Olympus Corporation

Oki Electric Industry Co., Ltd.

Mitsubishi Electric Corporation

Oki Semiconductor Co., Ltd.

Oki Electric Co., Ltd.

Tetsuya Tanabe

Yasuhiro Tanaka

Satoru Tanoi

Mitsushiro Yamaguchi

Takuya Hanashi

Seiji Kondo

Hidetaka Nakata

Kunio Hori

Kazutaka Nishikawa

Nobutaka Nasu

Hidetoshi Ikeda

Akira Tanabe

Kei Terada

Masanori Ozaki

Yoshihiro Marushita

Shuichi Hashidate

Masahiko Yoshida

Nobuhiko Morimoto

Yasuhiro Tokunaga

Shinichi Fukuzako

Shinya Ogiwara

Noriyoshi Sato

An optical analysis device includes a light source, a beam shaping unit, a relative movement unit, a photodetector, and a position detector. The light source unit generates a light beam. The beam shaping unit forms a flat beam portion. The relative movement unit is configured to cause the flat beam portion and a test sample including marker particles to relatively move in a minor axis direction of the flat beam portion. The photodetector is configured to detect a light intensity and a light emitting position in a plane orthogonal to the minor axis direction. The position detector is capable of detecting spatial positions of the marker particles on the basis of information on a relative movement amount of the flat beam portion, information on the light intensity and the light emitting position, and a change of the light intensity generated according to a relative movement of the flat beam portion.

Optical analysis device, optical analysis method, and recording medium

In the scanning molecule counting method of measuring light intensity from a light detection region while moving the position of the light detection region of a confocal or multiphoton microscope in a sample solution containing light-emitting particles, generating time series light intensity data and detecting each of signals of the light-emitting particles individually in the data, wherein the light-emitting particles are formed by binding to a particle to be observed a light-emitting probe which emits light through binding to the particle to be observed and in which a stochastic transition between a non-light-emitting state and a light-emitting state occurs in the unbound state, the moving speed of the position of the light detection region is adjusted to make the time during which the unbound probe is encompassed by the moving light detection region longer than an average lifetime during which the probe is in the light-emitting state.

Optical analysis method and optical analysis device using single light-emitting particle detection

There is provided an optical analysis technique of detecting light of a light-emitting particle in a sample solution in the scanning molecule counting method using the light measurement with a confocal or multiphoton microscope, for suppressing the scattering in detected results of signals of light of light-emitting particles smaller and achieving the improvement of accuracy. The inventive technique comprises moving the position of a light detection region along a predetermined route for multiple circulation times by changing the optical path of the optical system; detecting light from the light detection region and generating time series light intensity data during the moving of the light detection region and detecting individually a signal indicating light from each light-emitting particle existing in the predetermined route using the time series light intensity data obtained in the circulating movements of the light detection region of multiple times.

Optical analysis device, optical analysis method and computer program for optical analysis using single light-emitting particle detection

There is provided a microscopic observation technique capable of detecting a light-emitting object or a light-emitting particle moving in a thick sample by the scanning molecule counting method. In the inventive technique, the light from a light detection region of is detected the optical system of a confocal or multiphoton microscope is detected with while moving the light detection region in each observed subregion obtained by dividing a region to be observed into plural regions; the signal of the light from a light-emitting particle is individually detected; and the position of the light-emitting particle corresponding to the detected signal is determined in the region to be observed. The moving of the position of the light detection region in each observed subregion is performed continuously in at least two directions or and/or continuously multiple times in each observed subregion.

Optical microscope device, microscopic observation method and computer program for microscopic observation using single light-emitting particle detection technique

In the scanning molecule counting method using the light measurement with a confocal microscope or a multiphoton microscope, a measuring time is optimized with suppressing the scattering in a result small irrespective of light-emitting particle concentrations. In the inventive technique of detecting and analyzing the light from an light-emitting particle, there are repeated processes of detecting the light intensity from a light detection region with moving the position of the light detection region of an optical system in a sample solution by changing the optical path of the optical system of the microscope, and detecting the signals of the light of light-emitting particles individually, and based on the time taken for the number of the signals from the light-emitting particles to reach a predetermined number, the light-emitting particle concentration in the sample solution is determined.

Optical analysis method and optical analysis device using the detection of light from a single light-emitting particle

In the scanning molecule counting method using optical measurement with a confocal or multiphoton microscope, there is provided a technique of computing a light-emitting particle concentration which changes with time and detecting a concentration change velocity or a reaction velocity. The inventive optical analysis technique of detecting light of light-emitting particles in a sample solution generates time series light intensity data of light from a light detection region detected with moving the position of the light detection region of the microscope in the sample solution; measures successively an interval of generation times of signals of the light-emitting particles detected in the time series light intensity data; and determines the concentration or concentration change velocity of the light-emitting particles, using the successively measured signal generation time intervals.

There is provided optical analysis techniques in the scanning molecule counting method using the light measurement with a confocal or multiphoton microscope in which the measuring unit time in the light measurement is set to an appropriate value in order to surely detect an approximately bell shape profile of the signal of a light-emitting particle and avoid excessive increase data volume of time series light intensity data. The inventive optical analysis technique of detecting light of a light-emitting particle in a sample solution generates time series light intensity data of light from a light detection region detected during moving the position of the light detection region of a microscope in the sample solution and detects in the data a signal indicating light from each light-emitting particle individually. The measuring unit time is determined based on the size and the moving speed of the light detection region.

In the optical analysis technique of detecting an existence of a single particle in a sample solution with a confocal microscope or a multiphoton microscope according to the scanning molecule counting method of the present invention, the position of a light detection region is moved in the sample solution; the light intensity from the light detection region is measured so that light intensity data will be generated; a first occurrence probability in assuming a first condition that no single particles exist in the light detection region and a second occurrence probability in assuming a second condition that a single particle exists in the light detection region for a time variation of light intensity value on the light intensity data are computed; and a signal indicating each single particle is detected based on those occurrence probabilities, and thereby enabling improvements in the sensitivity and/or S/N ratio.

Optical analysis device, optical analysis method and computer program for optical analysis using single particle detection

There is provided a single particle detection technique based on a scanning molecule counting method, enabling individual detection of a single particle using light measurement with a confocal or multiphoton microscope, and quantitative observation of conditions or characteristics of the particle. The inventive technique of detecting a single particle in a sample solution detects light containing substantially constant background light from a light detection region with moving the position of the light detection region of the microscope in a sample solution to generate time series light intensity data; and detects individually a light intensity reduction occurred when a single particle which does not emit light (or a particle whose emitting light intensity in a detected wavelength band is lower than the background light) enters in the light detection region in the time series light intensity data as a signal indicating the existence of each single particle.

Single particle detection device, single particle detection method, and computer program for single particle detection, using optical analysis

There is provided a structure to make the setting of a criterion for eliminating noises easy in the scanning molecule counting method. In the inventive optical analysis technique of detecting light of a light-emitting particle in a sample solution, time series light intensity data of light from a light detection region detected with moving the position of the light detection region in the sample solution is generated, and a signal of a light-emitting particle individually is detected in the time series light intensity data, wherein a signal having a light intensity in a light intensity range set based upon a signal generation frequency integrated value distribution which is a distribution, obtained by using as a variable an intensity of a signal, of integrated values of generation frequencies of signals having an intensity not lower than the variable is extracted as the signal of the light-emitting particle.

In the scanning molecule counting method detecting light of a light-emitting particle in a sample solution using a confocal or multiphoton microscope, there is provided an optical analysis technique enabling the scanning in a sample solution with moving a light detection region in a broader area or along a longer route while making the possibility of detecting the same light-emitting particle as different particles as low as possible and remaining the size or shape of the light detection region unchanged as far as possible. In the inventive optical analysis technique, there are performed detecting light from the light detection region and generating time series light intensity data during moving the light detection region along the second route whose position is moved along the first route, and thereby, the signal indicating light from each light-emitting particle in a predetermined route is individually detected using the time series light intensity data.

There is provided an optical analysis technique enabling the detection of the condition or characteristic of a particle to be observed contained at a low concentration or number density in a sample solution using a light-emitting probe. The inventive optical analysis technique uses an optical system capable of detecting light from a micro region in a solution, such as an optical system of a confocal microscope or a multiphoton microscope, to detect the light from the light-emitting probe having bound to a particle to be observed while moving the position of the micro region in the sample solution (while scanning the inside of the sample solution with the micro region), thereby detecting individually the particle crossing the inside of the micro region to enable the counting of the particle(s) or the acquisition of the information on the concentration or number density of the particle.

Method of detecting sparse particles in a solution using a light-emitting probe

There is provided a way of enabling the discrimination or identification of the kind of a light-emitting particle corresponding to each pulse form signal in the scanning molecule counting method using the optical measurement by the confocal or multiphoton microscope. In the inventive technique, the position of a light detection region in a sample solution periodically along a predetermined route is moved in measuring the light intensity from the light detection region; and a signal of light from a light-emitting particle is detected individually. Then, an index value indicating a translational diffusional characteristic of one light-emitting particle in a plane perpendicular to the moving direction of the light detection region is determined based upon intensity values of signals of light of the same light-emitting particle for identifying a light-emitting particle.

The present invention is to provide a method for identifying a polymorphism. The method includes performing a nucleic acid chain extension reaction and identifying the polymorphism of the nucleic acid contained in the test nucleic acid sample. The extension reaction is conducted with use of a nucleic acid in a test nucleic acid sample as a template, a type I detection primer which hybridizes with a region including the polymorphic site of a nucleic acid whose polymorphic site nucleotide sequence consisting of a first nucleotide sequence, and a polymerase. The reaction is conducted with the presence of an inhibitory oligonucleotide, which hybridizes with the type I detection primer. The region of the type I detection primer to hybridize with the inhibitory oligonucleotide is located on the 5' side of the polymorphism detection site of the type I detection primer to hybridize with the polymorphic site.

Polymorphism identification method

The inventive technique of detecting and analyzing light from a light-emitting particle in accordance with the scanning molecule counting method using an optical measurement with a confocal microscope or a multiphoton microscope is characterized by detecting intensities of components of two or more wavelength bands of light from a light detection region of an optical system with moving the position of the light detection region in a sample solution by changing the optical path of the optical system of the microscope; detecting individually signals of the light from each light-emitting particle in the intensities of the components of the two or more wavelength bands of the detected light; and identifying a kind of light-emitting particle based on the intensities of the components of the two or more wavelength bands of the signals of the light of the detected light-emitting particle.

Growing community of inventors

Tokyo, Japan

Tetsuya Tanabe