CN102608100A - System and method for carrying out CARS (Coherent anti-Stokes Raman Scattering) imaging by using four-wave mixing signals generated by optical fiber - Google Patents
System and method for carrying out CARS (Coherent anti-Stokes Raman Scattering) imaging by using four-wave mixing signals generated by optical fiber Download PDFInfo
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Abstract
The invention relates to a system and method for carrying out CARS (Coherent anti-Stokes Raman Scattering) imaging by using four-wave mixing signals generated by optical fiber. The system comprises a laser source, multiple reflecting mirrors, an optical parametric oscillator, a precise displacement platform, two long-wave-pass dichroscopes, a coupling objective lens, optical fiber, a two-dimensional scanner, a focusing lens, a supporting sleeve, a sample, a band-pass filter, an imaging lens, a detector and a computer. A light beam which is output after the laser source sends a frequency-doubling light beam for pumping the optical parametric oscillator, and a base-frequency light beam sent by the laser source are respectively pumping light and Stokes light for CARS imaging, and the four-wave mixing signals which are excited after the pumping light and the Stokes light are overlapped on time and space and then transmitted in the optical fiber are probe light for CARS imaging. CARS signal light which is generated after the pumping light and the Stokes light are focused on a sample are not overlapped with the frequency of the pumping light and the frequency of the Stokes light and can be separated by the long-wave-pass dichroscopes. Relative to the standard CARS system, the system disclosed by the invention is not newly provided with any device, has no special requirement on the used devices and is simplified in system and saved in cost.
Description
Technical field
The present invention relates to medical imaging technology and coherent anti-stokes raman scattering imaging technique, especially relate to a kind of four-wave mixing signal that produces in the optical fiber that utilizes and carry out coherent anti-stokes raman scattering method for imaging and system.
Background technology
Coherent anti-stokes raman scattering (Coherent anti-Stokes Raman scattering; Abbreviation CARS) technology is a kind of novel non-marked molecular resonance imaging technique that developed rapidly since 1999; It is carried out to picture based on the Raman scattering resonance mechanism and enhancement mechanism of chemical bond; Can be carried out to picture to the particular chemical composition in the sample that is left intact; Comprise chemical bonds (being respectively applied for the detection of liposome, DNA, protein and hydrate) such as C-H, O-P-O, Amide I, H-O, make it have chemo-selective, the specificity that also promptly forms images.Because the coherence of CARS, the high several magnitude of Raman scattering signal that its signal intensity ratio is common, thus under medium excitation light intensity condition, just can obtain the image taking speed of video rate.In addition, CARS also has the characteristics of high detection sensitivity, sub-micron spatial resolution and three-dimensional imaging.Therefore, the CARS technology has been widely used in virus, cell, biological tissue and living small animal imaging, drug delivery process monitoring, and research field such as pathological changes diagnosis.
The CARS microscope is made up of bulky ultrafast pulse LASER Light Source and flying-spot microscope usually, has hindered it and on human body, has carried out the application of living imaging.Therefore, development by optical fiber pass light hand-held or in peep type CARS imaging technique, become the necessary condition that the CARS technology is applied to clinical diagnosis.In the CARS imaging process, the frequency that overlaps on the room and time is ω
pPumping (Pump) light beam and frequency be ω
s(＜ω
p) Stokes (Stokes) light beam by strong focusing on sample the time, the generation beat frequency is ω
p-ω
sThe stack electromagnetic field.As beat frequency ω
p-ω
sResonant frequency Ω with specific molecular
VibCoupling promptly satisfies Ω
Vib=ω
p-ω
sThe time, cause molecular oscillation and give off intensity very high, frequency is ω
As=2 ω
p-ω
sAnti-Stokes (Anti-Stokes) photon, also be CARS signal photon, satisfy phase-matching condition between wherein pumping, Stokes and the anti-Stokes photon.Absorbed two pump photons and a Stokes photon; An and anti-Stokes photon that inspires; Constituted a four-wave mixing (Four-wave mixing is called for short FWM) mechanism, visible CARS is a kind of four-wave mixing optics mechanism that satisfies phase matching.Pass the nonlinear optics formation method of light by optical fiber with other; Produce imaging different (the only single ultrafast pulse light beams of transmission in optical fiber) like two-photon fluorescence excitation imaging and second harmonic; The CARS imaging need be transmitted in pump light and the stokes light that overlaps on time and the space in same optical fiber, thereby can in optical fiber, inspire the FWM signal of non-phase matching inevitably.The frequency of FWM signal photon is ω
Fwm=2 ω
p-ω
sAnd the CARS signal that is produced by sample is identical and can not separated go out, thereby can in imaging results, form very strong ground unrest, and its intensity tends to flood sample signal.Therefore, must be suppressed at the FWM signal that produces by optical fiber under the normal CARS image-forming condition.
Chen of University of California and (the M Balu of Potma group; Et al.Fiber delivered probe for efficient CARS imaging of tissues.Optics Express; 2010; 18 (3): result of study 2380-2388) shows, in single mode fused quartz optical fiber, double-clad photon crystal optical fibre and the big mode field area photonic crystal fiber that they measured, has all observed very strong FWM signal.In order to eliminate of the influence of this signal to imaging results; They have used a long-pass dichroic mirror to come filtering FWM signal and pump light and stokes light are passed through at the output terminal of optical fiber; The CARS signal that their incident sample excitation go out by same long-pass dichroic mirror reflects to side direction; Collect by an other root multimode fiber then, thereby from pumping signal, separated the CARS signal come out.Owing to used more device, this method is difficult to be used under size-constrained based endoscopic imaging occasion.(the Z Wang of Wong group of Cornell University; Et al.Coherent anti-Stokes Raman scattering microscopy imaging with suppression of four-wave mixing in optical fibers.Optics Express; 2011; 19 (9): 7960-7970) proposed a kind of method that changes the exciting light polarization state and suppressed the FWM signal: before pump light and stokes light are coupled into polarization maintaining optical fibre; Change their polarization state earlier through wave plate, make the polarization state of the two mutually orthogonal, at this moment they just transmit in optical fiber and can not produce the FWM signal; The two is after optical fiber output; (to pump light is λ/2 wave plates with a dual wavelength wave plate; Stokes light then is the λ wave plate) make the two turn back to consistent polarization state again; Thereby can inspire the CARS signal behind the incident sample, realize the purpose of using same Optical Fiber Transmission exciting light beam not disturbed by the FWM signal with receiving sample CARS signal.
Can find out that from these methods the existing thinking that addresses this problem all is to manage to suppress this unfavorable factor of FWM.Do you because the FWM signal is stronger, can consider to utilize it to carry out the CARS imaging so? Three look CARS imaging techniques provide possibility for the realization of this imagination.In aforesaid CARS imaging process, the molecule absorption in the sample two pump photon (ω
p) and a Stokes photon (ω
s), this is double-colored CARS imaging technique.Pump photon (ω of molecule absorption in sample
p), a Stokes photon (ω
s) and a probe photon (ω
Pr) after also can produce CARS signal (ω
As=ω
p+ ω
Pr-ω
s), such mechanism has constituted three look CARS imaging techniques.Owing to need three kinds of different ultrafast pulse light beams of frequency, this cost and space-time that can greatly increase LASER Light Source overlaps the difficulty of regulating, and therefore three look CARS imaging techniques are used also seldom.If the FWM signal that in optical fiber, produces as probe light; It and carry out three look CARS imaging by the pump light of Optical Fiber Transmission and stokes light; Do not propose any new requirement to light source this moment, and realized the coincidence on time and space between these three exciting lights.In addition; The pulsed light beam (optical parametric oscillator that substituted volume is huge) that inspires when utilizing high power ultrafast pulse light beam in optical fiber, to transmit; Be used as the exciting light source of CARS imaging; Be the research direction (relating to the miniaturization of CARS system) of present CARS imaging, this point is that above-mentioned imagination provides strong theory support.
Summary of the invention
In order to overcome disadvantages of background technology, the purpose of this invention is to provide a kind of elimination by the method and system of the FWM signal that passes the generation of light optical fiber to the imaging results adverse effect, this adverse effect is the intrinsic problem in the optical-fiber type CARS imaging system.The non-phase matching FWM signal that these method and system inspire when utilizing pump light and stokes light in optical fiber, to transmit is as the probe light of CARS imaging; It and constituted three look CARS imaging mechanism together by the pump light of Optical Fiber Transmission and stokes light; This moment is not overlapping with the excitation light frequency by the CARS flashlight that sample produces; The output terminal (no bulk restriction) that receives signal at optical fiber can come out the CARS Signal Separation with a long-pass dichroic mirror, thereby has realized using same Optical Fiber Transmission exciting light beam not receive the purpose of FWM signal interference with receiving sample CARS signal.
In order to realize said purpose, one aspect of the present invention provides the system that a kind of four-wave mixing (FWM) signal that utilizes optical fiber to produce carries out coherent anti-stokes raman scattering (CARS) imaging, and the technical scheme that said system technical solution problem is adopted is:
Comprise LASER Light Source, first catoptron, second catoptron, optical parametric oscillator, the 3rd catoptron, the 4th catoptron, the 5th catoptron, the 6th catoptron, the 7th catoptron, the 8th catoptron, precision displacement table, the 9th catoptron, the first long-pass dichroic mirror, the tenth catoptron, coupling object lens, optical fiber, two-dimensional scanner, condenser lens, supporting sleeve, sample, the second long-pass dichroic mirror, band pass filter, imaging len, detector and computing machine, wherein:
The frequency multiplication light beam that sends from the output port a of LASER Light Source is after first catoptron and second mirror reflects, and input and pump optical parametric oscillator are the pump light of CARS imaging by the light beam of optical parametric oscillator output; The basic frequency beam that sends from the output port b of LASER Light Source is the stokes light of CARS imaging; Stokes light successively after the 3rd catoptron, the 4th catoptron, the 5th catoptron, the 6th catoptron, the 7th catoptron, the 8th catoptron and the 9th mirror reflects to the first long-pass dichroic mirror, this light path is the Stokes light path;
The 6th catoptron and the 7th catoptron in the Stokes light path are installed on the precision displacement table; Through regulating precision displacement table and the 8th catoptron and the 9th catoptron, make respectively from the pump light of optical parametric oscillator output and stokes light to overlap in time and space after through the first long-pass dichroic mirror from the 9th catoptron; Pump light that overlaps on time and the space and stokes light are coupled in the object lens coupled into optical fibres and transmit behind the tenth mirror reflects to the second long-pass dichroic mirror; When pump light that overlaps on time and the space and stokes light transmit, inspire the four-wave mixing signal of non-phase matching in optical fiber, the probe light of four-wave mixing signal as the CARS imaging; Be focused lens focus on sample and inspire the CARS signal from pump light, stokes light and the probe light of optical fiber output; The end of optical fiber is fixed on the two-dimensional scanner, and makes optical fiber stretch out a bit of semi-girder that becomes, and two-dimensional scanner and condenser lens are installed in the supporting sleeve;
The CARS flashlight that produces by sample and by the pump light of sample retroreflection or scattering, stokes light and probe light, be focused that lens are collected and coupled into optical fibres in transmit, then behind the object lens that are coupled to the second long-pass dichroic mirror; By the pump light of sample retroreflection or scattering, stokes light and the probe light transmission second long-pass dichroic mirror, the CARS flashlight by the second long-pass dichroic mirror reflects after received by detector behind band pass filter and the imaging len;
The two-dimensional scanner band that computer control is installed in the supporting sleeve optical fiber end resonance; Make the pump light, stokes light and the probe light that are focused lens focus on sample, carry out two-dimensional scan; Computing machine is control detector synchronous acquisition CARS signal and transfer to computing machine and handle simultaneously, thereby obtains the two-dimentional CARS imaging results of sample.
Described LASER Light Source is the high-frequency impulse near-infrared laser light source with 2～7ps or 102fs magnitude pulse width, the frequency multiplication light beam that said high-frequency impulse near-infrared laser light source has the basic frequency beam that sends from output port b, sends from output port a.
Described optical parametric oscillator is that the pulse width with LASER Light Source is complementary, wavelength is at the adjustable laser generator of near-infrared band.
The direction of the light beam of described the 5th catoptron to the six catoptrons and the light beam of the 7th catoptron to the eight catoptrons is all parallel with the displacement adjusting direction of precision displacement table.
The input end end face of described optical fiber is positioned on the front focal plane of coupling object lens.
The light-sensitive surface of described detector is positioned on the front focal plane of imaging len.
In order to realize said purpose, the present invention provides a kind of four-wave mixing signal that utilizes optical fiber to produce on the other hand and carries out the CARS method for imaging, may further comprise the steps:
Step S1: the stokes light of the basic frequency beam of LASER Light Source output as the CARS imaging, the lentor light frequency is designated as ω
sThe pump light of the light beam that produces behind the frequency multiplication light beam pump optical parametric oscillator of LASER Light Source output and export as the CARS imaging, the pumping light frequency is designated as ω
p
Step S2:, stokes light and pump light are spatially overlapped after passing through the first long-pass dichroic mirror through regulating the catoptron in the Stokes light path;
Step S3:, stokes light and pump light are overlapped in time after passing through the first long-pass dichroic mirror through regulating the precision displacement table in the Stokes light path;
Step S4: regulate the pump light wavelength of output through the temperature of regulating gain media in the optical parametric oscillator, make the difference on the frequency of pump light and stokes light and the resonant frequency Ω of testing molecule chemical bond
VibBe complementary, promptly satisfy: Ω
Vib=ω
p-ω
s
Step S5: the frequency that inspires when in optical fiber, transmitting the pump light that overlaps on time and the space and stokes light is ω
Fwm=2 ω
p-ω
sNon-phase matching four-wave mixing signal, as the probe light of CARS imaging, the probe light frequency is designated as ω
Pr
Step S6: after focusing on the sample through condenser lens pump light, stokes light and probe light, inspire the CARS flashlight that satisfies phase-matching condition, CARS signal light frequency is ω
As=ω
p+ ω
Pr-ω
s
Step S7: by the CARS flashlight of sample generation; And by the pump light of sample retroreflection or scattering, stokes light and probe light be focused that lens are collected and coupled into optical fibres in transmit; Fiber-optic output have only the CARS flashlight by the second long-pass dichroic mirror reflects to detector, the light signal that detector receives is converted into and transfers to computing machine behind the electric signal and handle;
Step S8: computer control two-dimensional scanner band optical fiber end resonance, makes focused beam on sample, carry out two-dimensional scan, thereby obtains the two-dimentional CARS imaging results of sample.
Compare with background technology, the beneficial effect that the present invention has is:
1, the present invention has eliminated in the optical-fiber type CARS imaging system by the adverse effect of the FWM signal that passes the generation of light optical fiber to imaging results.The fiberize of CARS system is the necessary condition of its clinical practice; And the pump light and the stokes light that are used for the CARS imaging can inspire non-phase matching FWM signal inevitably when optical fiber transmits; This signal has identical frequency with the CARS signal that sample produces and can not separated go out, and the strong sample signal that consequently usually floods of its intensity.
2, the present invention has realized using same Optical Fiber Transmission exciting light beam and has received sample CARS signal and the purpose that not disturbed by the FWM signal.The existing way that addresses this problem is to transmit exciting light beam with an optical fiber, and the FWM signal that optical fiber produces comes filtering with a long-pass dichroic mirror, receives the CARS signal of sample then with another root optical fiber.
3, relative standard CARS system configuration of the present invention increases any device newly, and the device that uses is had no specific (special) requirements, has greatly simplified system and has practiced thrift cost.Adopt two optical fiber to transmit exciting light beam respectively and increased more device newly, in size-constrained, peep under the condition and be difficult to be used with the method that receives signal beams; And the method for utilizing the dual wavelength wave plate need be introduced the polarization state that some wave plates and polaroid change the incident exciting light beam, and the dual wavelength wave plate needs specific customization.
4, the present invention is applicable to the CARS imaging system of any fiber type, for realizing the CARS based endoscopic imaging simple and effective solution route is provided.
Description of drawings
Fig. 1 is a system schematic of the present invention;
Fig. 2 a is the energy level synoptic diagram of double-colored CARS imaging;
Fig. 2 b is the energy level synoptic diagram of three look CARS imaging;
Fig. 3 is the inventive method process flow diagram.
Symbol description among the figure:
1 LASER Light Source, 2 first catoptrons, 3 second catoptrons,
4 optical parametric oscillators, 5 the 3rd catoptrons, 6 the 4th catoptrons,
7 the 5th catoptrons, 8 the 6th catoptrons, 9 the 7th catoptrons,
10 the 8th catoptrons, 11 precision displacement table, 12 the 9th catoptrons,
14 the tenth catoptrons, 15 coupling object lens, 16 optical fiber,
17 two-dimensional scanners, 18 condenser lenses, 19 supporting sleeves,
20 samples, 22 band pass filters, 23 imaging lens,
24 detectors, 25 computing machines,
13 first long-pass dichroic mirrors, 21 second long-pass dichroic mirrors.
Embodiment
Below in conjunction with accompanying drawing and embodiment the present invention is further described:
As shown in Figure 1, the system that the four-wave mixing signal that utilizes optical fiber to produce that the present invention proposes carries out the CARS imaging comprises: LASER Light Source 1, first catoptron 2, second catoptron 3, optical parametric oscillator 4, the 3rd catoptron 5, the 4th catoptron 6, the 5th catoptron 7, the 6th catoptron 8, the 7th catoptron 9, the 8th catoptron 10, precision displacement table 11, the 9th catoptron 12, the first long-pass dichroic mirror 13, the tenth catoptron 14, coupling object lens 15, optical fiber 16, two-dimensional scanner 17, condenser lens 18, supporting sleeve 19, sample 20, the second long-pass dichroic mirror 21, band pass filter 22, imaging len 23, detector 24 and computing machine 25.
LASER Light Source 1 is for having 2～7ps or 10
2The high-frequency impulse near-infrared laser light source of fs magnitude pulse width, the frequency multiplication light beam that it has the basic frequency beam that sends from output port b, sends from output port a.The frequency multiplication light beam is after first catoptron 2 and 3 reflections of second catoptron, and input is pump optical parametric oscillator 4 also.Optical parametric oscillator 4 is that the pulse width with LASER Light Source 1 is complementary, wavelength is at the adjustable laser generator of near-infrared band, by the light beam of its output pump light (ω as the CARS imaging
p).The basic frequency beam that sends from the output port b of LASER Light Source 1 is as the stokes light (ω of CARS imaging
s), successively after the 3rd catoptron 5, the 4th catoptron 6, the 5th catoptron 7, the 6th catoptron 8, the 7th catoptron 9, the 8th catoptron 10 and 12 reflections of the 9th catoptron to the first long-pass dichroic mirror 13, this light path has constituted the Stokes light path.
The 6th catoptron 8 and the 7th catoptron 9 in the Stokes light path are installed on the precision displacement table 11.Through regulating precision displacement table 11 and the 8th catoptron 10 and the 9th catoptron 12, make respectively from the pump light of optical parametric oscillator 4 outputs with from the stokes light of the 9th catoptron 12 then to overlap in time and space through the first long-pass dichroic mirror 13; Pump light that overlaps on time and the space and stokes light are coupled transmission in object lens 15 coupled into optical fibres 16 after the tenth catoptron 14 reflexes to the second long-pass dichroic mirror 21; When pump light that overlaps on time and the space and stokes light transmit, inspire the four-wave mixing signal of non-phase matching in optical fiber 16, the probe light of four-wave mixing signal as the CARS imaging; Being focused lens 18 from pump light, stokes light and the probe light of optical fiber 16 output focuses on the sample 20 and inspires the CARS signal; The end of optical fiber 16 is fixed on the two-dimensional scanner 17, and makes optical fiber 16 stretch out a bit of semi-girder that becomes, and two-dimensional scanner 17 is installed in the supporting sleeve 19 with condenser lens 18;
The pump light that the first long-pass dichroic mirror 13 reflects from optical parametric oscillator 4, and see through stokes light from the 9th catoptron 12.The direction of the light beam of the 5th catoptron 7 to the 6th catoptrons 8 and the light beam of the 7th catoptron 9 to the 8th catoptrons 10 is parallel with the displacement of precision displacement table 11 adjusting direction; So that when overlap regulating, can guarantee from the direction of the light beam of the 9th catoptron 12 to first long-pass dichroic mirrors 13 constant in the time of carrying out pump light and stokes light through precision displacement table 11.
Through regulating the 8th catoptron 10 and the 9th catoptron 12 in the Stokes light path, pump light and stokes light are spatially overlapped after passing through the first long-pass dichroic mirror 13.Pump light and stokes light are coupled transmission in object lens 15 coupled into optical fibres 16 after 14 reflections of the tenth catoptron and seeing through the second long-pass dichroic mirror 21, the input end end face of optical fiber 16 is positioned on the front focal plane of coupling object lens 15.When the pump light that overlaps on time and the space and stokes light transmitted in optical fiber 16, can inspire frequency was ω
Fwm=2 ω
p-ω
sNon-phase matching four-wave mixing signal, the probe light of this signal as CARS imaging, its frequency is designated as ω
Pr
Exciting light from optical fiber 16 outputs comprises pump light, stokes light and probe light, is focused lens 18 and focuses on the sample 20, inspires the CARS signal that satisfies phase-matching condition, and its frequency is ω
As=ω
p+ ω
Pr-ω
sThe end of optical fiber 16 is fixed on the two-dimensional scanner 17, and makes optical fiber 16 stretch out a bit of formation cantilever beam structure, and two-dimensional scanner 17 is installed in the supporting sleeve 19 with condenser lens 18.Computing machine 25 control two-dimensional scanners 17 make the exciting light beam that is focused lens 18 focusing on sample 20, carry out two-dimensional scan with the bracketed part resonance of optical fiber 16.
The CARS flashlight that produces by sample 20 and by the pump light of sample 20 retroreflections or scattering, stokes light and probe light, be focused that lens 18 are collected and coupled into optical fibres 16 in transmission, then behind the object lens 15 that are coupled to the second long-pass dichroic mirror 21.By the pump light of sample 20 retroreflections or scattering, stokes light and the probe light transmission second long-pass dichroic mirror 21, the CARS flashlight is reflected by the second long-pass dichroic mirror 21 after band pass filter 22 is received by detector 24 with imaging len 23 backs;
The two-dimensional scanner 17 that computing machine 25 controls are installed in the supporting sleeve 19 is being with optical fiber end resonance; Make the pump light, stokes light and the probe light that are focused lens 18 focusing on sample 20, carry out two-dimensional scan; Computing machine 25 control detector, 24 synchronous acquisition CARS signals also transfer to computing machine 25 and handle, thereby obtain the two-dimentional CARS imaging results of sample.The light-sensitive surface of described detector 24 is positioned on the front focal plane of imaging len 23.
The second long-pass dichroic mirror 21 sees through by the pump light of sample 20 retroreflections or scattering, stokes light and probe light, and reflection CARS flashlight is to feeler arm.In feeler arm, the CARS flashlight is focused on the light-sensitive surface of detector 24 by imaging len 23 through band pass filter 22 backs, and all kinds of parasitic lights that produce in the system are by band pass filter 22 filterings.
Fig. 2 a and Fig. 2 b are respectively double-colored and the energy level synoptic diagram of three look CARS imaging.In the double-colored CARS imaging process shown in Fig. 2 a, two pump photons of the molecule absorption in the sample 20 and a Stokes photon are as difference and the resonant frequency Ω of testing molecule of frequency of the two
VibCoupling promptly satisfies Ω
Vib=ω
p-ω
sThe time, causing molecular oscillation and giving off frequency is ω
As=2 ω
p-ω
sThe CARS signal.And in three look CARS imaging processes shown in Fig. 2 b, pump photon of the molecule absorption in the sample 20, a Stokes photon and a probe photon, the sample CARS flashlight frequency that inspire this moment is ω
As=ω
p+ ω
Pr-ω
sWith the exciting light zero lap; Can come out CARS signal extraction by the second long-pass dichroic mirror 21 at the output terminal that optical fiber 16 receives signal, thereby realized using same optical fiber 16 transmission exciting light beams not receive the purpose of FWM signal interference with receiving sample CARS signal.
As embodiment; LASER Light Source 1 can adopt the picoTRAIN IC-1064-10000 type laser instrument of Austrian High-Q Laser company; It can provide the fundamental frequency pulse output beam of 76MHz pulsed frequency, 7ps pulse width, 1064nm wavelength and the double frequency pulse output beam of 532nm wavelength.The Levante Emerald psec product of the German APE GmbH of optical parametric oscillator 4 employings company; The pulsed light beam of 532nm wavelength flows to optical parametric oscillator 4 as pump light; Can obtain adjustable in 680～990nm wavelength coverage, pulse width is the output beam of 5ps, as the pump light of CARS imaging.The stokes light that the 1064nm wavelength pulse light beam of being exported by LASER Light Source 1 forms images as CARS.The first long-pass dichroic mirror 13, the second long-pass dichroic mirror 21 and band pass filter 22 adopt Q1020LPXR, 640DCXR and the HQ560/50m type product of U.S. Chroma Technology company respectively, and detector 24 adopts R3896 type photomultiplier (PMT) detector of Japanese Bin Song company.All the other devices all can be buied from market.CH
2Chemical bond is often used in the imaging of biosome, and its resonant frequency is 2845cm
-1, need this moment to be adjusted to 816.8nm (12243cm to pump light
-1), stokes light stuck-at-064nm (9398cm
-1), the FWM signal that produces in the optical fiber is that probe light is 662.8nm (15088cm
-1), then the CARS signal light wavelength of sample generation is 557.6nm (17933cm
-1).
The four-wave mixing signal that utilizes optical fiber to produce that Fig. 3 illustrates the present invention's proposition carries out the CARS method for imaging, may further comprise the steps:
Step S1: the stokes light of the basic frequency beam of LASER Light Source output as the CARS imaging, the lentor light frequency is designated as ω
sThe pump light of the light beam that produces behind the frequency multiplication light beam pump optical parametric oscillator of LASER Light Source output and export as the CARS imaging, the pumping light frequency is designated as ω
p
Step S2:, stokes light and pump light are spatially overlapped after passing through the first long-pass dichroic mirror through regulating the catoptron in the Stokes light path;
Step S3:, stokes light and pump light are overlapped in time after passing through the first long-pass dichroic mirror through regulating the precision displacement table in the Stokes light path;
Step S4: regulate the pump light wavelength of output through the temperature of regulating gain media in the optical parametric oscillator, make the difference on the frequency of pump light and stokes light and the resonant frequency Ω of testing molecule chemical bond
VibBe complementary, promptly satisfy: Ω
Vib=ω
p-ω
s
Step S5: the frequency that inspires when in optical fiber, transmitting the pump light that overlaps on time and the space and stokes light is ω
Fwm=2 ω
p-ω
sNon-phase matching four-wave mixing signal, as the probe light of CARS imaging, the probe light frequency is designated as ω
Pr
Step S6: after focusing on the sample through condenser lens pump light, stokes light and probe light, inspire the CARS flashlight that satisfies phase-matching condition, CARS signal light frequency is ω
As=ω
p+ ω
Pr-ω
s
Step S7: by the CARS flashlight of sample generation; And by the pump light of sample retroreflection or scattering, stokes light and probe light be focused that lens are collected and coupled into optical fibres in transmit; Fiber-optic output have only the CARS flashlight by the second long-pass dichroic mirror reflects to detector, the light signal that detector receives is converted into and transfers to computing machine behind the electric signal and handle;
Step S8: computer control two-dimensional scanner band optical fiber end resonance, makes focused beam on sample, carry out two-dimensional scan, thereby obtains the two-dimentional CARS imaging results of sample.
Above-mentioned embodiment is used for the present invention that explains, rather than limits the invention.In the protection domain of spirit of the present invention and claim, any modification and change to the present invention makes all fall into protection scope of the present invention.
Claims (7)
1. the four-wave mixing signal that utilizes optical fiber to produce carries out the system of CARS imaging; It is characterized in that: comprise LASER Light Source (1), first catoptron (2), second catoptron (3), optical parametric oscillator (4), the 3rd catoptron (5), the 4th catoptron (6), the 5th catoptron (7), the 6th catoptron (8), the 7th catoptron (9), the 8th catoptron (10), precision displacement table (11), the 9th catoptron (12), the first long-pass dichroic mirror (13), the tenth catoptron (14), coupling object lens (15), optical fiber (16), two-dimensional scanner (17), condenser lens (18), supporting sleeve (19), sample (20), the second long-pass dichroic mirror (21), band pass filter (22), imaging len (23), detector (24) and computing machine (25), wherein:
The frequency multiplication light beam that sends from the output port a of LASER Light Source (1) is after first catoptron (2) and second catoptron (3) reflection, and input and pump optical parametric oscillator (4) are the pump light of CARS imaging by the light beam of optical parametric oscillator (4) output; The basic frequency beam that sends from the output port b of LASER Light Source (1) is the stokes light of CARS imaging; Stokes light successively after the 3rd catoptron (5), the 4th catoptron (6), the 5th catoptron (7), the 6th catoptron (8), the 7th catoptron (9), the 8th catoptron (10) and the 9th catoptron (12) reflection to the first long-pass dichroic mirror (13), this light path is the Stokes light path;
The 6th catoptron (8) in the Stokes light path and the 7th catoptron (9) are installed on the precision displacement table (11); Through regulating precision displacement table (11) and the 8th catoptron (10) and the 9th catoptron (12), make respectively from the pump light of optical parametric oscillator (4) output with from the stokes light that the 9th catoptron (12) comes to overlap in time and space through the first long-pass dichroic mirror (13) back; Pump light that overlaps on time and the space and stokes light are coupled transmission in object lens (15) coupled into optical fibres (16) after the tenth catoptron (14) reflexes to the second long-pass dichroic mirror (21); When pump light that overlaps on time and the space and stokes light transmit, inspire the four-wave mixing signal of non-phase matching in optical fiber (16), the probe light of four-wave mixing signal as the CARS imaging; Being focused lens (18) from pump light, stokes light and the probe light of optical fiber (16) output focuses on sample (20) and goes up and inspire the CARS signal; The end of optical fiber (16) is fixed on the two-dimensional scanner (17), and makes optical fiber (16) stretch out a bit of semi-girder that becomes, and two-dimensional scanner (17) and condenser lens (18) are installed in the supporting sleeve (19);
By the CARS flashlight of sample (20) generation with by the pump light of sample (20) retroreflection or scattering, stokes light and probe light; Be focused that lens (18) are collected and coupled into optical fibres (16) in transmission, then behind the object lens (15) that are coupled to the second long-pass dichroic mirror (21); By the pump light of sample (20) retroreflection or scattering, stokes light and the probe light transmission second long-pass dichroic mirror (21), the CARS flashlight is reflected by the second long-pass dichroic mirror (21) after band pass filter (22) and imaging len (23) back are received by detector (24);
The two-dimensional scanner (17) that computing machine (25) control is installed in the supporting sleeve (19) is being with optical fiber end resonance; Make the pump light, stokes light and the probe light that are focused lens (18) focusing on sample (20), carry out two-dimensional scan; Computing machine (25) is control detector (24) synchronous acquisition CARS signal and transfer to computing machine (25) and handle simultaneously, thereby obtains the two-dimentional CARS imaging results of sample.
2. the four-wave mixing signal that utilizes optical fiber to produce according to claim 1 carries out the system of CARS imaging; It is characterized in that: described LASER Light Source (1) is high-frequency impulse near-infrared laser light source with 2～7ps or 102fs magnitude pulse width, the frequency multiplication light beam that said high-frequency impulse near-infrared laser light source has the basic frequency beam that sends from output port b, sends from output port a.
3. the four-wave mixing signal that utilizes optical fiber to produce according to claim 1 carries out the system of CARS imaging, it is characterized in that: described optical parametric oscillator (4) is that the pulse width with LASER Light Source (1) is complementary, wavelength is at the adjustable laser generator of near-infrared band.
4. the four-wave mixing signal that utilizes optical fiber to produce according to claim 1 carries out the system of CARS imaging, it is characterized in that: the direction of the light beam of described the 5th catoptron (7) to the 6th catoptron (8) and the 7th catoptron (9) to the light beam of the 8th catoptron (10) is all parallel with the displacement adjusting direction of precision displacement table (11).
5. the four-wave mixing signal that utilizes optical fiber to produce according to claim 1 carries out the system of CARS imaging, it is characterized in that: the input end end face of described optical fiber (16) is positioned on the front focal plane of coupling object lens (15).
6. the four-wave mixing signal that utilizes optical fiber to produce according to claim 1 carries out the system of CARS imaging, and it is characterized in that: the light-sensitive surface of described detector (24) is positioned on the front focal plane of imaging len (23).
7. one kind is used the four-wave mixing signal that utilizes optical fiber to produce of the said CARS imaging system of claim 1 to carry out the CARS method for imaging, may further comprise the steps:
Step S1: the stokes light of the basic frequency beam of LASER Light Source output as the CARS imaging, the lentor light frequency is designated as ω
sThe pump light of the light beam that produces behind the frequency multiplication light beam pump optical parametric oscillator of LASER Light Source output and export as the CARS imaging, the pumping light frequency is designated as ω
p
Step S2:, stokes light and pump light are spatially overlapped after passing through the first long-pass dichroic mirror through regulating the catoptron in the Stokes light path;
Step S3:, stokes light and pump light are overlapped in time after passing through the first long-pass dichroic mirror through regulating the precision displacement table in the Stokes light path;
Step S4: regulate the pump light wavelength of output through the temperature of regulating gain media in the optical parametric oscillator, make the difference on the frequency of pump light and stokes light and the resonant frequency Ω of testing molecule chemical bond
VibBe complementary, promptly satisfy: Ω
Vib=ω
p-ω
s
Step S5: the frequency that inspires when in optical fiber, transmitting the pump light that overlaps on time and the space and stokes light is ω
Fwm=2 ω
p-ω
sNon-phase matching four-wave mixing signal, as the probe light of CARS imaging, the probe light frequency is designated as ω
Pr
Step S6: after focusing on the sample through condenser lens pump light, stokes light and probe light, inspire the CARS flashlight that satisfies phase-matching condition, CARS signal light frequency is ω
As=ω
p+ ω
Pr-ω
s
Step S7: by the CARS flashlight of sample generation; And by the pump light of sample retroreflection or scattering, stokes light and probe light be focused that lens are collected and coupled into optical fibres in transmit; Fiber-optic output have only the CARS flashlight by the second long-pass dichroic mirror reflects to detector, the light signal that detector receives is converted into and transfers to computing machine behind the electric signal and handle;
Step S8: computer control two-dimensional scanner band optical fiber end resonance, makes focused beam on sample, carry out two-dimensional scan, thereby obtains the two-dimentional CARS imaging results of sample.
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