CN111504978B - Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device - Google Patents

Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device Download PDF

Info

Publication number
CN111504978B
CN111504978B CN202010401969.5A CN202010401969A CN111504978B CN 111504978 B CN111504978 B CN 111504978B CN 202010401969 A CN202010401969 A CN 202010401969A CN 111504978 B CN111504978 B CN 111504978B
Authority
CN
China
Prior art keywords
module
spectrum
time
photons
sample
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN202010401969.5A
Other languages
Chinese (zh)
Other versions
CN111504978A (en
Inventor
王平
李昊政
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Wuhan Huayan Photoelectric Technology Co ltd
Huazhong University of Science and Technology
Original Assignee
Wuhan Huayan Photoelectric Technology Co ltd
Huazhong University of Science and Technology
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Wuhan Huayan Photoelectric Technology Co ltd, Huazhong University of Science and Technology filed Critical Wuhan Huayan Photoelectric Technology Co ltd
Priority to CN202010401969.5A priority Critical patent/CN111504978B/en
Publication of CN111504978A publication Critical patent/CN111504978A/en
Application granted granted Critical
Publication of CN111504978B publication Critical patent/CN111504978B/en
Priority to PCT/CN2021/093540 priority patent/WO2021228187A1/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G01N21/658—Raman scattering enhancement Raman, e.g. surface plasmons
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/01—Arrangements or apparatus for facilitating the optical investigation
    • G01N2021/0106—General arrangement of respective parts
    • G01N2021/0112—Apparatus in one mechanical, optical or electronic block
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G01N2021/653—Coherent methods [CARS]

Abstract

The invention belongs to the technical field of spectral measurement and spectral imaging, and discloses a pulse type time-delay dispersion spectral measurement method and device and a spectral imaging method and device; the spectral measurement method comprises the following steps: s1: exciting a sample to be detected by pulse laser and generating a transient spectrum; s2: different photons in the transient spectrum are subjected to time delay processing and are sequentially separated according to a time sequence; s3: photons in the spectrum over time are detected using a high-speed, highly sensitive photodetector. The invention adopts the instant pulse to excite the sample, concentrates the spectrum excited in the sample to the time period between the excitation light pulses, and then utilizes the single-channel high-sensitivity photoelectric detector to realize high-efficiency direct measurement on the spectrum spread in time without depending on multi-channel detection equipment to measure the spectrum; not only can efficiently utilize all photons in the spectrum, but also can improve the sensitivity of spectral measurement from the order of magnitude, thereby improving the spectral measurement efficiency.

Description

Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device
Technical Field
The invention belongs to the technical field of spectral measurement and spectral imaging, and particularly relates to a pulse type time delay dispersion spectral measurement method and device and a spectral imaging method and device.
Background
Spectroscopy has found widespread use in many fields such as biology, chemistry, materials, medicine, and in industrial production because of its molecular specificity and other analytical and measurement advantages. The fluorescence spectrum is derived from electronic state transition in atomic molecules, and can be applied to imaging, identification of atomic molecule components and states, and fluorescence labeling. However, the line width of the fluorescence spectrum line is wide and mostly ranges from 20nm to 100nm, so that the fluorescence imaging technology is difficult to realize more than five kinds of multicolor imaging. In contrast, the raman spectrum is derived from the vibrational state transition of molecules, and the raman fingerprint spectrum has good molecular specificity, can identify and characterize chemical components in a complex system, and can quantify the concentration of the molecules. And the Raman spectrum has narrower characteristic peak line width which is only 0.1 nm-1 nm, and the multicolor Raman imaging can be realized. By using the special Raman label, hundreds of colors or more of Raman labels can be developed, and more than one of hundreds of proteins and Ribonucleic acid (RNA) in organisms can be labeled, identified, positioned and imaged. However, the signal of the traditional raman is very weak, and the molecular concentration in the organism is low, so that the common raman is difficult to have wider application in biological imaging.
The detector of the common raman spectrometer usually adopts a CCD camera for spectrum acquisition. The CCD camera has low sensitivity, can suppress electronic noise only by a complex cooling system, has low data transmission speed and cannot realize high-speed signal acquisition. The sensitivity of a Photomultiplier (PMT) is far higher than that of a CCD (charge coupled device) pixel, but the use of a high-sensitivity Photomultiplier array to replace the CCD pixel requires 100-1000 independent PMT probes, and the system is expensive and complex and cannot be realized. Therefore, the Raman sensitivity is greatly limited only by the mode of performing Raman spectrum recording by the CCD spectrometer, and the development of Raman is limited for a long time.
Coherent Raman Scattering includes Coherent Anti-Stokes Scattering (CARS) and Stimulated Raman Scattering (SRS), and the Raman signal intensity of the Coherent Raman Scattering is 3 to 7 orders of magnitude higher than that of a common Raman signal. CARS and SRS are therefore widely used in the field of label-free bio-imaging. The CARS realizes the detection of Raman signals through a photomultiplier tube (PMT) and the SRS through a phase-locked amplification mode, and both have the advantages that high-speed and high-sensitivity Raman molecular imaging can be realized, but the disadvantage that the imaging can be only carried out on a single Raman peak. However, for the acquisition and imaging of raman spectra for detecting more characteristic peaks, SRS needs to be realized by scanning pulse laser wavelength and the like, and has slow speed and limited spectral range. Similarly, the acquisition of the CARS spectrum is not different from the traditional CCD spectrometer, and the sensitivity is low and the acquisition speed is slow. Since detectors with high quantum efficiency or gain amplification have not been available in spectroscopic detection, the detection of raman photons is very inefficient, meaning a significant waste of photon counts. Even using a single channel PMT or photon counter, time-sharing detection of photons of multiple wavelengths spatially separated by a grating also results in wasted photons.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide a method and a device for measuring a pulse type time-delay dispersion spectrum, and aims to solve the problem of low spectrum measurement efficiency caused by insufficient detection sensitivity of a spectrometer in the prior spectrum measurement technology.
The invention provides a pulse type time delay dispersion spectral measurement method, which comprises the following steps:
s1: exciting a sample to be detected by pulse laser and generating a transient spectrum;
s2: different photons in the transient spectrum are subjected to time delay processing and are sequentially separated according to a time sequence;
s3: photons in the spectrum over time are detected using a high-speed, highly sensitive photodetector.
In step S2, the different photons in the temporal spectrum are temporally delayed and dispersed by wavelength, frequency, phase, energy, polarization, wave vector direction or intensity.
In step S3, the high-speed and high-sensitivity photodetector employs a single-channel detection element, which includes a photomultiplier tube, a silicon-based photomultiplier tube, a photon counter, a single photon avalanche diode, or a modulation-demodulation photo-amplification detector.
The invention also provides a pulse type time delay dispersion spectrum measuring device, which comprises a pulse laser generating module, a time delay dispersion module and a photoelectric detection module, wherein the pulse laser generating module is used for generating pulse laser and irradiating the pulse laser to a sample to be measured to excite the sample to generate transient spectrum; the delay dispersion module is used for carrying out time delay processing on different photons in the instantaneous spectrum and sequentially separating the photons according to a time sequence; the photoelectric detection module is used for detecting photons coming in the spectrum along with time.
The invention also aims to provide an imaging method based on pulse type delay dispersion spectral measurement, aiming at solving the technical problem of slow spectral imaging speed caused by low spectral measurement efficiency in the prior art.
The invention provides a spectral imaging method based on the spectral measurement method.
The invention provides a pulse type time delay dispersion spectrum imaging device, comprising: the device comprises a pulse laser generating module, a reflecting module, a laser scanning module, an objective lens focusing module, a sample stage, a photon collecting module, a time delay dispersion module, a photoelectric detection module and an acquisition module; the pulse laser generating module is used for generating pulse laser; the reflection module is used for reflecting the pulse laser to the laser scanning module; the laser scanning module is used for scanning laser in space and realizing imaging; the objective lens focusing module is used for focusing laser on the sample; the sample stage is used for placing a sample and moving the sample to finish imaging; the photon collection module is used for collecting and outputting the instantaneous spectrum generated by the sample; the delay dispersion module is used for carrying out time delay processing on different photons in the instantaneous spectrum and sequentially separating the photons according to a time sequence; the photoelectric detection module is used for detecting photons coming from the spectrum along with time; the acquisition module is used for acquiring the detected photons.
The invention also provides a pulse type time delay dispersion spectral imaging device, comprising: the system comprises a double-channel pulse laser generating module, a first power regulating module, a second power regulating module, a light modulation module, a time delayer, a second reflection module, a dichroic mirror module, a laser scanning module, an objective lens focusing module, a sample stage, a photon collecting module, a time delay dispersion module, a photoelectric detection module and a collecting module; the dual-channel pulse laser generation module is used for generating two paths of pulse lasers of pump light and Stokes light; the first power adjusting module is used for adjusting the power of the Stokes light; the second power adjusting module is used for adjusting the power of the pump light; the optical modulation module is used for modulating the power or the phase or the polarization or the time delay of the Stokes light; the time delayer is used for registering the pulses of the pumping light and the Stokes light in time; the first reflection module is used for reflecting the pump light to the dichroic mirror module; the dichroscope module is used for reflecting the Stokes light and enabling the pumping light and the Stokes light to coincide in space; the laser scanning module is used for scanning laser in space and realizing imaging; the objective lens focusing module is used for focusing laser on the sample; the sample stage is used for placing a sample and moving the sample to finish imaging; the photon collection module is used for collecting and outputting the instantaneous spectrum generated by the sample; the delay dispersion module is used for carrying out time delay processing on different photons in the instantaneous spectrum and sequentially separating the photons according to a time sequence; the photoelectric detection module is used for detecting photons coming from the spectrum along with time; the acquisition module is used for acquiring the detected photons.
Furthermore, the photon collection module is a forward detection photon collection module or a backward detection photon collection module; the forward detection photon collection module is used for collecting and outputting an instantaneous spectrum generated by the sample in the direction of penetrating the sample; and the backward detection photon collection module is used for collecting and outputting the transient spectrum generated by the sample in the opposite direction of the exciting light.
As an embodiment of the present invention, the time-delay dispersion module includes a dispersion fiber, one end of which is connected to the photon collection module, and the other end of which is connected to the photoelectric detection module; when photons with different wavelengths are emitted out of the dispersion optical fiber outlet in sequence along with time, the photoelectric detection module performs photoelectric detection on the photons and generates a spectrum.
As another embodiment of the present invention, a time delay dispersion module includes: the device comprises a light splitting element, a conjugate module, a beam splitting module and a time delay reflection module; the light splitting element is used for splitting the photons collected and collimated by the photon collecting module, so that the photons with different wavelengths in the spectrum are diffracted to different angles in sequence and then emitted; the conjugate module is used for guiding the emergent light into the reflection module; the time delay reflection module is used for reflecting the guided light for multiple times and then returning the light to the original path, so that the path difference of the photons with different wavelengths in the spectrum back and forth in the reflection is different, and the photons with different wavelengths generate different time delays; the beam splitting module is used for transferring the returned delayed photons to the photoelectric detection module.
As another embodiment of the present invention, a time delay dispersion module includes: an optical wavelength divider, an optical fiber waveguide and an optical multiplexer; the optical wave splitter is used for distributing the photons collected and collimated by the photon collection module to optical fiber waveguides with different lengths; the optical fiber waveguides with different lengths enable the propagation distances of photons with different wavelengths to be different, and the photons with different wavelengths generate different time delays; the optical multiplexer is used for combining the multiple paths of photons after the time delay and outputting the combined photons to the photoelectric detection module.
In the spectral imaging method provided by the invention, along with the great improvement of the detection efficiency and the sensitivity of spectral measurement, the integration time of spectrum acquisition is reduced, and the spectral imaging speed is obviously improved. For example, in the case of raman spectroscopy, the acquisition of conventional raman spectroscopy requires 0.1 second, and in the case of raman spectroscopy imaging with 1000 by 1000 pixels, 1 million raman spectra are acquired, which requires 27.8 hours and the imaging speed is too slow. The new method can improve the spectrum collecting speed to 1 microsecond, so that a Raman spectrogram with million pixels can be completed in 1 second.
In summary, compared with the prior art, the invention has the following technical advantages:
(1) the invention concentrates the excitation laser to the extremely short transient pulse, adopts the transient pulse to excite the sample, concentrates the spectrum excited in the sample to the time period between the excitation light pulses, and realizes the high-efficiency direct measurement of the spectrum spread in time by using the single-channel high-sensitivity photoelectric detector without depending on the multi-channel detection equipment to measure the spectrum. The invention can adopt a more sensitive single-channel photoelectric detector, not only can efficiently utilize all photons in the spectrum, but also can improve the sensitivity of the spectrum measurement from the order of magnitude, and simultaneously, the invention also avoids the problem that the spectrum is measured by a multi-pixel, multi-channel, low-sensitivity and low-efficiency detector such as a CCD in the traditional spectrometer. Meanwhile, the excitation of the pulse laser and the detection time after the pulse are a period, the acquired spectrum in each period is completely the same, photons in the spectrum can be used for integration of repeated signals, the excitation light and the generated spectrum photons are completely used for spectrum measurement, and no waste is generated; thereby improving photon utilization rate and improving spectral measurement efficiency.
(2) The spectrum measuring device provided by the invention has a simpler structure, a high-precision grating is not needed, and the spectrum resolution is determined by the resolution of time delay dispersion; the use of various filters is reduced, because the excitation light and the spectrum are detected separately in time, and no crosstalk exists between different wavelengths; the full spectrum measurement can be realized by using a single-channel probe, and the segmented measurement or the moving of the grating is not needed; the excitation wavelength can be adjusted at will; the interference of background fluorescence can be eliminated through dual wavelengths, and the method is particularly suitable for being applied to Raman spectroscopy and CARS spectroscopy.
(3) The invention can measure the low wave number Raman spectrum. Because the traditional Raman spectrometer based on spatial dispersion needs to apply a long-pass boundary filter to suppress the scattering of the excitation laser in the spectrometer, the spectrum close to the wavelength of the excitation light is cut off; thus low wavenumber Raman spectrum (<500cm-1) And are often not measurable. The method of the time delay dispersion spectrum can avoid the use of the optical filter, thereby effectively collecting the Raman spectrum with low wave number.
(4) The invention is more advantageous for spectra generated by non-linear effects. Optical nonlinear effects produce spectra that typically require picosecond or femtosecond pulsed laser excitation. The shorter the pulse width and the higher the power density of the pulsed light, the stronger the signal of the nonlinear spectrum. Meanwhile, the spectrum can be generated instantaneously in picosecond or femtosecond time, when time gaps among pulses are used for delay spectrum collection, the spectrum can be divided into more parts, the overlapping part in time can be less, and the spectrum resolution ratio can be greatly improved.
(5) The single-channel high-speed high-sensitivity photoelectric detector provided by the invention comprises but is not limited to a photomultiplier tube (PMT), a silicon-based photomultiplier tube (SiPM), a photon counter, a Single Photon Avalanche Diode (SPAD), a modulation-demodulation photoelectric amplification detector and the like to realize spectrum detection. Because the detectors have the advantages of improving photoelectric amplification gain and suppressing signal noise by orders of magnitude compared with the traditional spectrograph, the invention greatly improves the sensitivity of spectral measurement and the photon utilization rate.
(6) In the invention, in a time delay dispersion module, photons with different wavelengths in a spectrum are sequentially diffracted to different angles for emergence by using a grating or other space dispersion elements, and then enter two nearly parallel reflectors for multiple reflection and return to the grating in the original path; the structure of the spectrometer can be greatly simplified, and various optical filters required in the traditional spectrometer are removed. And the other two types of delay dispersion modules use dispersion optical fibers or wave splitters, so that the grating can be further and completely removed, the structure is simpler, the miniaturization can be further realized, and the reliability is higher.
(7) The invention combines various specific Raman labeled probes, and the gene targets a plurality of specific proteins or specific RNAs of organisms to realize the identification and multicolor high-speed imaging of proteome and RNA transcriptome.
Drawings
FIG. 1 is a schematic diagram of a delayed dispersion spectroscopy measurement based on pulsed excitation, wherein (a) is a schematic diagram of a spectrum obtained according to a prior art spatial dispersion spectroscopy-based method, and (b) is a schematic diagram of a spectrum obtained according to a delayed dispersion spectroscopy method provided by the present invention;
FIG. 2 is an energy level transition diagram of the Raman process and the CARS process provided by the prior art;
FIG. 3 is a flow chart of an implementation of the pulse-type time-delay dispersion spectral measurement method provided by the present invention;
FIG. 4 is a schematic block diagram of a pulse-type time-delay dispersion spectrum measuring apparatus according to the present invention;
FIG. 5 is a schematic structural diagram of a delayed dispersion spontaneous Raman spectrum imaging apparatus based on pulse excitation according to an embodiment of the present invention;
FIG. 6 is a schematic structural diagram of a time-delay dispersion CARS spectral imaging device provided by an embodiment of the invention;
FIG. 7 is a schematic diagram of the reverse time-dispersive broad-spectrum pumping and Stokes light generation CARS provided by the present invention;
fig. 8 is a schematic structural diagram of three delay dispersion modules provided by an embodiment of the present invention, wherein (a) is a schematic diagram of a delay dispersion principle; (b) a schematic structural diagram of a delay dispersion module implemented based on an optical fiber delay dispersion principle provided in a first embodiment; (c) a schematic structural diagram of a delay dispersion module implemented based on a mirror delay dispersion principle according to a second embodiment; (d) the third embodiment provides a schematic structural diagram of a delay dispersion module implemented based on the wavelength division device delay dispersion principle.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
The invention provides a novel spectral measurement and spectral imaging method and system based on pulse excitation and time delay dispersion, and aims to solve the problems of low spectral measurement efficiency and low spectral imaging speed caused by insufficient detection sensitivity of a spectrometer in the existing spectral measurement and spectral imaging technology. The invention skillfully combines short pulse laser, time delay dispersion spectrum and high-speed and high-sensitivity detector, breaks through the concept of the traditional spatial dispersion spectrometer, realizes the time delay dispersion spectrometer on the time dimension, and can finish the spectrum measurement by only utilizing the high-sensitivity probe of one channel. The method for measuring the time-delay dispersion spectrum gets rid of the dilemma that the existing spatial dispersion spectrometer can only adopt an inefficient linear array camera or a low-sensitivity photoelectric conversion array to obtain the spectrum, and can greatly improve the measurement sensitivity by utilizing a high-gain or single-photon detector. The method comprises the steps of firstly utilizing short pulse light to excite a sample to generate an instantaneous spectrum, then separating photons in the spectrum in time according to wavelength or other spectral characteristics, and finally utilizing the photon detection advantages of a single high-speed high-sensitivity photoelectric detector to detect different photons in the spectrum along with the time.
As shown in fig. 3, the present invention provides a method for measuring a pulse-type time-delay dispersion spectrum, comprising the following steps:
s1: exciting a sample to be detected by pulse laser and generating a transient spectrum;
s2: different photons in the transient spectrum are subjected to time delay processing and are separated in sequence;
s3: photons in the spectrum over time are detected using a high-speed, highly sensitive photodetector.
In step S1, the short pulse laser (specifically, the pulse laser with a pulse width time of 1 microsecond to 1 attosecond) interacts with the sample to generate a transient spectrum (1 microsecond to 1 attosecond), which includes a fluorescence spectrum (for analyzing electronic state energy level transitions of molecular atoms and molecules), a raman scattering spectrum (for analyzing vibrational state energy level transitions of molecules), an absorption spectrum (for analyzing vibrational state energy level transitions of molecules), or other spectra (including but not limited to rotation spectrum, infrared absorption spectrum, etc., which can help analyze chemical and physical properties of atoms and molecules).
In step S2, the different photons in the temporal spectrum are temporally delayed back and forth in order of wavelength or other spectral characteristics (energy, frequency, intensity, polarization, wavefront, phase, wave-vector direction) and dispersed until the next laser pulse arrives. The spectrum utilizes the time gap between the excitation pulses to perform enough time expansion, photons with different wavelengths or other spectral characteristics reach the detector at different moments, and meanwhile, the high-speed photoelectric detector can be ensured to be capable of sequentially detecting the photons with different wavelengths or spectral characteristics according to time, and then the spectrum is reversely deduced according to the reaching time.
In step S3, photons of different wavelengths or spectral characteristics arriving at different delays may be detected using a single-channel high-speed high-sensitivity photodetector. The single-channel high-speed high-sensitivity photoelectric detector comprises but is not limited to a photomultiplier tube (PMT), a silicon-based photomultiplier tube (SiPM), a photon counter, a Single Photon Avalanche Diode (SPAD), a modulation-demodulation photoelectric amplification detector and the like.
In an embodiment of the invention, the single pulse excitation and the detection time after the pulse are a period, and the spectra acquired in each period are completely the same and can be used for integration of the repeated signals. The repetition frequency of the pulse laser is the frequency of spectrum detection per second, the pulse and signal acquisition need to be accurately synchronized, and the Raman spectrum after the pulse can be accurately superposed and integrated. The pulse laser outputs a trigger signal of the laser pulse to the time-delay spectrum acquisition card in real time, and the laser pulse and the spectrum acquisition are synchronized.
The invention avoids the defect that the pixel array in a low-sensitivity camera must be used as a spectrum detector in the traditional spectrometer, opens up a time-delay dispersion spectrum method in a time domain, and adopts a single-channel high-sensitivity detector to detect the spectrum in a time-sharing way. Because the invention adopts the transient pulse laser excitation spectrum, different wavelengths are obtained in a time-sharing way after time delay dispersion. Therefore, the method for detecting different wavelengths in the spectrum in a time-sharing manner does not waste photons due to small duty ratio in the common time-sharing multiplexing method, but avoids the problem that only one wavelength is detected by a single-channel probe and the rest wavelengths are wasted. On the contrary, the sensitivity of spectral measurement is greatly improved by utilizing the advantages of a single-probe detector (generally having single-photon detection capability), weak photons in a spectrum are efficiently utilized, the spectrum acquisition speed is improved in magnitude, and high-speed spectral imaging is realized.
As shown in fig. 4, the present invention also provides a pulse-type time-delay dispersion spectrum measuring apparatus, including: the device comprises a pulse laser generating module, a time delay dispersion module and a photoelectric detection module, wherein the laser generating module is used for generating short pulse laser with the wavelength of lambda l and irradiating the short pulse laser onto a sample to be detected for exciting the sample to be detected to generate transient spectrum; the time delay dispersion module is used for carrying out time delay processing on different photons in the instantaneous spectrum and sequentially separating the photons; the photoelectric detection module is used for detecting photons coming in the spectrum along with time.
In an embodiment of the present invention, the time-delay dispersion module time-delays and disperses the different photons in the temporal spectrum in order of wavelength or other spectral characteristics (energy, frequency, intensity, polarization, wavefront, phase, wave-vector direction) until the next laser pulse arrives. The time delay generated by the photons with different wavelengths or spectral characteristics in the time delay dispersion unit corresponds to the wavelengths thereof one by one.
As an embodiment of the present invention, the delay dispersion module may use a dispersion fiber to delay photons with different wavelengths in a spectrum, and an exit port of the fiber ensures that a high-speed detector sequentially detects photons with different wavelengths in time.
As another embodiment of the present invention, the time-delay dispersion module may use a grating to sequentially diffract photons with different wavelengths in a spectrum to exit at different angles, and then enter two nearly parallel mirrors to perform multiple reflection and return to the grating. Because the path difference between the mirrors for photons of different wavelengths in the spectrum going back and forth is different, photons of different wavelengths produce different time delays. The high-speed detector is ensured to detect photons with different wavelengths in turn according to time at the light exit port.
As another embodiment of the present invention, a wavelength division device may be used in the delay dispersion module to distribute photons with different wavelengths in a spectrum to different fiber waveguides, and due to different lengths of the fiber waveguides, propagation distances of the photons with different wavelengths are different, and finally, the light of different paths is merged and output through an optical multiplexer, so that the photons with different wavelengths in the spectrum have different delays, and the high-speed detector is ensured to sequentially detect the photons with different wavelengths in time.
In the embodiment of the invention, the short pulse laser generated by the laser generation module can excite the transient fluorescence spectrum in the sample; the delay dispersion module delays and disperses different photons in the instantaneous fluorescence spectrum from front to back in time according to the wavelength sequence until the next laser pulse arrives. The fluorescence spectrum utilizes the time gap between the excitation pulses to perform sufficient time expansion, and ensures that the high-speed photoelectric detector can sequentially detect photons with different wavelengths according to time.
As an embodiment of the invention, the short pulse laser generated by the laser generation module can excite the transient spontaneous Raman scattering spectrum in the sample. The delay dispersion module delays and disperses different photons in the instantaneous spontaneous Raman scattering spectrum from front to back in time according to the wavelength sequence until the next laser pulse comes. The raman spectroscopy utilizes the time gaps between the excitation pulses to spread over sufficient time to ensure that the high-speed photodetector can sequentially detect photons of different wavelengths in time.
The advantages of the invention on the measuring device are that the spectrum detection sensitivity is improved in magnitude, the structure of the spectrometer is simplified, the whole device can be further miniaturized, the carrying capacity is improved, and the manufacturing cost is reduced.
Since the spectral measurement in the prior art is too inefficient and too slow, if spectral imaging is achieved using the measurement in the prior art, a spectrum of 1000 x 1000 pixels would require 1 million spectra to be collected in a short time, several days, which is completely impractical. However, since the present invention can increase the efficiency and speed of spectral measurement by orders of magnitude, spectral imaging can be further achieved.
The invention also provides a spectral imaging method based on pulse type delay dispersion spectral measurement, which can solve the technical problem of low spectral imaging speed caused by low spectral measurement efficiency in the prior art. For example, the imaging method can be realized by conventional technical means such as moving a sample stage, or scanning a sample with laser to perform imaging. The specific implementation steps of the imaging method are prior art and are not described herein.
In the present invention, the spectral imaging mode of the sample in three-dimensional space can be applied to, but is not limited to, three-dimensional laser scanning or three-dimensional sample translation stage.
The invention can also be combined with Surface Enhanced Raman Scattering (SERS) to realize more sensitive Raman spectrum detection and imaging and realize the separation of Raman spectrum and background spectrum.
The spectral imaging method provided by the embodiment of the invention can also be combined with a specific Raman label, including but not limited to molecules generated by genetic modification and similar to HBI in a GFP protein structure; dye molecules with strong Raman signals and carbon-carbon double bonds or carbon-carbon triple bonds or other pi bonds, such as rhodamine 800 or similar molecules thereof and the like; genes are targeted to multiple specific proteins or specific RNAs of an organism, enabling the identification and multicolor imaging of proteomes and RNA transcriptomes.
In the embodiment of the invention, the Pump light (Pump) and the Stokes light (Stokes) are short pulse lasers which are overlapped in space and time, and transient Anti-Stokes Raman Scattering (CARS) spectra in the sample are excited due to a nonlinear effect. The delay dispersion module delays and disperses different photons in the instantaneous CARS spectrum from front to back in time according to the wavelength sequence until the next laser pulse arrives. The CARS spectrum utilizes the time gap between excitation pulses to perform sufficient time expansion, and ensures that the high-speed photoelectric detector can detect photons with different wavelengths in sequence according to time.
Wherein, pumping light (Pump) is a femtosecond laser with wide line width or a supercontinuum femtosecond laser; stokes light (Stokes) is a narrow-band pulsed laser, which can achieve CARS spectra.
Alternatively, both the Pump light (Pump) and Stokes light (Stokes) are wide linewidth femtosecond lasers, or both are supercontinuum femtosecond lasers; but the time dispersion of the two is just opposite, and the CARS spectrum can be generated. Because the method adopts two beams of femtosecond light, photons are not wasted, and the efficiency of CARS spectrum generation is higher.
To further illustrate the pulse-type time-delay dispersion spectroscopy measurement method, apparatus and spectral imaging method provided by the embodiments of the present invention, the following is detailed with reference to the accompanying drawings and specific examples:
example 1: time-delay dispersion spontaneous Raman spectrum measurement and spectrum imaging example based on pulse excitation.
FIG. 1 is a schematic diagram of a time-delay dispersion spectroscopy measurement based on pulsed excitation, wherein (a) is a schematic diagram of a spectrum obtained according to a prior art spatial dispersion spectroscopy-based method, and (b) is a schematic diagram of a spectrum obtained according to a time-delay dispersion spectroscopy method provided by the present invention; as can be seen in fig. 1 (a): the spectrum is dispersed and separated in space, a plurality of pixels of a camera array of the applied spectrometer are used for detecting different components of the spectrum, but the sensitivity of the array pixels is generally low, and the photon utilization rate is poor. As can be seen in fig. 1 (b): the spectra are separated in the time dimension, the dotted line is an exciting light short pulse, the solid line is the spectrum delayed in time, and the high-speed high-sensitivity single-channel photoelectric probe can detect the delayed spectral components in time sequence. The time-delay dispersion spectrum can be repeatedly measured and integrated in time.
In the invention, a sample is excited to generate an instantaneous spectrum by a repeated sequence of short pulse laser, photons with different wavelengths in the spectrum are delayed and separated in sequence according to the wavelength sequence on a time gap between excitation pulses by utilizing a delay dispersion module, and finally, the photons with different wavelengths are detected in sequence according to time by adopting a high-speed high-sensitivity photoelectric detector, and the spectrum is reversely pushed according to the arrival time. The detection time after a single pulse excitation and the pulse is one period, and the collected spectra in each period are completely the same and can be used for the integration of repeated signals. The repetition frequency of the pulse laser is the frequency of spectrum detection per second, the pulse and signal acquisition need to be accurately synchronized, and the spectrum after the pulse can be accurately superposed and integrated.
Fig. 2 shows energy level transition diagrams of the raman process and the CARS process provided by the prior art, wherein the raman process is: molecules in the sample are excited by the laser to generate Raman scattering photons; the CARS process comprises the following steps: molecules in the sample are subjected to phase-locked pulsed laser, Pump light and Stokes light (the energy difference between the two photons is equal to the transition energy of the molecular ground state), and generate Anti-Stokes photons. In the raman spectroscopy measurement, after laser light is scattered by a sample, as shown in fig. 2, the energy of the scattered photon and the photon energy of excitation light differ by a vibration energy level difference of one molecule. The molecular species and relative concentration in the sample can thus be identified by detecting the spectrum of the raman scattered photons.
The embodiment of the invention also provides a time-delay dispersion Raman spectrum measurement and spectrum imaging device based on pulse excitation, and referring to FIG. 5, various system parameters in the device can be selected according to actual conditions.
The pulse type time delay dispersion spectrum imaging device provided by the embodiment of the invention comprises: the system comprises a pulse laser generating module 1, a first reflecting module 102, a laser scanning module 103, an objective lens focusing module 104, a sample stage 105, a photon collecting module 106, a time-delay dispersion module 2, a photoelectric detection module 3 and an acquisition module 4; the pulse laser generating module 1 is used for generating pulse laser; the first reflection module 102 is configured to reflect the pulsed laser to the laser scanning module 103; the laser scanning module 103 is used for spatially scanning laser and realizing imaging; the objective lens focusing module 104 is used for focusing laser on the sample; the sample stage 105 is used for placing a sample; the photon collection module 106 is used for collecting the transient spectrum generated by the sample and outputting the transient spectrum after collimating the transient spectrum; the delay dispersion module 2 is used for carrying out time delay processing on different photons in the transient spectrum and sequentially separating the photons according to a time sequence; the photoelectric detection module 3 is used for detecting photons coming from the spectrum along with time; the acquisition module 4 is used to acquire the detected photons.
The imaging device works as follows: the pulse laser enters a three-dimensional or two-dimensional laser scanning module 103 and an objective lens focusing module 104 of the imaging microscope through a first reflection module 102, and scanning and imaging of the laser on a sample are completed. The laser scanning system may be a combination of scanning galvanometers and other laser scanning modes. The three-dimensional displacement sample stage 105 can also perform three-dimensional scanning of the sample. Raman photons generated by the interaction of the laser with the sample pass through the photon collection module 106 and enter the time-delay dispersion module 2 (see example 3 and fig. 8 for details). The single-channel high-speed high-sensitivity detector performs photoelectric detection, and the acquisition module 4 transmits the spectrum to a computer or other data terminals.
As an embodiment of the present invention, the photon collection module 106 may be a forward detection photon collection module or a backward detection photon collection module (dotted line in fig. 5); the forward detection photon collection module is used for collecting and outputting an instantaneous spectrum generated by the sample in the direction of penetrating the sample; and the backward detection photon collection module is used for collecting and outputting the transient spectrum generated by the sample in the opposite direction of the exciting light.
Specifically, the pulse laser spontaneous Raman light source can emit laser pulses with the repetition frequency of 1MHz and the pulse width of 1ns, the average power of the laser is 100 mW-200 mW, and the wavelength is 532 nm. The pulse laser enters a three-dimensional or two-dimensional laser scanning module 103 and an objective lens focusing module 104 of the imaging microscope through a first reflection module 102, and scanning and imaging of the laser on a sample are completed. The laser scanning system can be a scanning galvanometer or other laser scanning modesAnd (4) combining. The three-dimensional displacement sample stage 105 can also perform three-dimensional scanning of the sample. Raman photons generated by the interaction between the laser and the sample enter the core delay dispersion module 2 through the photon collection module 106, which may be a confocal collection system or an optical fiber collection system (see example 3 and fig. 8 for details), and thus delay dispersion of different wavelength components in the raman photons is achieved. The delay dispersion module can also be entered by backward probing (dashed part in fig. 5). Since the repetition rate of the laser pulses is 1MHz, the time gap between each pulse is 1000 ns. The time-delay dispersion module collects the Raman Stokes photons, the wavelength of which is usually 533 nm-680 nm and approximately covers Raman shift of 0-4000 cm-1The delay is divided into 1000 segments in 1000 ns. After time delay, photons with different wavelengths enter a single-channel high-speed high-sensitivity detector in sequence according to the wavelength and time to carry out photoelectric detection. The detector's rate of utilization is 1GHz, samples a wavelength every 1 ns. A raman spectrum can be acquired every 1 microsecond emanating from each pulse impulse. Through accurate time sequence control, 100 ten thousand Raman spectra can be collected for 1 second to carry out integral superposition, and the Raman spectrum with higher signal-to-noise ratio is obtained. Finally, the spectrum is transmitted to a computer or other data terminal through the acquisition module 4. Similarly, sample imaging of raman spectra can be achieved by waiting for the spectra to be acquired after each pulse or pulses, moving the laser to a new position, and acquiring a new raman spectrum. The invention realizes the spectral measurement and collection only by the single-channel high-sensitivity probe, and the utilization efficiency of Raman photons and the Raman spectral imaging speed can be greatly improved.
The example is equally applicable to measurement of fluorescence spectra and fluorescence spectral imaging.
Example 2: time-lapse dispersive CARS spectroscopic measurement and CARS spectroscopic imaging examples.
When the excitation laser is two synchronous and phase-locked pulse light sources omegaP、ωsWherein ω isPAnd ωsTypically pump light and stokes light; when ω isPAnd ωsWhen the energy difference between photons is consistent with the molecular vibration energy level difference, anti-Stokes will be generatedPhoton omegaCARSAs shown in fig. 2.
The embodiment of the invention provides a method for delayed dispersion CARS spectral measurement and CARS spectral imaging, please refer to FIG. 6, and various system parameters can be selected according to actual conditions.
The pulse type time delay dispersion spectrum imaging device provided by the embodiment of the invention comprises: the system comprises a dual-channel pulse laser generation module 200, a first power regulation module 201, a second power regulation module 202, a light modulation module 203, a time delay device 204, a second reflection module 206, a dichroic mirror module 205, a laser scanning module 103, an objective lens focusing module 104, a sample stage 105, a photon collection module 106, a time delay dispersion module 2, a photoelectric detection module 3 and an acquisition module 4; the dual-channel pulse laser generation module 200 is used for generating two paths of pulse lasers of pump light and Stokes light; the first power adjusting module 201 is used for adjusting the power of stokes light; the second power adjusting module 202 is configured to adjust the power of the pump light; the optical modulation module 203 is used for modulating the power or phase or polarization or time delay of the stokes light; the time delay 204 is used to temporally register the pulses of the pump light and the stokes light; the second reflection module 206 is configured to reflect the pump light to the dichroic mirror module 205; the dichroic mirror module 205 is configured to reflect the stokes light, and spatially coincide the pump light with the stokes light; the laser scanning module 103 is used for spatially scanning laser and realizing imaging; the objective lens focusing module 104 is used for focusing laser on the sample; the sample stage 105 is used for placing a sample; the photon collection module 106 is used for collecting the transient spectrum generated by the sample and outputting the transient spectrum after collimating the transient spectrum; the delay dispersion module 2 is used for carrying out time delay processing on different photons in the transient spectrum and sequentially separating the photons according to a time sequence; the photoelectric detection module 3 is used for detecting photons coming from the spectrum along with time; the acquisition module 4 is used to acquire the detected photons.
The imaging device works as follows: the dual-channel pulse CARS laser source emits two beams of femtosecond lasers: pump light omegaPumpAnd Stokes light omegaStokesStokes pulse laser passes through the first power regulating module201 are then modulated by the light modulation module 203. The time delayer 204 can adjust the time delay of the stokes light, the dichroic mirror module 205 completely spatially combines the stokes light with the pump light, and both the time delayer 204 and the dichroic mirror module 205 are used to ensure that the stokes light and the pump light pulses completely temporally and spatially coincide. The two beams of femtosecond light enter a three-dimensional or two-dimensional laser scanning module 103 and an objective lens focusing module 104 of the imaging microscope to complete the scanning imaging of the laser to the sample. The three-dimensional displacement sample stage 105 can also perform three-dimensional scanning of the sample. CARS photons generated by the interaction of the laser and the sample enter the time-delay dispersion module 2 after passing through the photon collection module 106. The core component, time-delay dispersion module 2 (see example 3 and fig. 8 for details), performs time-delay dispersion of different wavelength components in raman photons. The delayed photons with different wavelengths enter the single-channel high-speed high-sensitivity detector 3 in sequence according to the wavelength and the time to carry out photoelectric detection. Finally, the spectrum is transmitted to a computer or other data terminal through the acquisition module 4.
As an embodiment of the present invention, the photon collection module 106 may be a forward detection photon collection module, which is used for collecting and outputting an instantaneous spectrum generated by the sample in a direction of transmitting the sample, or a backward detection photon collection module (shown in a dotted line in fig. 6); and the backward detection photon collection module is used for collecting and outputting the transient spectrum generated by the sample in the opposite direction of the exciting light.
Specifically, the dual-channel pulse CARS laser source can emit two beams of femtosecond laser with repetition frequency of 1MHz and phase locking, pump light omegaPumpAnd Stokes light omegaStokesThe average power of the laser is 100 mW-1000 mW, and the wavelengths are respectively 800 nm-960 nm of the supercontinuum and 1040nm of the narrow band. The stokes pulse laser light is modulated by the optical modulation module 203 after passing through the first power adjustment module 201. The time delayer 204 can adjust the time delay of the stokes light, the dichroic mirror module 205 completely spatially combines the stokes light with the pump light, and both the time delayer 204 and the dichroic mirror module 205 are used to ensure that the stokes light and the pump light pulses completely temporally and spatially coincide. Two femtosecond laser beamsAnd entering a three-dimensional or two-dimensional laser scanning module 103 and an objective lens focusing module 104 of the imaging microscope to complete the scanning and imaging of the sample by the laser. The laser scanning system may be a combination of scanning galvanometers and other laser scanning modes. The three-dimensional displacement sample stage 105 can also perform three-dimensional scanning of the sample. CARS photons generated by the interaction of the laser and the sample enter the time-delay dispersion module 2 through the photon collection module 106 (see example 3 and FIG. 8 for details), and time-delay dispersion of different wavelength components in the Raman photons is completed. It is also possible to enter the delay dispersion module by backward probing (dashed line). Since the repetition rate of the laser pulses is 1MHz, the time gap between each pulse is 1000 ns. The time-delay dispersion module collects Raman Stokes photons, the wavelength of which is 650 nm-891 nm generally, and approximately covers the Raman shift of 2885cm-1～801cm-1The delay is divided into 1000 segments in 1000 ns. After time delay, photons with different wavelengths enter a single-channel high-speed high-sensitivity detector in sequence according to the wavelength and time to carry out photoelectric detection. The detector's rate of utilization is 1GHz, samples a wavelength every 1 ns. A raman spectrum can be acquired every 1 microsecond emanating from each pulse impulse. Through accurate time sequence control, 100 ten thousand Raman spectra can be collected for 1 second to carry out integral superposition, and the Raman spectrum with higher signal-to-noise ratio is obtained. Finally, the spectrum is transmitted to a computer or other data terminal through the acquisition module 4. Similarly, sample imaging of raman spectra can be achieved by waiting for the spectra to be acquired after each pulse or pulses, moving the laser to a new position, and acquiring a new raman spectrum. According to the invention, the CARS spectral measurement and collection are realized only by the single-channel high-sensitivity probe, and the utilization efficiency of CARS photons and the Raman spectral imaging speed can be greatly improved.
In addition, both the Pump light (Pump) and Stokes light (Stokes) may be wide linewidth femtosecond laser (10fs), or both may be supercontinuum femtosecond laser, but the difference of the central wavelengths of the two covers the vibration spectrum of the molecule and the time dispersion is just opposite, as shown in fig. 7, the CARS spectrum may also be generated. The pump light and the Stokes light are wide-line-width pulse lasers, and the photon energy difference of the pump light and the Stokes light covers the vibration spectrum omega of moleculesiAnd the time dispersion is just opposite, ensuring that only one molecular vibration is excited at a time to generate CARS photons.
Example 3: three specific structural examples of delay dispersion modules.
Embodiments of the present invention provide three examples of producing time-delay dispersion spectra. Please refer to fig. 8, and various system parameters and methods may be selected according to actual situations. The principle is shown in fig. 8(a), the instantaneous spectrum wavelength generated by the pulse laser is overlapped in time, and photons in the spectrum can be separated in time and wavelength and detected by using a time delay dispersion method.
Fig. 8(b) shows a structure of a delay dispersion module implemented based on the principle of optical fiber delay dispersion according to a first embodiment, where the optical fiber delay dispersion module includes: one end of the dispersion optical fiber 21 is connected with the photon collection module 106, and the other end is connected with the photoelectric detection module 3; the photon collection module 106 collects and couples all photons into the dispersive fiber 21. When photons with different wavelengths are emitted out at the outlet of the optical fiber in sequence along with time, the high-speed and high-sensitivity photoelectric detection module 3 performs photoelectric detection on the photons to generate a spectrum. The method is straightforward and does not require gratings or other dispersive elements.
Fig. 8(c) shows a structure of a delay dispersion module implemented based on the mirror delay dispersion principle according to a second embodiment, where the mirror delay dispersion module includes: the light splitting element 22, the conjugate module 23, the beam splitting module 24 and the time delay reflection module 25 are arranged in sequence; the light splitting element 22 is configured to split the photons collected by the photon collecting module 106, so that the photons with different wavelengths in the spectrum are diffracted to different angles in sequence and then emitted; the conjugate module 23 is configured to guide the outgoing light into the delay reflection module 25; the time-delay reflection module 25 is used for reflecting the guided light for multiple times and then returning the light to the original path, so that the path difference of the photons with different wavelengths in the spectrum from reflection to reflection is different, and the photons with different wavelengths generate different time delays; the beam splitting module 24 is used for transferring the returned delayed photons to the photo detection module 3.
The distance between the spectroscopic element 22 and the conjugate module 23 is the focal length of the conjugate module 23, the conjugate module 23 includes two lenses, and the distance between the two lenses is the sum of the focal lengths of the two lenses. The beam splitting module 24 is disposed at an arbitrary position between the light splitting element 22 and the time-delay reflection module 25.
In the embodiment of the present invention, the light splitting element 22 may employ a spatial dispersion element such as a grating or a prism. The working process of the mirror delay dispersion module is detailed by taking a grating as an example as follows:
after the photon collection module 106 collects and collimates the photons, all the photons enter a grating or other spatial dispersion elements for splitting, and the photons with different wavelengths in the spectrum are sequentially diffracted to different angles for emission. Then the light is guided into two nearly parallel delayed reflection modules 25 through a conjugate module 23 composed of two lenses to be reflected for multiple times and then returned. The included angle between the two reflecting plates in the time-delay reflecting module 25 is 0.001-1 degree; preferably, the angle is 0.01 degrees.
Because the path difference between the mirrors for photons of different wavelengths in the spectrum going back and forth is different, photons of different wavelengths produce different time delays. The delayed photons are transferred to the photoelectric detection module 3 by the beam splitting module 24 to be detected when returning. The high-speed detector is ensured to detect photons with different wavelengths in turn according to time at the light exit port. This particular example may achieve greater delay through the optical path difference of the different photons.
Fig. 8(d) shows a structure of a delay dispersion module implemented based on the wavelength division device delay dispersion principle according to a third embodiment, where the wavelength division device delay dispersion module includes: an optical wavelength splitter 26, an optical fiber waveguide 27, and an optical multiplexer 28, which are arranged in this order; the photon collection module 106 collects and couples all photons into the optical wavelength splitter 26. The optical wavelength splitter 26 distributes photons of different wavelengths in the spectrum into fiber optic waveguides 27 of different lengths; due to the different lengths of the optical fiber waveguides, the propagation distances of photons with different wavelengths are different, and finally, the light of different paths is merged and output to the photoelectric detection module 3 through the optical multiplexer 28.
The optical splitter 26 can split photons with different wavelengths into 100-1000 channels. Accordingly, 100 to 1000 optical fiber waveguides having different lengths may be provided in the optical fiber waveguide 27.
The optical fiber time delay dispersion module has simple structure and few required elements, and can be miniaturized as much as possible; the reflector time-delay dispersion module and the wave division device time-delay dispersion module can provide more dispersion time-delay, and are beneficial to improving the spectral resolution.
In addition, in the present invention, the single-channel high-speed high-sensitivity photodetection module 3 in all embodiments includes, but is not limited to, a photomultiplier tube (PMT), a silicon-based photomultiplier tube (SiPM), a photon counter, a Single Photon Avalanche Diode (SPAD), a modulation-demodulation photoelectric amplification detector, and the like. Compared with a multi-channel detector, the single-channel detector has higher photoelectric sensitivity and data transmission speed, and can also have larger detection area, thereby improving the detection efficiency of photons.
It will be understood by those skilled in the art that the foregoing is only a preferred embodiment of the present invention, and is not intended to limit the invention, and that any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the scope of the present invention.

Claims (12)

1. A method for measuring pulse type time delay dispersion spectrum is characterized by comprising the following steps:
s1: exciting a sample to be detected by pulse laser and generating a transient spectrum;
s2: different photons in the transient spectrum are subjected to time delay dispersion treatment and are sequentially separated according to a time sequence;
s3: the high-speed high-sensitivity photoelectric detector is adopted to detect photons in the spectrum along with time in a time-sharing mode, and the detected spectrum in each period is completely the same.
2. The method of claim 1, wherein in step S2, different photons in the temporal spectrum are delayed in time and dispersed by wavelength, frequency, phase, energy, polarization, wavevector direction, or intensity.
3. The method for spectral measurement according to claim 1 or 2, wherein in step S3, the high-speed and high-sensitivity photodetector employs a single-channel detection element including a photomultiplier tube, a silicon-based photomultiplier tube, a photon counter, a single photon avalanche diode, or a modem photo-amplification detector.
4. A spectral imaging method realized based on the spectral measurement method of claim 1.
5. A pulse type time delay dispersion spectrum measuring device is characterized by comprising a pulse laser generating module (1), a time delay dispersion module (2) and a photoelectric detection module (3);
the pulse laser generation module (1) is used for generating pulse laser and enabling the pulse laser to be incident on a sample to be tested to excite the sample to generate transient spectrum;
the time delay dispersion module (2) is used for carrying out time delay dispersion treatment on different photons in the transient spectrum and sequentially separating the photons according to a time sequence;
the photoelectric detection module (3) is used for detecting photons in a spectrum in a time-sharing manner along with time, wherein the detected spectrum in each period is completely the same.
6. A pulsed time-lapse, dispersive spectral imaging apparatus, comprising: the device comprises a pulse laser generating module (1), a first reflecting module (102), a laser scanning module (103), an objective lens focusing module (104), a sample stage (105), a photon collecting module (106), a time delay dispersion module (2), a photoelectric detection module (3) and an acquisition module (4);
the pulse laser generation module (1) is used for generating pulse laser;
the first reflection module (102) is used for reflecting the pulse laser to the laser scanning module (103);
the laser scanning module (103) is used for spatially scanning laser and realizing imaging;
the objective lens focusing module (104) is used for focusing laser on the sample;
the sample stage (105) is used for placing a sample and moving the sample to complete imaging;
the photon collection module (106) is used for collecting and outputting the instantaneous spectrum generated by the sample;
the time-delay dispersion module (2) is used for carrying out time-delay processing on different photons in the instantaneous spectrum and sequentially separating the photons according to a time sequence;
the photoelectric detection module (3) is used for detecting photons coming in the spectrum along with time;
the acquisition module (4) is used for acquiring the detected photons.
7. A pulsed time-lapse, dispersive spectral imaging apparatus, comprising: the system comprises a dual-channel pulse laser generation module (200), a first power regulation module (201), a second power regulation module (202), a light modulation module (203), a time delayer (204), a second reflection module (206), a dichroic mirror module (205), a laser scanning module (103), an objective lens focusing module (104), a sample stage (105), a photon collection module (106), a time-delay dispersion module (2), a photoelectric detection module (3) and an acquisition module (4);
the dual-channel pulse laser generation module (200) is used for generating two paths of pulse lasers of pump light and Stokes light;
the first power adjusting module (201) is used for adjusting the power of Stokes light;
the second power adjusting module (202) is used for adjusting the power of the pump light;
the optical modulation module (203) is used for modulating the power or the phase or the polarization or the time delay of Stokes light;
the time delayer (204) is for temporally registering the pulses of the pump light and the stokes light such that the pump light is temporally coincident with stokes light pulses;
the second reflection module (206) is configured to reflect the pump light to the dichroic mirror module (205);
the dichroic mirror module (205) is configured to reflect the stokes light and spatially coincide the pump light with the stokes light;
the laser scanning module (103) is used for spatially scanning laser and realizing imaging;
the objective lens focusing module (104) is used for focusing laser on the sample;
the sample stage (105) is used for placing a sample and moving the sample to complete imaging;
the photon collection module (106) is used for collecting and outputting the instantaneous spectrum generated by the sample;
the time-delay dispersion module (2) is used for carrying out time-delay processing on different photons in the instantaneous spectrum and sequentially separating the photons according to a time sequence;
the photoelectric detection module (3) is used for detecting photons coming in the spectrum along with time;
the acquisition module (4) is used for acquiring the detected photons.
8. Spectral imaging apparatus according to claim 6 or 7, wherein said photon collection module (106) is a forward-detection photon collection module or a backward-detection photon collection module;
the forward detection photon collection module is used for collecting and outputting an instantaneous spectrum generated by the sample in the direction of penetrating the sample;
and the backward detection photon collection module is used for collecting and outputting the instantaneous spectrum generated by the sample in the opposite direction of the exciting light.
9. The spectral imaging apparatus according to claim 7, wherein a CARS spectrum is generated when the pump light and the Stokes light are both broad linewidth femtosecond lasers or supercontinuum femtosecond lasers, and the difference in the central wavelengths of the pump light and the Stokes light covers the vibration spectrum of a molecule, and the temporal dispersions of the pump light and the Stokes light are exactly opposite.
10. The arrangement according to any of claims 6, 7, 9, wherein the delay-dispersion module (2) comprises: the dispersion optical fiber (21) is connected with the photon collection module (106) at one end and connected with the photoelectric detection module (3) at the other end; when photons with different wavelengths are emitted out sequentially at the outlet of the dispersion optical fiber (21) along with time, the photoelectric detection module (3) performs photoelectric detection on the photons and generates a spectrum.
11. The arrangement according to any of claims 6, 7, 9, wherein the delay-dispersion module (2) comprises: the device comprises a light splitting element (22), a conjugate module (23), a beam splitting module (24) and a time delay reflection module (25);
the light splitting element (22) is used for splitting the photons collected by the photon collecting module (106), so that the photons with different wavelengths in the spectrum are diffracted to different angles in sequence and then emitted;
the conjugation module (23) is used for guiding emergent light into the time delay reflection module (25);
the time-delay reflection module (25) is used for reflecting the guided light for multiple times and then returning the light back to the original path, so that the path difference of the photons with different wavelengths in the spectrum back and forth in the reflection is different, and the photons with different wavelengths generate different time delays;
the beam splitting module (24) is used for transferring the returned delayed photons to the photoelectric detection module (3).
12. The arrangement according to any of claims 6, 7, 9, wherein the delay-dispersion module (2) comprises: an optical wavelength splitter (26), an optical fiber waveguide (27), and an optical multiplexer (28);
the optical wave splitter (26) is used for distributing the photons collected by the photon collecting module (106) into the optical fiber waveguides (27) with different lengths;
the optical fiber waveguides (27) with different lengths enable the propagation distances of photons with different wavelengths to be different, and the photons with different wavelengths can generate different time delays;
the optical multiplexer (28) is used for combining the multiple paths of photons after the time delay and outputting the multiple paths of photons to the photoelectric detection module (3).
CN202010401969.5A 2020-05-13 2020-05-13 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device Active CN111504978B (en)

Priority Applications (2)

Application Number Priority Date Filing Date Title
CN202010401969.5A CN111504978B (en) 2020-05-13 2020-05-13 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device
PCT/CN2021/093540 WO2021228187A1 (en) 2020-05-13 2021-05-13 Pulse-type delay dispersion spectrum measurement method and apparatus, and spectral imaging method and apparatus

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202010401969.5A CN111504978B (en) 2020-05-13 2020-05-13 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device

Publications (2)

Publication Number Publication Date
CN111504978A CN111504978A (en) 2020-08-07
CN111504978B true CN111504978B (en) 2021-03-30

Family

ID=71868421

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202010401969.5A Active CN111504978B (en) 2020-05-13 2020-05-13 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device

Country Status (2)

Country Link
CN (1) CN111504978B (en)
WO (1) WO2021228187A1 (en)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111504978B (en) * 2020-05-13 2021-03-30 华中科技大学 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device
CN115165101A (en) * 2022-07-14 2022-10-11 华中科技大学 Single photon sensitivity ultrafast spectral measurement and spectral imaging device and method
CN116148227B (en) * 2023-04-23 2023-07-28 广东大湾区空天信息研究院 Time-resolved spectrum rapid measurement system and method
CN116818101A (en) * 2023-05-22 2023-09-29 中国航天三江集团有限公司 Non-resonant-eliminating modulation-free stable CARS detection system and method
CN117217321B (en) * 2023-11-09 2024-01-30 合肥硅臻芯片技术有限公司 Quantum image generation device and method

Family Cites Families (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4164373A (en) * 1978-01-12 1979-08-14 The United States Of America As Represented By The United States Department Of Energy Spectrometer employing optical fiber time delays for frequency resolution
JP3922993B2 (en) * 2002-09-30 2007-05-30 沖電気工業株式会社 Optical transmitter, optical receiver, and optical transmission system
CN100576946C (en) * 2006-06-22 2009-12-30 天津大学 The novel full light serial-parallel conversion method that is used for the high-speed light packet switching
WO2008127434A2 (en) * 2006-11-28 2008-10-23 The Regents Of The University Of California Time-resolved and wavelength-resolved spectroscopy for characterizing biological materials
CN101055265B (en) * 2007-05-16 2010-05-19 南京大学 Surface and near-surface stratification chromatography for functional gradient material
CN101532877B (en) * 2009-03-03 2014-05-07 中国海洋大学 Pulse spectrograph of single receiving device
WO2014168734A1 (en) * 2013-03-15 2014-10-16 Cedars-Sinai Medical Center Time-resolved laser-induced fluorescence spectroscopy systems and uses thereof
CN104375374B (en) * 2014-11-25 2017-02-08 北京理工大学 Ultra-fast laser continuous imaging device and method based on frequency domain space-time transformation
US10211587B2 (en) * 2015-03-31 2019-02-19 Versitech Limited Spatial chirped cavity for temporally stretching/compressing optical pulses
CN107167457A (en) * 2017-05-23 2017-09-15 北京理工大学 The confocal CARS micro-spectrometers method and device of transmission-type
JP2019207338A (en) * 2018-05-30 2019-12-05 オリンパス株式会社 Observation device and objective lens
CN109799671A (en) * 2019-03-05 2019-05-24 北京理工大学 Multifrequency pulse ultrafast laser continuous imaging device and method based on super continuous spectrums
CN110351000B (en) * 2019-08-15 2020-09-15 中国科学院半导体研究所 All-optical serial-parallel conversion system based on wavelength division multiplexing technology
CN110579462B (en) * 2019-09-18 2020-07-10 华中科技大学 Time-resolved broad-spectrum CARS spectral imaging device based on high-repetition-frequency femtosecond laser
CN110755042B (en) * 2019-10-21 2021-01-01 华中科技大学 Time pulse light sheet tomography method and system for realizing large-volume high-resolution
CN111504978B (en) * 2020-05-13 2021-03-30 华中科技大学 Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device

Also Published As

Publication number Publication date
WO2021228187A1 (en) 2021-11-18
CN111504978A (en) 2020-08-07

Similar Documents

Publication Publication Date Title
CN111504978B (en) Pulse type time-delay dispersion spectral measurement method and device and spectral imaging method and device
Dobryakov et al. Femtosecond pump/supercontinuum-probe spectroscopy: Optimized setup and signal analysis for single-shot spectral referencing
US8693004B2 (en) Dual-etalon cavity ring-down frequency-comb spectroscopy with broad band light source
US8089625B2 (en) Time-resolved and wavelength-resolved spectroscopy for characterizing biological materials
JP2012032183A (en) Sample observation device and sample observation method
KR20120001533A (en) A device for measuring fluorescence lifetime and a method thereof
CN110836883A (en) SPAD-based time-dependent Raman-fluorescence lifetime spectrometer
CN111122535B (en) High-spectrum rapid imaging measurement system for molecular vibration mode
US20100014076A1 (en) Spectrometric apparatus for measuring shifted spectral distributions
Michalet et al. High-throughput single-molecule fluorescence spectroscopy using parallel detection
CN114460061A (en) Time-gated Raman spectrum system and Raman spectrum correction method
CN115855252B (en) Single photon sensitivity ultrafast spectrum measurement and spectrum imaging device and method
JP2000283960A (en) Micro-chip electrophoretic device
US20220187126A1 (en) Broadband pulsed light source apparatus
CN112113939A (en) Fluorescence lifetime imaging method and device based on spectral technology
CN115046987B (en) Time-gated Raman spectrum system and time synchronization compensation method thereof
EP4027120B1 (en) Apparatus and method for measuring spectral components of raman scattered light
US20210239590A1 (en) Apparatus and Method for Cyclic Flow Cytometry Using Particularized Cell Identification
CN101446546A (en) Dispersion direct reading spectroscopic detector
CN219038184U (en) Time resolution Raman spectrum device
CN109580582A (en) A kind of laser Raman spectrometer based on compound Amici Prism group
CN113203727B (en) Spectrum measuring device and method
CN116148227B (en) Time-resolved spectrum rapid measurement system and method
CN219015470U (en) Time-gated Raman spectrum system
Waddell et al. A fiber-optic-based multichannel time-correlated single-photon-counting device with subnanosecond time resolution

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant