Provided herein are methods for classifying or characterizing a biological sample in vivo or ex vivo in real-time using time-resolved spectroscopy and/or electrical stimulation. A biological sample may produce a responsive fluorescence signal when irradiated by a light excitation signal or pulse at a predetermined wavelength. The responsive fluorescence signal may be recorded. The intensity of the excitation wavelength may be recorded and used to normalize the recorded responsive fluorescence signal. The biological sample may produce a responsive electrical signal in response to electrical stimulation. Raw fluorescence decay data may be generated from the responsive fluorescence signal and pre-processed. The pre-processed raw fluorescence decay data may be de-convolved to remove an instrument response function therefrom and generate true fluorescence decay data. The biological sample may be characterized in response to the responsive fluorescence signal, the responsive electrical signal, the normalized responsive fluorescence signal, and/or the true fluorescence decay data.

A method of estimating background radiation in spectral data. The method may comprise, iteratively, fitting an analytical curve, such as a spline curve, to reference data, determining an allowable deviation of the reference data from the analytical curve and clipping data points of the reference data or the spectral data that are more than the allowable deviation above the analytical curve to provide the reference data for the next iteration until termination criterion is met. The reference data is initially based upon the spectral data. The method may comprise generating estimates of background radiation of the spectral data, each estimate based upon fitting a different order polynomial to the spectral data, and selecting an order of polynomial to use for estimating background radiation and/or one of the estimates of the background radiation. The method may further comprise estimating the noise in the spectral data from the reference data.

Systems and methods according to exemplary embodiments of the present disclosure can be provided that can efficiently detect the amplitude and phase of a spectral modulation. Such exemplary scheme can be combined with self-interference fluorescence to facilitate a highly sensitive depth localization of self-interfering radiation generated within a sample. The exemplary system and method can facilitate a scan-free depth sensitivity within the focal depth range for microscopy, endoscopy and nanoscopy.

A spectroscopic measurement apparatus includes a light source, an integrator, a spectroscopic detector, and an analysis unit. The integrator includes an internal space in which a measurement object is disposed, a light input portion for inputting light to the internal space, a light output portion for outputting light from the internal space, a sample attachment portion for attaching the measurement object, and a filter attachment portion for attaching a filter unit. The filter unit has a transmission spectrum in which an attenuation rate for excitation light is larger than an attenuation rate for up-conversion light, and attenuates the light output from the light output portion. The analysis unit analyzes luminous efficiency of the measurement object on the basis of the transmission spectrum data and the spectroscopic spectrum data acquired by the spectroscopic detector.

An imaging device includes a light splitting unit which splits first light from a subject into second light and third light, first and second imaging units, and an arithmetic unit. The first light includes the second light having infrared light and at least one of green light and blue light, and the third light having red light or the green light. The first imaging unit includes a first and a second light reception regions. The first light reception region generates at least one of the group consisting of a B signal according to the blue light and a G signal according to the green light. The second light reception region generates an IR signal according to the infrared light. The arithmetic unit generates a visible light image signal from the R signal, the G signal, and the B signal and generates an infrared light image signal from the IR signal.

In a first aspect, a method includes: providing a sample, the sample including a first nucleotide and a second nucleotide; contacting the sample with a first fluorescent dye and a second fluorescent dye, the first fluorescent dye emitting first emitted light within a first wavelength band responsive to a first excitation illumination light, the second fluorescent dye emitting second emitted light within a second wavelength band responsive to a second excitation illumination light; simultaneously collecting, using one or more image detectors, multiplexed fluorescent light comprising the first emitted light and the second emitted light, the first emitted light being a first color channel corresponding to the first wavelength band and the second emitted light being a second color channel corresponding to the second wavelength band; and identifying the first nucleotide based on the first wavelength band of the first color channel and the second nucleotide based on the second wavelength band of the second color channel.

Provided herein are devices, systems, and methods for characterizing a biological sample in vivo or ex vivo in real-time using time-resolved spectroscopy. A light source generates a light pulse or continuous light wave and excites the biological sample, inducing a responsive fluorescent signal. A demultiplexer splits the signal into spectral bands and a time delay is applied to the spectral bands so as to capture data with a detector from multiple spectral bands from a single excitation pulse. The biological sample is characterized by analyzing the fluorescence intensity magnitude and/or decay of the spectral bands. The sample may comprise one or more exogenous or endogenous fluorophore. The device may be a two-piece probe with a detachable, disposable distal end. The systems may combine fluorescence spectroscopy with other optical spectroscopy or imaging modalities. The light pulse may be focused at a single focal point or scanned or patterned across an area.

A method to normalize at least one of a population of subordinate clinical diagnostic analyzer to a master clinical diagnostic analyzer such that an assay result from a subordinate clinical diagnostic analyzer can be converted to the equivalent result of the master clinical diagnostic analyzer by using a simple multiplicative factor when the assay executed on each analyzer uses a common fluorescently labeled dye. Also a method to re-normalize a subordinate clinical diagnostic analyzer assay result to a master clinical diagnostic analyzer assay result by using a simple multiplicative factor when the assay executed on the subordinate clinical diagnostic analyzer uses a different fluorescently labeled dye than the assay executed on the master clinical diagnostic analyzer.

First and second filter magazines in each of which plural filters having different transmission wavelengths from each other are arranged in a row are provided, and the first and second filter magazines are arranged next to each other in one direction. A light detection unit in which plural photomultipliers of first and second photomultipliers, each of which detects light that has passed through at least one of the filters included in the first and second filter magazines, are arranged in the arrangement direction of the filters is provided, and the light detection unit is placed in the one direction in such a manner to be parallel to the first and second filter magazines. The apparatus is configured in such a manner that the first and second filter magazines and the light detection unit are movable in the arrangement direction of the filters.

The invention provides systems for characterizing a biological sample by analyzing emission of fluorescent light from the biological sample upon excitation and methods for using the same. The system includes a laser source, collection fibers, a demultiplexer and an optical delay device. All references cited herein are incorporated by reference in their entirety as though fully set forth. Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of-ordinary skill in the art in which this invention belongs.