CN107037031A - The confocal CARS micro-spectrometers method and device of reflection type differential - Google Patents
The confocal CARS micro-spectrometers method and device of reflection type differential Download PDFInfo
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Abstract
The invention belongs to microspectrum imaging detection technical field, it is related to a kind of confocal CARS micro-spectrometers method and device of reflection type differential.The core concept of the present invention is twin-laser as light source activation Reyleith scanttering light and the CARS light for being loaded with sample spectral characteristic, lossless separation is carried out to Reyleith scanttering light and CARS light using dichroic optical system, wherein Reyleith scanttering light carries out geometry detection and positioning, and CARS light carries out spectrographic detection.The present invention utilizes accurate this corresponding characteristic in differential confocal curve zero crossing and focal position, accurate capture and localized excitation light spot focus position, the spectrographic detection of high-precision geometry detection and high-space resolution is realized, a kind of method and apparatus of achievable sample microcell high-space resolution spectrographic detection are constituted.By combining CARS microtechnics, the time of what is inspired the be loaded with Raman diffused light of sample message is short than traditional spontaneous Raman effect, and energy quick nondestructive is detected to sample.The present invention has the advantages that accurate positioning, high-space resolution, lossless detection, spectral detectivity are high, is that microscopic spectrum detection and dimensional measurement provide a kind of new approach.
Description
Technical field
The invention belongs to microspectrum technical field of imaging, it is related to a kind of confocal CARS micro-spectrometers of reflection type differential
Method and device, available for microcell anti-Stokes scattering (CARS) spectrum of all kinds of samples of quick detection, can be achieved high spatial
Resolution imaging and detection.
Technical background
Light microscope is widely used in biomedical sector and materials science field, and quick with modern science
Development, the requirement to micro-imaging is also imaged from structure imaging turning function.Nineteen ninety, confocal Raman spectra microtechnic into
Work(application, greatly improves the possibility for exploring the specific structural constituent of small items and pattern.It is by confocal microscopy and drawing
Graceful spectral technique is combined, the high-resolution tomography feature for possessing confocal microscopy, has nodestructive testing and spectrum analysis concurrently again
A kind of ability, it has also become technological means of important material structure measurement and analysis, is widely used in physics, chemistry, biological doctor
The fields such as, material science, petrochemical industry, food, medicine, criminal investigation.
Traditional spontaneous Raman scattering imaging technique causes its transmission signal extremely weak due to Raman scattering self character, even if
With the laser excitation of high intensity, the good spectrum picture of a CONT is obtained, the action time grown still is needed very.This length
Time effect limits application of the Raman microscopy in biological field.Coherent based on coherent Raman effect is drawn
Graceful scattering (CARS) process can largely strengthen Raman signal, so as to realize quick detection.Coherent Raman effect is logical
Cross the light for being excited to excite molecule is locked in vibration level, the intensity of the vibration signal that this method is produced and exciting light it is strong
Non-linear relation is spent into, very strong signal, also referred to as coherent nonlinear Raman spectrum can be produced.There is very strong energy to turn for it
Change efficiency, the time for exposure is short, the infringement to sample is also smaller, at the same it scattering have certain directionality, easily with it is miscellaneous
Astigmatism is separated.
The generation of coherent anti-stokes raman scattering (CARS) is a third-order nonlinear optical processes, and it needs pumping
Light, stokes light and detection light.In general, the quantity in order to reduce light source, simplifies process, commonly use pump light and replace detection
Light, the relation between them is as shown in Fig. 2 as pump light (wp) and stokes light (ws) frequency difference and Raman active point
When the vibration frequency of son matches, CARS light w will be inspiredas, wherein was=2wp-ws.The generation process of CARS light is comprising specific
Raman active molecule vibration mode and cause interaction of the molecule from ground state to the incident field of excitation state vibrational transition
Process, its energy level schematic diagram is as shown in Figure 3.Fig. 3 (a) represents resonance Raman and the enhancing of off-resonance single photon to CARS processes
Contribution, Fig. 3 (b) represents that resonance Raman and non resonant two photon strengthen the contribution to CARS processes；When the frequency difference between Ep and Es
When matching with the vibration frequency of raman active molecule, the signal inspired obtains resonant check；
Traditional CARS microscopies using two single wavelength lasers, can only obtain the spectral information of specific frequency spectrum mostly, and
And tradition CARS microscopies do not emphasize system focus ability, cause actual spectrum detecting location to be often in defocus position.
Even if defocus position can also inspire the Raman spectrum of sample and by the spectrometer detection after pin hole, but intensity can not be reasonable
Characterize the correct spectral signal intensity of the point.In CARS microscopic systems, only when system is accurately focused, optimal sky could be obtained
Between resolving power and best spectroscopic probing capabilities.
Above-mentioned reason limits the ability that CARS microscopic systems detect microscopic spectrum, constrains it in finer microscopic spectrum
Test and the application in analysis occasion.Based on above-mentioned situation, the present invention proposes the strong of the sample surfaces scattering for arriving systematic collection
In sample Raman diffused light 103~106Reyleith scanttering light again carries out detected with high accuracy, it is organically blended with spectrum investigating system, enters
The synchronizing detection of row spatial positional information and spectral information, with the differential confocal of high spectral resolution that is realizing high-space resolution
CARS collection of illustrative plates is imaged and detected.
The core concept of patent of the present invention is as sharp from super continuous spectrums pulse laser and unicast long-pulse laser
Light emitting source, expands excitation spectrum scope, improves excitation of spectra intensity；Confocal detection light path is divided into two parts, two detections
The photoelectricity point probe of light path is respectively placed in before Jiao defocused equidistant position to realize differential detection, according to differential principle, realizes
Bipolarity actual zero point tracking measurement, accurately focuses, realizes high-space resolution；After accurately focusing, spectrographic detection is carried out, is obtained most
Good spectrally resolved ability.
The content of the invention
The purpose of the present invention is that, to overcome the deficiencies in the prior art, a kind of reflection type differential of high-space resolution of proposition is confocal
CARS micro-spectrometers method and its device.
The present invention is achieved by the following technical solutions.A kind of confocal CARS micro-spectrometer sides of reflection type differential
Method, comprises the following steps：
A) super continuous spectrums laser is sent by super continuous spectrums laser (3), after bandpass filter (4) by one two to
The Single wavelength laser that Look mirror (5) is sent with single wavelength laser (2) converges, by adjust light path make two light beams sequential it is consistent,
Space coincidence (Single wavelength laser envelope is in continuous spectrum laser)；Mixed light beam is by half-reflection and half-transmission prism (6) by microcobjective (8)
It is focused on sample (9), inspires Reyleith scanttering light and be loaded with the CARS light of sample (9) spectral characteristic；
B) CARS light and Reyleith scanttering light are divided into two beams after dichroscope (11), wherein the light beam comprising CARS light enters light
Probe unit (12) is composed, light beam of another beam comprising Reyleith scanttering light enters differential confocal probe unit (18)；In spectrographic detection unit
(12) in, the light beam comprising CARS light first passes through bandpass filter (13), filters out the non-CARS interference light in light beam, then passes through
First condenser (14), which is assembled, enters spectrometer (17), obtains CARS spectral informations I (λ)；The light beam of another Shu Zaiyou Reyleith scanttering lights
Two beams are divided into by half-reflection and half-transmission prism (19), carry out respectively it is burnt before, defocused detection, wherein light beam passes through the 3rd condenser
(20) filtered out after by the second pin hole (21) before Jiao after veiling glare by the first photodetector (22) detection, Jiao before obtaining
Signal I (u0-uM), another light beam by after the 4th condenser (23) by filtering out veiling glare positioned at the 3rd defocused pin hole (24)
Detected afterwards by the second photodetector (25), Jiao signal I (u after acquisition0+uM), wherein uMIt is that the first pin hole and the second pin hole are relative
In the offset distance of object focal point.
C) by preceding burnt signal I (u0-uM) and rear Jiao signal I (u0+uM) normalization subtract each other after, obtain differential confocal curve I
(uM), using " zero crossing " of differential confocal response curve with measuring objective focus positions accurately corresponding characteristic, pass through " zero passage
Point " triggering carrys out accurate capture and excites hot spot focal position, realizes the geometry detection and space orientation of high-space resolution.
D) sample (9) surface location (exciting light spot focus) is obtained by computer disposal, controls Z-direction translation stage
(7) drive microcobjective (8) mobile, make Laser Focusing on sample (9), the point is obtained by spectrographic detection unit (12)
CARS spectral informations.
E) signal of Reyleith scanttering light is individually handled, the three dimension scale tomography of high-space resolution can be achieved；Individually processing
CARS spectral signals, can obtain spectrum picture；Rayleigh signal and CARS spectral signals are handled simultaneously, and high-space resolution can be achieved
Collection of illustrative plates tomography, that is, realize the collection of illustrative plates imaging and detection of sample geometric position information and spectral information.
Particularly, in the methods of the invention, excitation beam is identical by two beam frequencies, the pulse laser composition of time consistency,
Wherein a branch of is unicast Long Pulse LASER, and as pump light and detection light, another beam is super continuous spectrums pulse laser, is used as this support
Ke Si light.
Particularly, in the methods of the invention, addition pin hole (15) may filter that veiling glare before spectrometer (17), improves spectrum and visits
The signal to noise ratio of survey.
Particularly, in the methods of the invention, excitation beam includes the light beams such as line polarisation, rotatory polarization, radial polarisation light
With the structure light beam generated by technologies such as pupil filterings, system spectrum Signal-to-Noise and system lateral resolution are thus improved.
Particularly, in the methods of the invention, by matching the optical filter of different spectral bands, the Stokes of different spectral coverage is selected
Light, it is possible to achieve the spectrographic detection of different spectral coverage；Wherein, the optical filtering bands of a spectrum of bandpass filter (4) and bandpass filter (13) are closed
It is symmetrical in the centre wavelength of single wavelength laser (2).
Particularly, in the methods of the invention, laser emission element (1) can also be brilliant with single wavelength laser (2) plus photon
Body optical fiber (34) carries out spectrum widening realization, in addition, the spectrometer (17) in spectrographic detection unit (12) is substituted for into photoelectricity point
Detector, rotatory polarization piece (38) can realize that spectral scan is exported, and then excite CARS spectrum and visited by photoelectricity point probe
Measure CARS spectral signals；
Include the invention provides a kind of confocal CARS micro-spectrometers device of reflection type differential：Excitation of spectra unit,
Laser emission element (1) positioned at excitation of spectra unit incident direction, the dichroic list positioned at excitation of spectra unit exit direction
First (11), the spectrographic detection unit (12) positioned at dichroic unit (11) transmission direction, positioned at dichroic unit (11) reflection side
To differential confocal probe unit (18), control whole system computer (39).Laser emission element (1) is by unicast long pulse
Laser (2), super continuous spectrums pulse laser (3), bandpass filter (4) and dichroscope (5) composition；Excitation of spectra unit by
Half-reflection and half-transmission prism (6), iris filter (32), Z-direction translation stage (7), microcobjective (8), testing sample (9) and X-Y translations
Platform (10) is constituted；Dichroic unit (11) is dichroscope (11)；Spectrographic detection unit (12) is by bandpass filter (13), first
Condenser (14), the first pin hole (15), second condenser lens (16) and spectrometer (17) composition；Laser differential confocal probe unit
(18) gathered by half-reflection and half-transmission prism (19), the 3rd condenser (20), the second pin hole (21), the first photodetector (22), the 4th
Light microscopic (23), the 3rd pin hole (24), the second photodetector (25) composition, wherein, the second pin hole (21) is located at the 3rd condenser
(20) before burnt, distance is S1, and the 3rd pin hole (24) is defocused positioned at the 4th condenser (23), and distance is S2, S1=S2.
In apparatus of the present invention, the angle of dichroscope (11) and Amici prism (6) the reflected beams axis is variable, may be selected
Appropriate angle is to meet the structure design of the present apparatus, so as to improve the flexibility of system.
In apparatus of the present invention, can by computer (39) control switch different multiples microcobjective, subsidiary and
Observation.
In apparatus of the present invention, Z-direction translation stage (7), X-Y translation stages (10), spectrographic detection unit (12), differential confocal are visited
Survey unit (18) and realize coordinated signals and measurement by computer (39).
Beneficial effect
The inventive method, contrast prior art has following innovative point：
1st, the present invention organically combines laser differential confocal microtechnic and CARS spectrographic detections technology, utilizes differential confocal
Curve " zero point " position excites hot spot focal position and detected with the accurate corresponding characteristic in microcobjective focal position, accurate catch
Spectral information, so as to realize spectrum tomography and the detection of high-space resolution.
2nd, using dichroscope is by Reyleith scanttering light and is loaded with the CARS light of sample message and is separated in the present invention, wherein Rayleigh
Light realizes that geometric position is detected into differential confocal probe unit, and CARS light realizes that CARS spectrum are visited into spectrographic detection unit
Survey, the spectral information of light spot focus is excited by precisely focusing capture, system spectrum detectivity is improved.In addition, two to
Color light-dividing device can adjustment angle as needed, facilitate structure to debug.
3rd, the present invention structurally and functionally combines differential confocal microscopic system and CARS spectrum imaging systems, can be real
The tomography of existing sample microcell geometric parameter, can realize the spectrographic detection of sample microcell, can also realize collection of illustrative plates tomography again.
The inventive method, contrast prior art has following remarkable advantage：
1st, the differential confocal detection mode that the present invention is used, utilizes the differential characteristic for subtracting each other noise cancellation, environment resistant interference
Ability is strong.
2nd, the present invention carries out high accuracy positioning using differential confocal technology to measurement focal beam spot, and focusing position is carried out
Real-time follow-up, eliminates the ambient influnence such as temperature and disturbance, realizes that accurate correspondence minimum excites focusing to the detection of CARS spectrums all the time
The sample spectra of spot area, greatly improves the microscopical microscopic spectrum detectivities of existing CARS and geometric position detection energy
Power.
3rd, the present invention is used as excitation source using super continuous laser matching Single wavelength laser, it is possible to achieve broad band CARS light
Spectrum detection.
Brief description of the drawings
Fig. 1 is Figure of abstract, i.e., the present invention is basic implements figure；
Fig. 2 is coherent (CARS) light excitation principle figure；
Fig. 3 is CARS light and pump light, the graph of a relation of stokes light；
Fig. 4 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential；
Fig. 5 is differential confocal response curve；
Fig. 6 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential for improving spectrographic detection；
Fig. 7 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential for adding iris filter；
Fig. 8 is the micro- method of testing schematic diagrames of the dichroscope direction confocal CARS of adjustable reflection type differential；
Fig. 9 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential of single laser；
Figure 10 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential that time domain is scanned；
Figure 11 is that the confocal CARS microscopic test devices figure of reflection type differential, i.e. embodiment are schemed；
In figure, 1- laser emission elements, 2- Single wavelengths LASER Light Source, 3- super continuous spectrums LASER Light Source, 4- bandpass filters,
5- dichroscopes, 6- Amici prisms, 7-Z to translation stage, 8- microcobjectives, 9- samples, 10-X-Y translation stages, 11- bis- to
Look mirror, 12, spectrographic detection unit, 13- bandpass filters, the condensers of 14- first, the pin holes of 15- first, 16- second condenser lens,
17- spectrometers, 18- differential confocals probe unit, 19- Amici prisms, the condensers of 20- the 3rd, the pin holes of 21- second, the light of 22- first
Burnt hot spot before electric explorer, the condensers of 23- the 4th, the pin holes of 24- the 3rd, the photodetectors of 25- second, 26- subtraction circuits, 27-,
Burnt curve, 31- non-cumulative curves, 32- iris filters, 33- polarization spectro ribs after burnt hot spot, 30- after burnt curve, 29- before 28-
Mirror, 34- photonic crystal fibers, the speculums of 35- first, 36- retarders, the speculums of 37- second, 38- polarizers, 39- light
Electric point probe, 40- computers；
Embodiment
The present invention is described in further detail with reference to the accompanying drawings and examples.
Fig. 4 is the confocal CARS microspectrums detection method schematic diagram of reflection type differential.First, from a unicast long pulse
Laser (2) sends pump light (detection light) as pump light source and probe source by it, then from a frequency with one of
The super continuous spectrums pulse laser (3) of cause is obtaining requiring wavelength model as Stokes light source after bandpass filter (4)
Interior continuous spectrum laser is enclosed, by adjusting optical texture, two beam laser is passed through dichroscope (5) time consistency, space coincidence；
Mixed light first passes through an Amici prism (6), after pass through a high x Microscope Objective being fixed on Z-direction translation stage (7)
(8) focus on sample (9)；Because in the case of tightly focused, the condition of phase matched is easily met, and inspires Reyleith scanttering light
With the CARS light for being loaded with sample spectral characteristic；CARS light and Reyleith scanttering light arrive separately at spectrographic detection after dichroscope (11)
Unit (12) and differential confocal probe unit (18)；Wherein, spectrographic detection unit (12) carries out spectrographic detection to CARS light, differential
Confocal detection unit (18) carries out geometric position detection to Reyleith scanttering light.
In differential confocal probe unit (18), it is s that the second pin hole (21), which is located at the 3rd condenser (20) focus front distance,1
Position, by the first photodetector (22) obtain before burnt signal (28)；It is burnt that 3rd pin hole (24) is located at the 4th condenser (23)
Distance is s after point2Position (s1=s2), the burnt signal (30) after the second photodetector (25) acquisition；By preceding burnt signal and rear Jiao
Signal makes the difference, that is, obtains differential confocal curve (31), i.e. Fig. 5.
As shown in fig. 6, in spectrographic detection unit (12), the first pin hole of addition (15) filtering veiling glare, then gathered by second
Light microscopic (16), which assembles CARS light, enters spectrometer (17), thus improves the signal to noise ratio of spectrographic detection；
Iris filter (32) is added between Amici prism (6) and microcobjective (8) and produces structure light beam, will be obtained more
The spectrographic detection effect of high spatial resolution, i.e. Fig. 7.
In basic implementation of the invention, the angle of dichroscope (11) and Amici prism (6) the reflected beams axis can
Become, the position of corresponding confocal detection unit (18) can be adjusted, i.e. Fig. 8, this structure design is conducive to the inventive method
Realization.
Fig. 9 is the micro- method of testing schematic diagrames of the confocal CARS of reflection type differential of single laser, it is therefore an objective to by Laser emission
The twin-laser input of unit is changed to single laser input, reduces cost；Unicast long-pulse laser (2) sends Single wavelength and swashed
Light, through polarization splitting prism (33) light splitting, transmissive portion enters photonic crystal fiber (34) and carries out widening and filtered with band logical
Wave plate (4) carries out particular requirement wavelength interception, the speculum of reflecting part first (35), retarder (36) and the second reflection
Mirror (37) is coupled with the continuous spectrum laser after broadening at dichroscope (5) place afterwards, and output space is consistent, time consistency mixed
Closing light beam, the CARS excitations of spectra are carried out to sample.Wherein, the effect of optical time delay line (36) is to ensure two beam laser sequential
Overlap.
Spectrometer (17) in spectrographic detection unit (12) in Fig. 9 is substituted for photoelectricity point probe (39), in polarization point
Polarizer (38), i.e. pie graph 10 are added between light prism (33) and photonic crystal fiber (34)；By rotatory polarization piece (38),
Change the polarization state of light beam, the light beam for exporting photonic crystal fiber (34) realizes that spectral scan is exported, with reference to optical time delay line
(36) it can realize that time domain scans the CARS excitations of spectra, and then realize the CARS spectral measurements of broad band.
Embodiment
In the present embodiment, the picosecond laser that wavelength is 532nm is used as pump light source and probe source, using heavy
Consistent super continuous spectrums picosecond laser adds 550~650nmm bandpass filters as Stokes light source to complex frequency therewith,
Outgoing is mixed under conditions of space coincidence, time consistency is met, high power is passed through after obtaining structure light beam by iris filter
Microcobjective tightly focused now meets phase-matching condition on sample, inspire wave-length coverage 450~515nm it is anti-this
Lentor light (CARS) and the Reyleith scanttering light that wavelength is 532nm.
As shown in figure 11, it is the confocal CARS micro-spectrometers device of reflection type differential, its testing procedure is as follows：
First, in laser emission element (1), the continuous spectrum laser that super continuous spectrums laser (3) is sent is filtered by band logical
Mating plate (4) obtains 550~650 broad band laser after filtering, then send one-wavelength laser with Single wavelength (532nm) laser (2)
Converge at dichroscope (5) place, form mixed light beam, wherein, this two beams laser repetition rate is consistent, reaches dichroscope (5)
Time consistency, light beam can be completely superposed (the complete envelope of pump spot is in Stokes hot spot) after converging；Mixed light beam passes through
After Amici prism (6), iris filter (32) is reached, structure light beam is produced；Structure light beam passes through on Z-direction translation stage (7)
Microcobjective (8) tightly focused on sample (9), inspire Reyleith scanttering light and being loaded with sample (9) spectral characteristic
CARS light.
Now, Sample Scan can be accomplished by the following way：Z-direction is realized by the PZT (7) on microcobjective (8)
Scanning, realizes that the displacement of x-y directions is scanned, or add in the light path after laser emitting by the objective table (10) under sample (9)
Plus vibration mirror scanning structure, realize x-y scanning directions.
The light beam that sample is reflected back includes stokes light λs, pump light λ0, Reyleith scanttering light λ0, CARS light λas；Wherein,
CARS light λasWith stokes light λsInto spectrographic detection unit (12), 532nm pump light and Reyleith scanttering light is through dichroscope
(11) it is reflected into differential confocal probe unit (18).In spectrographic detection unit (12), by stokes light λsWith CARS light
λasThe bandpass filter (13) that the light of mixing first passes through 450~515nm only retains CARS light afterwards, then passes through the first condenser
(14) assemble by the first pin hole (15), filter and assembled again by second condenser lens (16) into spectrometer (17) after ambient light, from
And detect and obtain CARS spectrum I (λ), wherein λ is inspired the wavelength of CARS light by sample (9) stimulated luminescence.Differential
In confocal detection unit (18), Reyleith scanttering light λ0Half-reflection and half-transmission Amici prism (19) light splitting is first passed through, a semi-gloss passes through the 3rd
Condenser (20) is assembled by the pin hole (21) before Jiao, is finally detected by the first photodetector (22), is obtained preceding Jiao
Signal I (u0+uM)；Another semi-gloss is assembled by positioned at defocused pin hole (24) by the 4th condenser (23), finally by the second light
Electric explorer (25) is detected, Jiao signal I (u after obtaining0-uM).Differential subtraction module 26 is a subtraction circuit, can be by before
Burnt signal makes the difference with rear burnt signal, obtains differential wave:
I(uM)=I (uo+uM)-I(uo-uM)
Wherein, I (u0+uM) responded for preceding Jiao, I (uo-uM) responded for rear Jiao, I (uM) responded for differential confocal；uoFor axial direction
Skew, uMFor pin hole axial offset." zero point " of laser differential confocal response curve (31) and the focused spot of excitation beam
Accurate correspondence, the elevation information on sample (9) surface is obtained by " zero point " of response curve (31), with reference to connection translation stage
(10) positional information of displacement transducer feedback reconstructs the three-dimensional surface shape I of sample (9) by computer (40)
(x,y,z)。
Computer (40) control Z-direction translation stage (7) makes laser vernier focusing on sample (9) surface, and inspiring can be just
The CARS spectrum I (λ) of sample spectral characteristic is really characterized, is gathered by spectrographic detection unit (12), will by computer (40)
Positional information I (x, y, z) and spectral information I (λ) fusions, complete the three-dimensionalreconstruction and Spectrum Data Fusion I of sample (9)
(x,y,z,r).In addition, computer system is also extended through in whole system, computer (40) is used to realize to Z-direction translation stage (7), X-
The acquisition process and Data Fusion of the Bit andits controls of Y translation stages (10), differential confocal signal and CARS spectral signals.
More than, along laser emitting direction, it is sequentially placed laser emission element (1), Amici prism (6), in Amici prism (6)
Transmission direction places iris filter (32), Z-direction translation stage (7), microcobjective (8), sample (9), X-Y translation stages
(10) dichroscope (11), is placed in Amici prism (6) reflection direction, spectrographic detection is placed in dichroscope (11) transmission direction
Unit (12), differential confocal probe unit (18) is placed in dichroscope (11) reflection direction.Laser emission element (1) includes position
In the super continuous spectrums laser (3) of dichroscope (5) transmission direction, bandpass filter (4) and dichroscope (5) reflection direction
Unicast long-pulse laser (2)；Be sequentially placed in spectrographic detection unit (12) bandpass filter (13), the first condenser (14),
First pin hole (15), second condenser lens (16), spectrometer (17)；In differential confocal probe unit (18), in Amici prism
(19) transmission direction is sequentially placed the 3rd condenser (20), the second pin hole (21), the first photodetector (22), in Amici prism
(19) reflection direction is sequentially placed the 4th condenser (23), the 3rd pin hole (24), the second photodetector (25)；In whole system
In, unicast long-pulse laser (2), super continuous spectrums laser (3), X-Y translation stages (10), Z-direction translation stage (7), spectrometer
(17), the first photodetector (22), the second photodetector (25), subtraction circuit (26) are controlled by computer (40), are
The three dimensional local information and spectral information that system is obtained also carry out fusion treatment by computer (40).
The embodiment of the present invention is described above in association with accompanying drawing, but these explanations can not be understood to limitation
The scope of the present invention, protection scope of the present invention is limited by appended claims, any in the claims in the present invention base
The change carried out on plinth is all protection scope of the present invention.
Claims (10)
1. a kind of confocal CARS micro-spectrometers method and device of reflection type differential, it is characterised in that comprise the following steps：
A) super continuous spectrums laser is sent by super continuous spectrums laser (3), passes through a dichroscope after bandpass filter (4)
(5) the Single wavelength laser sent with single wavelength laser (2) converges, and makes that two light beams sequential is consistent, space by adjusting light path
Overlap (Single wavelength laser envelope is in continuous spectrum laser)；Mixed light beam is assembled by half-reflection and half-transmission prism (6) by microcobjective (8)
On sample (9), inspire Reyleith scanttering light and be loaded with the CARS light of sample (9) spectral characteristic；
B) CARS light and Reyleith scanttering light are divided into two beams after dichroscope (11), wherein the light beam comprising CARS light is visited into spectrum
Unit (12) is surveyed, light beam of another beam comprising Reyleith scanttering light enters differential confocal probe unit (18)；In spectrographic detection unit (12)
In, the light beam comprising CARS light first passes through bandpass filter (13), filters out the non-CARS interference light in light beam, then passes through first
Convergent mirror (14), which is assembled, enters spectrometer (17), obtains CARS spectral informations I (λ)；The light beam of another Shu Zaiyou Reyleith scanttering lights passes through
Half-reflection and half-transmission prism (19) is divided into two beams, carry out respectively it is burnt before, defocused detection, wherein light beam by the 3rd convergent mirror (20) after
Filtered out by the second pin hole (21) before Jiao by the first photodetector (22) detection after veiling glare, burnt signal I before obtaining
(u0-uM), another light beam by after the 4th convergent mirror (23) by being filtered out positioned at the 3rd defocused pin hole (24) after veiling glare by
Two photodetectors (25) are detected, Jiao signal I (u after acquisition0+uM), wherein uMIt is the first pin hole and the second pin hole relative to object lens
The offset distance of focus.
C) by preceding burnt signal I (u0-uM) and rear Jiao signal I (u0+uM) normalization subtract each other after, obtain differential confocal curve I (uM), profit
With the accurate corresponding characteristic of " zero crossing " of differential confocal response curve and measurement objective focus positions, by " zero crossing " trigger come
Accurate capture excites hot spot focal position, realizes the geometry detection and space orientation of high-space resolution.
D) sample (9) surface location (exciting light spot focus) is obtained by computer disposal, controls Z-direction translation stage (7) band
Dynamic microcobjective (8) is mobile, makes Laser Focusing on sample (9), the point is obtained by spectrographic detection unit (12)
CARS spectral informations.
E) signal of Reyleith scanttering light is individually handled, the three dimension scale tomography of high-space resolution can be achieved；Individually handle CARS light
Spectrum signal, can obtain spectrum picture；Rayleigh signal and CARS spectral signals are handled simultaneously, and the collection of illustrative plates layer of high-space resolution can be achieved
Analysis imaging, that is, realize the collection of illustrative plates imaging and detection of sample geometric position information and spectral information.
2. a kind of confocal CARS micro-spectrometers method of reflection type differential according to right 1, it is characterised in that exciting light
Beam is identical by two beam frequencies, the pulse laser composition of time consistency, wherein a branch of is unicast Long Pulse LASER, is used as pump light
With detection light, another beam is super continuous spectrums pulse laser, is used as stokes light.
3. a kind of confocal CARS micro-spectrometers method of reflection type differential according to right 1, it is characterised in that spectrometer
(17) addition pin hole (15) may filter that veiling glare before, improve the signal to noise ratio of spectrographic detection.
4. a kind of confocal CARS micro-spectrometers method of reflection type differential according to right 1, it is characterised in that exciting light
Beam includes the light beams such as line polarisation, rotatory polarization, radial polarisation light and the structure light beam generated by technologies such as pupil filterings, thus
Improve system spectrum Signal-to-Noise and system lateral resolution.
5. a kind of confocal CARS micro-spectrometers method of reflection type differential according to right 1, it is characterised in that by
Optical filter with different spectral bands, selects the stokes light of different spectral coverage, it is possible to achieve the spectrographic detection of different spectral coverage；Wherein,
The optical filtering bands of a spectrum of bandpass filter (4) and bandpass filter (13) are symmetrical on the centre wavelength of single wavelength laser (2).
6. a kind of confocal CARS micro-spectrometers method of reflection type differential according to right 1, it is characterised in that the present invention
Laser emission element (1) can also carry out spectrum widening reality with single wavelength laser (2) plus photonic crystal fiber (34) in method
Existing, in addition, the spectrometer (17) in spectrographic detection unit (12) is substituted for into photoelectricity point probe, rotatory polarization piece (38) can be with
Realize that spectral scan is exported, and then excite CARS spectrum and CARS spectral signals are obtained by the detection of photoelectricity point probe.
7. a kind of confocal CARS micro-spectrometers device of reflection type differential includes：Excitation of spectra unit, positioned at excitation of spectra list
The laser emission element (1) of first incident direction, the dichroic unit (11) positioned at excitation of spectra unit exit direction, positioned at two to
The spectrographic detection unit (12) of color element (11) transmission direction, the differential confocal positioned at dichroic unit (11) reflection direction are detected
Unit (18), the computer (39) for controlling whole system.Laser emission element (1) is by unicast long-pulse laser (2), super continuous
Compose pulse laser (3), bandpass filter (4) and dichroscope (5) composition；Excitation of spectra unit by half-reflection and half-transmission prism (6),
Iris filter (32), Z-direction translation stage (7), microcobjective (8), testing sample (9) and X-Y translation stages (10) composition；Dichroic
Unit (11) is dichroscope (11)；Spectrographic detection unit (12) is by bandpass filter (13), the first convergent mirror (14), the first pin
Hole (15), the second convergent mirror (16) and spectrometer (17) composition；Laser differential confocal probe unit (18) is by half-reflection and half-transmission prism
(19), the 3rd convergent mirror (20), the second pin hole (21), the first photodetector (22), the 4th convergent mirror (23), the 3rd pin hole
(24), the second photodetector (25) is constituted, wherein, the second pin hole (21) is located at before the 3rd convergent mirror (20) Jiao, and distance is S1,
3rd pin hole (24) is defocused positioned at the 4th convergent mirror (23), and distance is S2, S1=S2.
8. a kind of confocal CARS micro-spectrometers device of reflection type differential as claimed in claim 6, it is characterised in that two
It is variable to Look mirror (11) and the angle of Amici prism (6) the reflected beams axis.
9. a kind of confocal CARS micro-spectrometers device of reflection type differential as claimed in claim 6, it is characterised in that can lead to
Cross the microcobjective of computer (39) control switching different multiples, subsidiary and observation.
10. a kind of confocal CARS micro-spectrometers device of reflection type differential as claimed in claim 6, it is characterised in that Z-direction
Translation stage (7), X-Y translation stages (10), spectrographic detection unit (12), differential confocal probe unit (18) are real by computer (39)
Existing coordinated signals and measurement.
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