illumination light in a first spectrum where the first spectrum includes at least a portion of the visible spectrum. The illuminator also outputs target image illumination light in a second spectrum, where the second spectrum includes non-visible light with a wavelength the same as a wavelength in an emission from a fluorophore.

The present invention provides plasmonic crystals comprising three-dimensional and quasi comprising three-dimensional distributions of metallic or semiconducting films, including multi-layered crystal structures comprising nanostructured films and film arrays. Plasmonic crystals of the present invention include precisely registered and deterministically selected nonplanar crystal geometries and spatial distributions providing highly coupled, localized plasmonic responses in thin film elements and / or nanostructures of the crystal. Coupling of plasmonic responses provided by three-dimensional and quasi-three dimensional plasmonic crystal geometries and structures of the present invention generates enhanced local plasmonic field distributions useful for detecting small changes in the composition of an external dielectric environment proximate to a sensing surface of the plasmonic crystal. Plasmonic crystal structures of the present invention are also useful for providing highly localized excitation and / or imaging of fluorophores proximate to the crystal surface.

Provided is a durable tissue pH / pCO2 and / or tissue oxygen sensitive probe of sufficient strength to withstand direct tissue pressures in vivo, the probe comprising one or more sensor chambers within a biocompatible, gas-permeable membrane containing together in a single chamber, or in separate chambers, respectively, a pH sensitive fluorophor from which pCO2 level(s) are calculated when the fluorophor is excited and the resulting fluorescence is measured and / or an oxygen sensitive phosphor solution producing oxygen quenchable phosphorescence when excited. Further provided is a tissue pH / pCO2 and / or tissue oxygen detection and measurement system comprising the probe, and methods for use of the probe and the system to directly, rapidly and accurately measure tissue pH / pCO2 and / or tissue oxygen levels in a patient without reliance on blood vessels or fluid protection of the probe.

A color display comprising an image of all chromosomes or portions of chromosomes of a cell, each of the chromosomes or portions of chromosomes being painted with a different fluorophore or a combination of fluorophores, the image presenting the chromosomes or portions of chromosomes in different distinctive colors, wherein each of the chromosomes or portions of chromosomes is associated with one of the different distinctive colors.

Disclosed is a compact, microprocessor-controlled instrument for fluorometric assays in liquid samples, the instrument having a floating stage with docking bay for receiving a microfluidic cartridge and a scanning detector head with on-board embedded microprocessor operated under control of a ODAP daemon resident in the detector head for controlling source LEDs, emission signal amplification and filtering in an isolated, low noise, high-gain environment within the detector head. Multiple optical channels may be incorporated in the scanning head. In a preferred configuration, the assay is validated using dual channel optics for monitoring a first fluorophore associated with a target analyte and a second fluorophore associated with a control. Applications include molecular biological assays based on PCR amplification of target nucleic acids and fluorometric assays in general, many of which require temperature control during detection.

Fluorescent biosensor molecules, fluorescent biosensors and systems, as well as methods of making and using these biosensor molecules and systems are described. Embodiments of these biosensor molecules exhibit fluorescence emission at wavelengths greater than about 650 nm. Typical biosensor molecules include a fluorophore that includes an iminium ion, a linker moiety that includes a group that is an anilinic type of relationship to the fluorophore and a boronate substrate recognition / binding moiety, which binds glucose. The fluorescence molecules modulated by the presence or absence of polyhydroxylated analytes such as glucose. This property of these molecules of the invention, as well as their ability to emit fluorescent light at greater than about 650 nm, renders these biosensor molecules particularly well-suited for detecting and measuring in-vivo glucose concentrations.

The present invention relates to methods and apparatuses for analyzing molecules, particularly polymers, and molecular complexes with extended or rod-like conformations. In particular, the methods and apparatuses are used to identify repetitive information in molecules or molecular ensembles, which is interpreted using an autocorrelation function in order to determine structural information about the molecules. The methods and apparatuses of the invention are used for, inter alia, determining the sequence of a nucleic acid, determining the degree of identity of two polymers, determining the spatial separation of specific sites within a polymer, determining the length of a polymer, and determining the velocity with which a molecule penetrates a biological membrane.

An active CMOS biosensor chip for fluorescent-based detection is provided that enables time-gated, time-resolved fluorescence spectroscopy. In one embodiment, analytes are loaded with fluorophores that are bound to probe molecules immobilized on the surface of the chip. Photodiodes and other circuitry in the chip are used to measure the fluorescent intensity of the fluorophore at different times. These measurements are then averaged to generate a representation of the transient fluorescent decay response unique to the fluorophores. In addition to its low-cost, compact form, the biosensor chip provides capabilities beyond those of macroscopic instrumentation by enabling time-gated operation for background rejection, easing requirements on optical filters, and by characterizing fluorescence lifetime, allowing for a more detailed characterization of fluorophore labels and their environment. The biosensor chip can be used for a variety of applications including biological, medical, in-the-field applications, and fluorescent lifetime imaging applications.

A molecular construct capable of fluorescent resonance energy transfer (FRET), comprising a linker peptide, a donor fluorophore moiety and an acceptor fluorophore moiety, wherein the linker peptide is a substrate of a botulinum neurotoxin selected from the group consisting of synaptobrevin, syntaxin and SNAP-25, or a fragment thereof capable being cleaved by the botulinum neurotoxin, and separates the donor and acceptor fluorophores by a distance of not more than 10 nm, and wherein emission spectrum of the donor fluorophore moiety overlaps with the excitation spectrum of the acceptor fluorophore moiety; or wherein the emission spectra of the fluorophores are detectably different. Also provided are isolated nucleic acid expressing the construct, kits comprising said construct and cell lines comprising said nucleic acid. Further provided are methods of detecting a BoNT using the above described construct via FRET, and methods for detecting a BoNT using surface plasmon resonance imaging.

The present invention provides a technology called Pulse-Multiline Excitation or PME. This technology provides a novel approach to fluorescence detection with application for high-throughput identification of informative SNPs, which could lead to more accurate diagnosis of inherited disease, better prognosis of risk susceptibilities, or identification of sporadic mutations. The PME technology has two main advantages that significantly increase fluorescence sensitivity: (1) optimal excitation of all fluorophores in the genomic assay and (2) “color-blind” detection, which collects considerably more light than standard wavelength resolved detection. Successful implementation of the PME technology will have broad application for routine usage in clinical diagnostics, forensics, and general sequencing methodologies and will have the capability, flexibility, and portability of targeted sequence variation assays for a large majority of the population.

A method to detect the presence or amount of at least one molecule in a sample which employs a derivative of luciferin or a derivative of a fluorophore is provided.

Methods are provided for cancer and pre-cancer detection by increased uptake of fluorophore glucose or deoxyglucose conjugates in cancerous and pre-cancerous cells relative to normal cells.

Improved methods, systems and apparatus relating to wide field fluorescence and reflectance imaging are provided, including improved methods, systems and apparatus relating to removal of background signals such as autofluorescence and / or fluorophore emission cross-talk; distance compensation of fluorescent signals; and co-registration of multiple signals emitted from three dimensional tissues.