CN103439254B - One kind of confocal Raman spectroscopy method and apparatus for laser spectroscopic pupil - Google Patents

One kind of confocal Raman spectroscopy method and apparatus for laser spectroscopic pupil Download PDF

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CN103439254B
CN103439254B CN201310404307.3A CN201310404307A CN103439254B CN 103439254 B CN103439254 B CN 103439254B CN 201310404307 A CN201310404307 A CN 201310404307A CN 103439254 B CN103439254 B CN 103439254B
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pupil
system
spectral
detection
confocal
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CN103439254A (en
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王允
赵维谦
刘大礼
盛忠
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北京理工大学
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Abstract

本发明属于显微光谱成像技术领域,将共焦显微技术与拉曼光谱探测技术相结合,涉及一种分光瞳激光共焦拉曼光谱测试方法及装置。 The present invention belongs to the technical field microspectroscopy imaging, confocal microscopy and Raman spectroscopy detection technology, relates to a laser confocal Raman spectroscopic pupil spectroscopy method and apparatus. 利用共焦拉曼光谱探测中遗弃的瑞利散射光构建分光瞳共焦显微成像系统,实现样品三维几何位置的高分辨成像与探测;利用分光瞳共焦显微成像系统的“极值点”来控制光谱探测系统能精确捕获物镜聚焦点处激发的拉曼光谱信息,继而实现“图谱合一”的分光瞳共焦拉曼光谱高空间分辨成像与探测。 Construction of high resolution optical spectral imaging and detection pupil confocal microscopy imaging system, the position of the sample using three-dimensional geometric abandoned spectral detection of Raman confocal Rayleigh scattering; pupil confocal microscopy using spectroscopic imaging system "extreme point" to control the spectral detection system to accurately capture the information of the Raman spectrum at the excitation focal point of the objective lens, then carrying out the "Atlas one" spectral confocal Raman spectroscopy pupil high spatial resolution imaging and detection. 本发明为微区三维几何位置与光谱的高空间分辨探测提供新的技术途径,可广泛应用于物理、化学、生物医学、材料科学、环境科学、石油化工、地质、药物、食品、刑侦和珠宝检定等领域,可对样品进行无损伤鉴定和深度光谱分析。 The present invention is a three-dimensional geometric positions spectral domains provide high spatial resolution to detect new way, can be widely used in physical, chemical, biomedical, materials science, environmental science, petrochemical, geology, pharmaceutical, food, forensic and jewelry test fields, the sample can be identified and no damage depth spectroscopy.

Description

一种分光瞳激光共焦拉曼光谱测试方法与装置 One kind of confocal Raman spectroscopy method and apparatus for laser spectroscopic pupil

技术领域 FIELD

[0001] 本发明属于显微光谱成像技术领域,将共焦显微技术与拉曼光谱探测技术相结合,涉及一种“图谱合一”的高空间分辨光谱测试成像方法及装置,用于对样品的微区光谱进行高分辨测试等。 [0001] The present invention belongs to the technical field microspectroscopy imaging, confocal microscopy and Raman spectroscopy detection technology, relates to a "map one" high spatial resolution imaging spectroscopy method and apparatus for sample High resolution spectra were micro testing.

背景技术 Background technique

[0002] 激光共焦拉曼光谱技术通过入射激光引起分子(或晶格)产生振动而损失(或获得)部分能量,使散射光频率发生变化,通过对散射光进行分析,来探知分子的组分、结构及相对含量等,激光共焦拉曼光谱技术亦被称为分子探针技术。 [0002] Confocal Raman spectroscopy molecules caused by the incident laser (or lattice) vibration loss (or gain) portion of the energy, the frequency of scattered light is changed by light scattering analysis to ascertain the group of molecules points, and the relative content of other structures, confocal laser Raman spectroscopy technique also known as molecular probe. 该技术既继承了共焦显微术的高分辨层析成像特征,又可以对样品进行光谱分析,它不仅可以观测样品同一层面内不同微区的拉曼光谱信号,还能分别观测样品内深度不同的各个层面的拉曼信号,对被测样品进行断层扫描,从而在不损伤样品的情况下达到进行“光学切片”的效果。 This technique not only inherited the high-resolution tomography characterized in confocal microscopy, and samples can be spectroscopy, Raman spectra can be observed not only signal the different domains of the same level of the sample, observations were also different depths within the sample Raman signal levels of each of the samples tested for tomography, so that the sample without damaging the effect achieved for "optical sections" of. 激光共焦拉曼光谱测试技术作为一种极其重要的材料结构测量与分析的基本技术手段,广泛应用于物理、化学、生物医学、材料科学、环境科学、石油化工、地质、药物、食品、刑侦和珠宝检定等领域,可对样品进行无损伤鉴定和深度光谱分析,同时,还可以进行样品扫描和低温分析、材料的光致发光研宄等。 Confocal Raman spectroscopy technology as a very important basic material structure measurement techniques and analysis, widely used in physics, chemistry, biomedicine, materials science, environmental science, petrochemical, geological, pharmaceutical, food, criminal investigation and jewelery fields assay, samples can be identified and no damage to the depth of spectral analysis, can also be scanned and low sample analysis, a Subsidiary photoluminescent material and the like.

[0003] 传统共焦拉曼光谱测试仪的原理如图1所示,光源系统发出激发光束透过偏振分光镜、四分之一波片和聚焦物镜后,聚焦在被测样品上,激发出载有样品光谱特性的拉曼散射光;通过三维扫描系统移动被测样品,使对应被测样品不同区域的拉曼散射光再次通过四分之一波片并被偏振分光镜反射,第一聚光镜将偏振分光棱镜反射的光进行会聚,利用位于第一针孔后面的拉曼光谱探测系统测得载有被测样品光谱信息的拉曼散射光谱。 [0003] The principle of confocal Raman spectroscopy conventional tester shown in Figure 1, a light source system emits excitation beam passes through a polarization beam splitter, the quarter-wave plate and the focusing objective is focused on the sample under test, stimulate contains a sample spectrum characteristic Raman scattered light; test sample moves through the three-dimensional scanning system, different regions of the corresponding test sample Raman scattered light through the quarter-wave plate and the reflective polarization beam splitter, a first condenser lens again the light reflected by the PBS converging Raman spectroscopy using a detection system located behind the first pinhole measured Raman scattering spectrum containing spectral information the test sample.

[0004] 由于激发出的拉曼散射光十分微弱,现有激光共焦拉曼光谱探测仪为了减小拉曼散射光的能量损失,系统中选取较大的针孔,通常在Φ150μπι〜Φ 200 μm之间,由于系统利用共焦方式进行焦点定位,针孔尺寸直接影响共焦轴向定位曲线的半高宽,针孔尺寸较大导致系统定焦精度降低,即降低了空间分辨力。 [0004] Since the excitation of the Raman scattered light is very weak, the conventional confocal laser Raman spectroscopy detector in order to reduce energy loss of the Raman scattered light, the larger the system selected pinholes typically Φ150μπι~Φ 200 between [mu] m, since the system utilizes a confocal manner for positioning the focus, the pinhole size directly affects the axial positioning of the confocal curve FWHM, the larger the size of the pinhole resulting in reduced accuracy of fixed-focus system, i.e., reducing the spatial resolution. 并且系统只能进行光谱探测,模式单一,限制了其应用领域。 Spectrum detection system and only a single mode, which limits its field of application.

[0005]同时,样品散射的拉曼散射光只有反射的瑞利光束强度的10_3〜10 _6倍,而现有共焦拉曼光谱探测仪器只利用了微弱的拉曼散射光进行光谱探测而遗弃强于拉曼散射光13〜10 6倍的瑞利光束,利用拉曼散射光直接进行成像,存在系统灵敏度较低的不足。 [0005] Meanwhile, _6 10_3~10 sample times the Rayleigh scattering of the beam intensity of Raman scattered light is only reflected, whereas conventional confocal Raman spectroscopy detection apparatus uses only a weak Raman scattered light spectrum detection and abandonment Raman scattered light intensity at 13~10 6 times the Rayleigh light, Raman scattering light using direct imaging, the presence of low system sensitivity insufficient.

[0006] 此外,为了获得精确、丰富的测量信息,拉曼光谱成像时既需较长时间的单点拉曼光谱探测,又需进行多点拉曼光谱探测,因此拉曼光谱成像需要较长的时间。 [0006] Further, in order to obtain accurate measurement information rich, Raman spectroscopy requires both imaging longer a single point of detection of the Raman spectrum, and the need for multi-probe Raman spectroscopy, Raman spectroscopy thus requires a longer imaging time. 但是,仪器长时间成像过程中受环境温度、振动、空气抖动等的影响较大,易使仪器系统产生漂移,从而导致样品被探测位置离焦;由于现有共焦拉曼光谱探测技术不具备实时焦点跟踪和位置矫正能力,因而在整个成像过程中,无法保证其激发光斑的位置处在物镜焦点位置,实际激发光斑远大于物镜聚焦光焦斑,其结果制约了可探测区域的微小化,限制了共焦拉曼光谱仪器的微区光谱探测能力。 However, the image forming apparatus during a greater impact for a long time by the environmental temperature, vibration, shake or the like of air, the system easy to make the instrument drift, causing the sample to detect the defocus position; confocal Raman spectroscopy because conventional detection techniques do not have real-time tracking and focus position correction capability, thus the entire imaging process, can not guarantee a position in which the excitation spot of the objective lens focal position of the objective lens is much larger than the actual spot of the excitation light focused focal spot, as a result of miniaturization detectable restricted area, It limits the ability to detect the spectral domains of confocal Raman spectrometer.

发明内容 SUMMARY

[0007] 本发明的目的是为克服已有技术的不足,提供一种共焦显微技术与拉曼光谱探测相结合的分光瞳激光共焦拉曼光谱测试方法及装置。 [0007] The object of the present invention is to overcome the disadvantages of prior art, provide one kind of spectral pupil laser confocal microscopy and Raman spectroscopy to detect the combination of confocal Raman spectroscopy method and apparatus. 利用共焦拉曼光谱探测中遗弃的瑞利散射光构建分光瞳共焦显微成像系统,实现样品三维几何位置的高分辨成像与探测;利用分光瞳共焦显微成像系统的“极值点”来控制光谱探测系统能精确捕获物镜聚焦点处激发的拉曼光谱信息,继而实现“图谱合一”的分光瞳共焦拉曼光谱高空间分辨成像与探测。 Construction of high resolution optical spectral imaging and detection pupil confocal microscopy imaging system, the position of the sample using three-dimensional geometric abandoned spectral detection of Raman confocal Rayleigh scattering; pupil confocal microscopy using spectroscopic imaging system "extreme point" to control the spectral detection system to accurately capture the information of the Raman spectrum at the excitation focal point of the objective lens, then carrying out the "Atlas one" spectral confocal Raman spectroscopy pupil high spatial resolution imaging and detection.

[0008] 本发明的目的是通过下述技术方案实现的。 [0008] The object of the present invention is achieved by the following technical solutions.

[0009] 一种分光瞳激光共焦拉曼光谱测试方法,包括以下步骤: [0009] A laser confocal Raman spectroscopic pupil spectroscopy method, comprising the steps of:

[0010] 在测量物镜的光瞳面上放置照明光瞳和收集光瞳。 [0010] In the measurement of the objective lens pupil plane of the illumination pupil and the collection placed pupil. 光源系统发出能够激发出拉曼光谱的激发光束。 The light source system emits an excitation beam can excite the Raman spectrum. 激发光束透过照明光瞳、测量物镜后,聚焦在被测样品上,激发出载有被测样品光谱特性的拉曼散射光,同时反射出瑞利光。 Excitation beam passes through the illumination pupil, measured after the objective lens is focused on the sample under test, the test sample containing the excitation spectral characteristics of Raman scattered light, Rayleigh light while reflecting. 拉曼散射光和瑞利光经测量物镜和收集光瞳后到达二向色分光系统,二向色分光系统对拉曼散射光和瑞利光进行无损分离。 Raman scattered light and the Rayleigh light collected by the objective lens and the pupil measurement reaches dichroic spectroscopy system, the Raman scattered light and the Rayleigh light two pairs of non-destructive separation dichroic spectroscopy system.

[0011] 经二向色分光系统反射的瑞利光进入分光瞳激光共焦探测系统。 [0011] by the two reflected by the dichroic beam splitter splitting system Rayleigh light enters the pupil confocal laser detection system. 分光瞳激光共焦探测系统利用探测器横向偏移可使分光瞳共焦显微系统的响应特性曲线产生相移的特性,实现对被测样品微区几何位置的探测。 Confocal spectral pupil detection system can utilize laterally offset detector spectral response of the pupil confocal microscopy system generated characteristic curve of phase shift, to achieve detection of the geometrical position of the micro test sample. 具体过程为:对接收瑞利光的分光瞳激光共焦探测系统获得的探测光斑进行处理,得到探测区域,测得反映样品凹凸变化的分光瞳激光共焦响应I (U),即可进行高空间分辨的三维尺度层析成像探测,其中,u为轴向归一化光学坐标。 The specific process is: the spectral pupil Rayleigh light received laser confocal detection spot detection system available, to give the detection region, the sample was measured to reflect changes in spectral irregularities pupil laser confocal response I (U), to high spatial resolved detection tomography three dimensions, wherein, u is a normalized optical axial coordinate.

[0012] 与此同时,经二向色分光系统透射的拉曼散射光进入拉曼光谱探测系统,利用分光瞳激光共焦响应曲线的“极值点”与测量物镜焦点位置精确对应这一特性,将分光瞳激光共焦响应曲线的“极值点”精确对应测量物镜(5)的焦点,通过寻找“极值点”来精确捕获激发光斑焦点位置的光谱信息,实现高空间分辨的光谱探测。 [0012] Meanwhile, the two spectral color transmission system to enter the Raman spectroscopy Raman scattered light detection system, using a confocal laser pupil spectral response curve of the "extreme point" and the focal position of the objective lens correspond precisely measuring the characteristics , the spectral response confocal laser pupil "extreme point" curve corresponds precisely focus measurement objective (5) and searching the "extreme point" capture spectral information to accurately position the focal spot of the excitation, to achieve high spatial resolution of the spectral detection . 测量过程中可以对被测样品进行实时、精确地跟踪定焦,通过计算机处理系统控制三维扫描系统的姿态以保证被测样品在整个测量过程中始终处于焦点位置,可抑制因环境温度和振动等因素导致被测样品离焦而产生的误差,提高测量精度。 Measurement process may be performed in real-time, accurately track focusing on a test sample, the attitude control system of three-dimensional scanning by the computer processing system to ensure that the test sample remains in focus throughout the measurement process, due to environmental temperature can be suppressed, and vibration factors leading to defocus errors generated test sample to improve measurement accuracy.

[0013] 当只对接收瑞利光的分光瞳激光共焦探测系统获得的探测光斑进行处理时,能够进行高空间分辨率的三维尺度层析成像探测; [0013] When the detection spot of the laser receiver only pupil Rayleigh light spectral confocal detection system was subjected to treatment, can be performed in three dimensions with high spatial resolution tomography detection;

[0014] 当只对接收拉曼散射光的拉曼光谱探测系统获得的光谱信号进行处理时,能够进行高空间分辨的光谱探测; [0014] When the spectra of the Raman spectrum signal detection system only receives Raman scattered light obtained processing, spectrum detection can be performed in high spatial resolution;

[0015]当同时对接收瑞利光的分光瞳激光共焦探测系统获得的探测光斑和接收拉曼散射光的拉曼光谱探测系统获得的光谱信号进行处理时,能够进行高空间分辨的微区图谱层析成像,实现对被测样品“图谱合一”的分光瞳激光共焦拉曼光谱高空间分辨成像与探测。 [0015] When the spectral flare while the spectral signal detection pupil confocal laser Rayleigh light receiving detection system is obtained and the received Raman scattered light detection system of the Raman spectrum obtained for processing, can be micro pattern of high spatial resolution tomography, to achieve the "Atlas one" test sample spectral pupil laser confocal Raman spectroscopy high spatial resolution imaging and detection.

[0016] 特别的,在本发明方法中,所述照明光瞳和收集光瞳可以是圆形、D形或者其他形状。 [0016] In particular, in the process of the present invention, the illumination pupil and the collection pupil can be circular, D-shaped, or other shapes.

[0017] 在本发明方法中,激发光束可以是线偏振光束、圆偏振光束等偏振光束;还可以是由光瞳滤波技术生成的结构光束。 [0017] In the method of the present invention, the excitation beam may be linearly polarized beam, the polarization beam circularly polarized beam and the like; may also be generated by the beam structure pupil filtering techniques. 偏振光束与光瞳滤波技术结合可以压缩测量聚焦光斑尺寸,提高分光瞳激光共焦探测系统的横向分辨力。 Polarized light beam to the pupil filtering technologies focus measurement spot size may be compressed to improve the spectral confocal laser pupil lateral resolution of the detection system.

[0018] 本发明方法可以探测包括荧光、布里渊散射光、康普顿散射光等散射光谱。 [0018] The method of the present invention may comprise detecting fluorescence, Brillouin scattered light, scattered light, etc. Compton scattering spectrum.

[0019] 在本发明方法中,仅通过计算机系统软件处理即可实现对不同NA值的测量物镜的匹配,而无需重新对系统进行任何硬件装调。 [0019] In the method of the present invention, only the matching processing can be realized with different values ​​of NA of the objective lens is measured by the computer system software without any hardware re-assembly and adjustment system.

[0020] 本发明提供了一种分光瞳激光共焦拉曼光谱测试装置,包括产生激发光束的光源系统、测量物镜、照明光瞳、收集光瞳、三维扫描系统、位移传感器、二向色分光系统、拉曼光谱探测系统、分光瞳激光共焦测量系统、数据处理单元和计算机处理系统。 [0020] The present invention provides a confocal Raman spectroscopy pupil laser spectroscopic apparatus, comprising a light source for producing the excitation beam system, measurement objective, illumination pupil, the pupil collect, three-dimensional scanning system, a displacement sensor, a dichroic beam splitter system, detection system, Raman spectroscopy, confocal laser pupil spectroscopic measurement system, data processing unit and a computer processing system.

[0021] 其中,照明光瞳和收集光瞳放置在测量物镜的光瞳面上。 [0021] wherein the illumination pupil and the collection pupil placed at the pupil plane of the objective lens is measured. 被测样品固定在三维扫描装置的载物台上。 The test sample was fixed on a stage of the three-dimensional scanning apparatus. 照明光瞳和测量物镜依次放置在光源系统的光束出射方向上,照明光瞳与激发光束同轴,二向色分光系统放置在收集光瞳之后,拉曼光谱探测系统放置在二向色分光系统的透射方向上,分光瞳激光共焦探测系统在二向色分光装置的反射方向上,数据处理单元用于对拉曼光谱探测系统、分光瞳激光共焦探测系统和位移传感器采集到的数据进行融合处理。 Illumination pupil of the objective lens and measuring sequentially placed on the exit direction of the beam of the light source system, and the excitation light illumination pupil coaxially disposed two dichroic spectroscopy system pupil after collection, the Raman spectrum detection system is placed dichroic spectroscopy system in the direction of transmission, spectral confocal laser detection system pupil in the reflecting direction of the dichroic beam splitter means, a data processing unit for detection of the Raman spectroscopy system, the spectral data detection system pupil confocal laser displacement sensor and to focus collected for the integration process. 被测样品固定在三维扫描系统的载物台上。 The test sample was fixed on a stage of the three-dimensional scanning system. 计算机处理系统用于对三维扫描系统、拉曼光谱探测系统、分光瞳激光共焦探测系统以及数据处理单元进行协调和控制。 The computer processing system for three-dimensional scanning system, Raman spectroscopy detection system pupil spectral confocal laser detection system and a data processing unit for coordinating and controlling.

[0022] 在本发明装置中,数据处理单元包括分割焦斑探测模块、极大值运算模块,用于处理图像采集系统得到的探测光斑,得到分光瞳激光共焦响应曲线,从而得到被测样品的位置信息;包括数据融合模块,用于融合位置信息Ku)和光谱信息I(r),完成样品三维重构及光谱信息融合I (X,y, Z, r)。 [0022] In the apparatus of the present invention, the data processing unit includes dividing the focal spot detection module, the maximum value calculation means for processing the image acquisition system to detect spots obtained, to obtain the spectral response curve confocal laser pupil, thereby obtaining a test sample position information; includes a data fusion module for fusing the position information Ku) and the spectral information I (r), and complete three-dimensional reconstructed samples fusion spectral information I (X, y, Z, r).

[0023] 在本发明装置中,拉曼光谱探测系统可以是普通拉曼光谱探测系统,包括沿光路依次放置的第一聚光镜、位于第一聚光镜焦点位置的光谱探测器,以及位于光谱探测器之后的探测器,用于被测样品的表层拉曼光谱探测。 [0023] In the apparatus of the present invention, the Raman spectrum detection system may be an ordinary Raman spectrum detection system are disposed in order along the optical path including a first condenser, a first condenser lens spectrum detector located in the focal position, and after the probe of the spectrum detectors, surface Raman spectrum for a test sample probe. 还可以是共焦拉曼光谱探测系统,包括沿光路依次放置的第一聚光镜、位于第一聚光镜焦点位置的第一针孔、位于第一针孔后的光谱探测器,以及位于光谱探测器之后的探测器,可以有效滤除焦点之外的杂散光,提高拉曼光谱探测的信噪比。 May also be a confocal Raman spectroscopy detection system are disposed in order along the optical path including a first condenser, a first pinhole located at a first focal position of the condenser lens, the spectral detector positioned after the first pinhole, and the detectors of the spectrum after detector, stray light can be effectively filtered out of focus, to improve the Raman spectral detection SNR.

[0024] 在本发明装置中,可通过在光源系统和照明光瞳之间增加光束调制系统,包括沿光路依次放置的第三聚光镜、位于第三聚光镜焦点处的第二针孔、以及焦点位于第二针孔处的第四聚光镜,用于对光源系统发出的激发光束进行调制,以得到更高空间分辨力和更好的激发效果。 [0024] In the apparatus of the present invention may be increased by a beam modulation system between the light source and the illumination pupil system, comprising sequentially positioned along the optical path of the third condenser lens, a second pinhole located at the focal point of the third condenser lens, and a focus located a fourth condenser lens second pinhole for the excitation light beam emitted from the light source system is modulated in order to obtain a higher spatial resolution and better excitation effect.

[0025] 在本发明装置中,可通过增加图像放大系统,放大图像采集系统探测到的艾利斑,包括沿光路依次放置的第二聚光镜、与第二聚光镜共焦点的图像放大系统以及位于图像放大系统焦点处的图像采集系统,以提高分光瞳激光共焦测量装置的采集精度。 [0025] In the apparatus of the present invention, it can be detected by increasing the image magnification system, an enlarged image acquisition system Airy, including along the optical path are sequentially disposed a second condenser, a second condenser lens confocal image magnification system and of the image an enlarged image acquisition system at the focus system, in order to improve the spectral acquisition accuracy pupil laser confocal measurement device.

[0026] 有益效果 [0026] Advantageous Effects

[0027] 本发明方法对比已有技术具有以下创新点: [0027] comparative prior art method of the present invention has the following innovations:

[0028] 1、本发明将分光瞳激光共焦显微技术与拉曼光谱探测技术有机结合,融合了分光瞳激光共焦显微技术的高精度物镜聚焦点位置跟踪捕获能力,可使共焦拉曼光谱显微镜探测到精确对应最小激发聚焦光斑区域的样品光谱特性,大幅提高了现有共焦拉曼光谱显微镜的微区拉曼光谱探测能力,这是区别于现有拉曼光谱探测技术的创新点之一; [0028] 1, the present invention is the spectroscopic pupil confocal laser microscopy and Raman spectroscopy combined organic detection technology, integration of spectral confocal laser microscopy pupil of the objective lens focal point position precision tracking capturing ability, can confocal microscope Raman spectroscopy detected accurately correspond to the minimum focused spot excitation spectral characteristics of the sample area, a substantial increase in the ability to detect the Raman spectrum of conventional micro confocal Raman spectroscopy microscope, which is different from the one of the innovations of the conventional Raman spectroscopy detection technology ;

[0029] 2、利用二向色分光装置对瑞利光和拉曼散射光进行无损分离,瑞利光进入分光瞳激光共焦探测系统,拉曼散射光进入拉曼光谱探测系统,提高了系统拉曼光谱探测灵敏度。 [0029] 2, using two pairs of the Raman scattered light and the Rayleigh light nondestructive separation dichroic beam splitter means, Rayleigh light entering the spectroscopic detection system pupil confocal laser, Raman spectroscopy Raman scattered light enters the detection system, increase the Raman system spectral detection sensitivity. 这是区别于现有拉曼光谱探测技术的创新点之二; This innovation is different from the prior art two-probe Raman spectroscopy;

[0030] 3、由于采用分割焦斑的方法来获取信号,可通过改变在图像探测系统探测焦面上所设置的微小区域的参数以匹配不同的样品的反射率,从而可以扩展其应用领域;还可以仅通过计算机系统软件处理即可实现对不同NA值的测量物镜的匹配,而无需重新对系统进行任何硬件装调,有利于实现仪器的通用性。 [0030] 3, since the method of dividing the focal spot to acquire the signal, may be different to match the reflectance of the sample by changing the parameters of the microscopic region disposed in the focal plane of the detection system detects that the image, which can extend its field of application; can also be treated only by a computer software system can be realized matching values ​​measured for different NA of the objective lens, without any hardware re-assembly and adjustment system, it is conducive to the versatility of the instrument. 这是区别于现有拉曼光谱探测技术的创新占之二.V、、、----, This is different from the existing Raman spectroscopy detection technology and innovative accounting of the two .V ,,, ----,

[0031] 4、本发明仅通过软件切换处理便可实现分光瞳激光共焦显微探测、激光共焦拉曼光谱探测、分光瞳激光共焦拉曼光谱探测多模式切换与处理。 [0031] 4, only the present invention can be realized by software switching process Confocal Microscopic spectral pupil detection, the detection laser confocal Raman spectroscopy, confocal laser Raman spectroscopic pupil multi-spectral detection and mode switching process. 这是区别于现有拉曼光谱探测技术的创新点之四。 This is the innovation of the existing Raman spectroscopy detection technique is different from four.

[0032] 本发明方法具有如下特点: [0032] The method of the present invention has the following characteristics:

[0033] 1、融合分光瞳激光共焦探测技术和拉曼光谱探测技术,利用分光瞳激光共焦系统对焦点的精确定位,进行焦点跟踪测量,大幅提高拉曼光谱探测的空间分辨力; [0033] 1, fused pupil spectroscopic detection techniques and confocal laser Raman spectroscopy detection technique, using a confocal laser spectroscopic pupil precise positioning system focus, the focus tracking measurement, a substantial increase in the spatial resolution of the Raman spectroscopy detection;

[0034] 2、可实现量程范围与分辨能力的有效兼顾,同时通过设置在焦斑上所取微小区域的参数,以匹配不同反射率的被测样品,应用范围得到扩展; [0034] 2, can be effective both measuring range and the resolving power, while by setting the parameters in the region of the focal spot taken minute, to match the different reflectivities of the test sample, expanded range of applications;

[0035] 3、系统具有分光瞳激光共焦显微探测、激光共焦拉曼光谱探测和分光瞳激光共焦拉曼光谱探测三种工作模式,仅需通过软件操作便可实现三种模式的切换; [0035] 3, the spectroscopic system having a pupil detection laser confocal microscopy, laser confocal Raman spectroscopy and the spectroscopic detection pupil confocal laser Raman spectroscopy to detect three operating modes, switching the three modes can be realized only by software operation;

[0036] 4、二向色分光系统的使用增强拉曼光谱探测系统所接收到的拉曼光谱,提高拉曼光谱探测的信噪比,使共焦拉曼光谱显微镜的微区光谱探测能力显著提高,也可以降低系统对激发光源的光强要求。 [0036] 4, two dichroic spectroscopy system using enhanced Raman spectroscopy Raman spectra of the received detection systems, improved signal to noise ratio of the Raman spectrum detection, so that the spectra of confocal Raman microscope probe microanalysis capability significantly increase, the system can also reduce the intensity of the excitation light source is required.

附图说明 BRIEF DESCRIPTION

[0037] 图1为共焦拉曼光谱成像方法示意图; [0037] Figure 1 is a schematic view of a confocal Raman spectroscopic imaging method;

[0038] 图2为本发明所述分光瞳激光共焦拉曼光谱测试方法示意图; [0038] Fig 2 a schematic view of a total power of the Raman spectroscopic method spectra pupil laser of the present invention;

[0039] 图3为D形分光瞳激光共焦拉曼光瞳测试示意图; [0039] FIG. 3 is a D-shaped laser confocal Raman spectroscopic pupil test schematic pupil;

[0040] 图4为本发明所述分光瞳激光共焦拉曼光谱测试装置示意图; [0040] Fig 4 a schematic view of the spectroscopic pupil laser power Raman spectroscopy apparatus of the present invention were;

[0041] 图5为具有共焦光谱探测系统的分光瞳激光共焦拉曼光谱测试装置示意图; [0041] FIG. 5 is a schematic diagram of a confocal Raman spectroscopy device having a spectral confocal laser spectroscopy pupil detection system;

[0042] 图6为具有光束调制系统的分光瞳激光共焦拉曼光谱测试装置示意图; [0042] FIG. 6 is a schematic view of a confocal Raman spectroscopy apparatus having the laser beam spectroscopic pupil modulation systems;

[0043] 图7为具有探测焦斑放大系统的分光瞳激光共焦拉曼光谱测试装置示意图; [0043] FIG. 7 is a schematic view of a confocal Raman spectroscopy device having a spectral detection laser focal spot pupil amplifier system;

[0044] 图8为分光瞳激光共焦拉曼光谱测试方法与装置实施例示意图; [0044] FIG. 8 is a Raman spectroscopic pupil confocal laser spectroscopy method and apparatus schematic diagram of embodiment;

[0045] 图9为分光瞳激光共焦拉曼光谱测试方法的分光瞳激光共焦响应曲线及拉曼光谱响应曲线示意图。 [0045] FIG. 9 is a confocal laser spectroscopic spectral pupil pupil laser Raman spectra method confocal focus response curve and the Raman spectrum schematic response curve.

[0046] 其中,1-光源系统、2-测量物镜、3-照明光瞳、4-收集光瞳、5-被测样品,6- 二向色分光系统、7-拉曼光谱探测系统、8-第一聚光镜、9-光谱探测器、10-探测器、11-分光瞳激光共焦探测系统、12-第二聚光镜、13-图像采集系统、14-探测器区域、15-三维扫描系统、16-位移传感器、17-数据处理单元、18-分割焦斑探测模块、19-极值运算模块、20-数据融合模块、21-计算机处理系统、22-第一针孔、23-光束调制系统、24-第三聚光镜、25-第二针孔、26-第四聚光镜、27-图像放大系统、28-分光瞳激光共焦响应曲线、29-拉曼光谱响应曲线、30-偏振分光镜、31-四分之一波片、32-聚焦物镜。 [0046] wherein, l-source system, measurement objective 2-, 3- illumination pupil, the pupil collected 4-, 5- test sample, 6 dichroic spectroscopy system, the Raman spectroscopy detection system 7-, 8 - a first condenser lens, 9- spectral detector, 10 detector, 11-pupil spectral confocal laser detection system, 12 a second condenser lens, 13 an image acquisition system, 14-detector region, a three-dimensional scanning system 15, 16- displacement sensor, 17 a data processing unit 18 dividing the focal spot detection module, 19 computing module extrema, data fusion module 20, a computer processing system 21, a first pinhole 22-, 23- beam modulation system 24- third condenser lens, a second pinhole 25, a fourth condenser lens 26-, 27- image magnification system, 28- pupil confocal spectral response curve, the Raman spectral response curves 29-, 30- polarization beam splitter, 31- quarter-wave plate, 32- focusing objective.

具体实施方式 Detailed ways

[0047] 下面结合附图和实施例对本发明作进一步详细说明。 Drawings and embodiments of the present invention will be further described in detail [0047] below in conjunction.

[0048] 一种分光瞳激光共焦拉曼光谱测试方法,包括以下步骤: [0048] A laser confocal Raman spectroscopic pupil spectroscopy method, comprising the steps of:

[0049] 如图2所不,照明光瞳3和收集光瞳4放置在测量物镜2的光瞳面上。 [0049] FIG. 2 is not, the illumination pupil and the collection pupil 4 3 placed on the pupil plane of the objective lens 2 is measured. 光源系统I发出激发光束,激发光束透过照明光瞳3、测量物镜2后,聚焦在被测样品5上,激发出载有被测样品5光谱特性的拉曼散射光,同时反射出瑞利光。 The light source emits excitation light system I, the excitation beam is transmitted through the illumination pupil 3, measured after the objective lens 2, is focused on the test specimen 5, containing the test sample excited 5 spectral characteristic Raman scattered light, Rayleigh light while reflecting the . 拉曼散射光和瑞利光经测量物镜2和收集光瞳4后到达二向色分光系统6。 Raman scattered light and the Rayleigh light was measured and the objective lens 2 reaches the collection pupil dichroic spectroscopy system 6 4. 二向色分光系统6对拉曼散射光和瑞利光进行无损分离。 = 6 Raman scattered light and Rayleigh light nondestructive separation dichroic spectroscopy system. 经二向色分光系统6反射的瑞利光进入分光瞳激光共焦探测系统11,分光瞳激光共焦探测系统11利用探测器横向偏移可使分光瞳共焦显微系统的轴向响应特性曲线产生相移的特性,实现对被测样品5微区几何位置的探测。 By two reflected by the dichroic beam splitter 6 Rayleigh light entering the spectroscopic system pupil laser confocal detection system 11, the spectroscopic detection system pupil confocal laser probe 11 laterally offset can use axial confocal microscopy system pupil spectral response curve generated with shifting features that enable detection of the geometrical position of the test sample 5 domains. 经二向色分光系统6透射的拉曼散射光进入拉曼光谱探测系统7,利用分光瞳激光共焦响应曲线28的“极值点”与焦点位置精确对应这一特性,通过寻找“极值点”来精确捕获激发光斑焦点位置的光谱信息,实现高空间分辨的拉曼光谱探测。 Transmitted through two dichroic spectroscopy system 6 Raman spectroscopy Raman scattered light enters the detection system 7, by a spectral confocal laser pupil response "extreme point" of the curve 28 corresponds exactly with the focal position of this feature, by finding the "extrema point "capture spectral information to accurately position the focal spot of the excitation, Raman spectrum achieve high spatial resolution of the probe.

[0050] 一种分光瞳激光共焦拉曼光谱测试装置,如图4所示,包括光源系统1、测量物镜2、照明光瞳3、收集光瞳4、二向色分光系统6、拉曼光谱探测系统7、分光瞳激光共焦探测系统11、位移传感器16、数据处理单元17、三维扫描系统15和计算机处理系统21。 [0050] A laser confocal Raman spectroscopic spectrum pupil test device shown in Figure 4, the system includes a light source 1, measurement objective 2, 3 illumination pupil, the pupil collected 4, 6 two dichroic spectroscopy system, the Raman spectrum detection system 7, pupil spectral confocal laser detection system 11, the displacement sensor 16, the data processing unit 17, a three-dimensional scanning system 15 and the computer processing system 21.

[0051] 其中,照明光瞳3和收集光瞳4放置在测量物镜2的光瞳面上。 [0051] wherein the illumination pupil and the collection pupil 4 3 placed on the pupil plane of the objective lens 2 is measured. 照明光瞳3和测量物镜2依次位于光源系统I的激发光束出射方向上,照明光瞳3与激发光束同轴;二向色分光系统6位于收集光瞳4之后。 3 on the illumination pupil of the objective lens 2 and the measuring system I is successively positioned light source excitation beam exit direction, illumination pupil 3 and the excitation light beam coaxial; two pupil of the collector 6 after the color splitting system 4.

[0052] 拉曼光谱探测系统7位于二向色分光系统6的透射方向上;拉曼光谱探测系统7包括第一聚光镜8、光谱探测器9和探测器10。 [0052] Raman spectroscopy detection system 7 located on the two transmission directions dichroic beam splitter system 6; 7 Raman spectroscopy detection system comprising a first converging mirror 8, 9 and the detector 10 the spectral detector. 其中,光谱探测器9的探测面位于第一聚光镜8的焦点处,探测器10位于光谱探测器9之后。 Wherein the detection surface spectral detector 9 is positioned at a first focal condenser lens 8, the detector 10 is located after 9 spectroscopy probe. 探测器10将得到的光谱信号发送给数据处理单元17。 The resulting spectroscopic signal detector 10 is sent to the data processing unit 17.

[0053] 分光瞳激光共焦探测系统11位于二向分光系统6的反射方向上;分光瞳激光共焦探测系统11包括第二聚光镜12和图像采集系统13,其中,图像采集系统13的探测面位于第二聚光镜12的焦点处。 [0053] Spectrophotometric pupil confocal laser detection system 11 is located on the two reflection direction of the spectroscopic system 6; spectroscopic pupil laser confocal detection system 11 includes a second condenser lens 12 and the image acquisition system 13, the detection surface wherein the image acquisition system 13 It is located at the second focal point of the condenser lens 12.

[0054] 数据处理单元17用于处理拉曼光谱探测系统7和分光瞳激光共焦探测系统11所得到的信息。 [0054] The data processing unit 17 and the beam splitter 7 for pupil detection laser Raman spectroscopy system information processing system 11 to detect the resulting confocal. 包括分割焦斑探测模块18、极值运算模块19、数据融合模块20。 Dividing the focal spot detection module 18 comprising, extrema calculation module 19, a data fusion module 20. 三者关联关系为:分割焦斑探测模块18将图像采集系统13采集到的艾利斑进行分割并探测,得到的信号进入极值运算模块19进行极值求取,得到的样品测量点位置信息进入数据融合模块20。 Three association of: dividing collected 13 Airy focal spot detection module 18 divides the image acquisition system and detecting the resulting signal enters the arithmetic module 19 extrema extrema is obtained, the sample measurement point position information obtained enter data fusion module 20. 数据融合模块20还接收探测器10得到的光谱信号,并融合光谱信息和位置信息。 Spectral data fusion module 20 also receives signals obtained by the detector 10, and fused spectral information and position information.

[0055] 计算机处理系统21负责协调和控制整个装置的运作。 [0055] The computer processing system 21 is responsible for coordinating and controlling the operation of the entire device. 计算机处理系统21与位移传感器16、三维扫描系统15、数据融合模块20相连接;图像采集系统13和分割焦斑探测模块18相连接,数据融合模块21与探测器10相连接; The computer processing system 21 and the displacement sensor 16, a three-dimensional scanning system 15, 20 is connected to a data fusion module; image acquisition system 13 and split the focal spot detection module 18 is connected to a data fusion module 21 is connected to the detector 10;

[0056] 其中,通过计算机处理系统21控制三维扫描系统15移动被测样品5,使不同区域瑞利光及对应该区域被测样品5的拉曼散射光通过测量物镜2和收集光瞳4 ;位移传感器16用于将被测样品5的实时位置反馈至计算机处理系统21为被测样品5的运动提供依据。 [0056] wherein the computer processing system 21 is controlled by three-dimensional scanning system 15 moves the test sample 5, so that different regions of the Rayleigh light and 2 and the pupil should be collected Raman scattered light test sample by measuring the area 5 of the objective lens 4; displacement sensor 16 for the position of the test sample 5 is fed back to the real-time computer processing system 21 as a moving test sample 5 provide a basis.

[0057] 特别的,可以将圆形照明光瞳和收集光瞳替换为其他形状(如D形,构成D形分光瞳激光共焦拉曼光谱测试方法示意图,如图3所示)。 [0057] In particular, it can be circular illumination pupil and the collection pupil Alternatively (confocal Raman spectroscopy methods schematic, such as shown D-shaped, D-shaped configuration of the laser beam splitter 3 pupil) other shapes.

[0058] 特别的,本发明装置还可以包括第一针孔22,构成具有共焦光谱探测系统的分光瞳激光共焦拉曼光谱测试装置,如图5所示。 [0058] In particular, the present invention may further comprise a first pinhole means 22, constituting a confocal Raman spectroscopy device having a spectral confocal laser spectroscopy pupil detection system, as shown in FIG. 第一针孔位于第一聚光镜8的焦点位置。 A first pinhole located at a first focal position of the condenser lens 8.

[0059] 特别的,本发明装置还可以在光源系统I和照明光瞳3之间增加光束调制系统23,如图6所示。 [0059] In particular, the apparatus of the present invention can be further increased and the light source system I 3 between the illumination pupil beam modulation system 23, as shown in FIG. 光束调制系统23包括沿光路依次放置的第三聚光镜24、位于第三聚光镜24焦点处的第二针孔25、焦点位于第二针孔25处的第四聚光镜26,构成具有光束调制系统的分光瞳激光共焦拉曼光谱测试装置。 Beam modulation system 23 comprising sequentially positioned along the optical path of the third condenser lens 24, a second pinhole 24 at the focal point in the third converging mirror 25, the focus is on the fourth condenser lens 25 at the second pinhole 26, a beam splitter having a beam modulation system composed of confocal Raman Spectroscopy pupil test device.

[0060] 特别的,本发明装置还可以在分光瞳激光共焦探测系统11中增加图像放大系统27,用于放大图像采集系统13探测到的艾利斑,如图7所示。 [0060] In particular, the apparatus of the present invention may also be in a spectral confocal laser detection system 11, the pupil image magnification system 27 increases, the image acquisition system 13 for amplifying the detected Airy, as shown in FIG. 图像放大系统27与第二聚光镜12共焦点,且图像采集系统13位于图像放大系统27焦点处,以提高分光瞳激光共焦测量装置的采集精度。 Image magnification system 27 and a second confocal converging lens 12 and the image acquisition system 13 located at the focus of an image magnification system 27, in order to improve the accuracy of the spectral pupil were collected laser power measuring means.

[0061] 实施例 [0061] Example

[0062] 本实施例中,二向色分光系统6为陷波滤波器,光谱探测器9为拉曼光谱探测器,图像采集系统13为CCD,图像放大系统27为放大物镜。 [0062] In this embodiment, the dichroic beam splitter 6 is a notch filter system, the spectral detector 9 is a Raman spectroscopy detector, the CCD image pickup system 13, an enlarged image of the objective lens system 27 is an enlarged.

[0063] 如图8所示,分光瞳激光共焦拉曼光谱探测过程如下: [0063] 8, confocal laser Raman spectroscopic spectrum of the pupil detection process is as follows:

[0064] 首先,由激光器构成的光源系统I发出可激发出被测样品拉曼光谱的激发光,激发光经第三聚光镜24会聚后进入第二针孔25成为点光源,经第四聚光镜26准直扩束后,形成平行的激发光束。 [0064] First, a configuration of a laser light source system I emit excitation light excited Raman spectra of a test sample, the excitation light by the third condenser lens 24 converges into the second pinhole 25 becomes a point light source, a condenser lens 26 through the fourth collimated beam expander, formed of parallel excitation beam. 激发光束经过照明光瞳3、测量物镜2后,聚焦在被测样品5上,激发出载有被测样品5光谱特性的拉曼散射光。 After illumination pupil excitation beam 3, after the measurement objective 2, 5 is focused on the test sample, the test sample containing the excitation spectral characteristics 5 Raman scattered light.

[0065] 然后,通过计算机系统21控制三维扫描系统15移动被测样品5,使不同区域瑞利光及对应该区域被测样品5的拉曼散射光通过测量物镜2和收集光瞳4,陷波滤波器6将瑞利反射光和拉曼散射光分离。 [0065] Then, the control system of three-dimensional scanning system 21 by the computer 15 moves the test sample 5, so that different regions of the Rayleigh light and 4, the notch should test sample Raman scattered light 5 by measuring the area of ​​the objective lens 2 and the collection pupil reflected light and Rayleigh filter 6 separating Raman scattered light.

[0066] 经陷波滤波器6反射的瑞利光进入分光瞳激光共焦测量系统11,经第二聚光镜12会聚后进入放大物镜27,被放大后的光斑被CCD13探测,CCD13探测到的光斑进入分割焦斑探测模块18,在探测焦斑中心设置微小区域探测区域14,测得这个区域的响应为I (U),极值运算模块19将得到的响应进行极值求取,得到分光瞳激光共焦响应曲线28的“极值点”;分光瞳激光共焦响应曲线28的“极值点”与激发光束的聚焦焦点精确对应,通过响应曲线28的“极值点”来获得样品表面的高度信息,结合位移传感器16反馈的位置信息重构出被测样品5的表面三维形貌。 [0066] reflected by the notch filter 6 Rayleigh light enters the pupil spectral confocal laser measuring system 11, after entering the second condensing lens 12 condensing lens 27 enlarged spot is detected after being amplified CCD 13, CCD 13 detects the light spot into the dividing the focal spot 18 in response to the detection module, detecting minute region disposed in the detection region 14 the center of the focal spot, as measured in response to this region is I (U), extrema calculation module 19 obtains extreme value is obtained, to obtain spectroscopic laser pupil confocal response "extreme point" of the curve 28; pupil confocal spectral response "extreme point" of the curve 28 and the excitation light beam focused focal correspond precisely to obtain the sample surface in response to "extreme point" of the curve 28 altitude information, the position information of the displacement sensor 16 in conjunction with feedback reconstruct a three-dimensional surface topography of the test specimen 5.

[0067] 经陷波滤波器6透射的拉曼散射光进入拉曼光谱探测系统7,经第一聚光镜8及位于其焦点上的第一针孔22后进入拉曼光谱探测器9以及其后的探测器10,测得载有被测样品5光谱信息的拉曼散射光谱响应曲线I (r) 29,其中r为被测样品5受激发光所激发出拉曼散射光的波长。 [0067] transmitted by the notch filter 6 Raman spectroscopy Raman scattered light enters the detection system 7, and 8 via the first condenser lens in its first focus on the pinhole 22 enters the Raman spectroscopy probe 9 backward and thereafter detector 10, a test sample containing the measured spectrum information 5 Raman scattering spectral response I (r) 29, the test sample 5 wherein r is excited by the excitation light wavelength of the Raman scattered light.

[0068] 只对接收瑞利光的分光瞳激光共焦探测系统11获得探测光斑进行处理得到探测区域14分光瞳激光共焦响应I (U),通过其“极值点”精确捕获激发光斑的焦点位置,系统可以进行高空间分辨的三维尺度层析成像;只对接收拉曼散射光的拉曼光谱探测系统7获得的光谱响应I (r) 29进行处理时,系统可以进行拉曼光谱探测;同时对接收瑞利光的探测器11获得的分光瞳激光共焦响应I (U)和拉曼散射光的拉曼光谱探测系统7获得的光谱信号I (r)进行处理时,系统可以进行高空间分辨的微区图谱层析成像,即实现被测样品5几何位置信息和光谱信息的高空间分辨的“图谱合一”探测效果; [0068] Only the spectroscopic pupil Rayleigh light received laser confocal detection system 11 obtains the spot detection processing to obtain the detection area 14 in response to the spectral confocal laser pupil I (U), through which the "extreme point" of accurately capturing the excitation focus spot position, the system may be a high spatial resolution of tomography three dimensions; only 29 processing, the system can detect the Raman spectrum of the Raman scattered light spectrum of the received Raman spectrum obtained in response to the detection system 7 I (r); spectroscopic signal I (r) while the Raman spectrum spectroscopic detection system pupil Rayleigh light detector of the laser receiver 11 obtains a confocal response I (U) 7 and the Raman scattered light obtained when the processing system may be a high spatial resolution micro image atlas chromatography, i.e., to achieve the geometrical position information of the test sample 5 and spectral information of high spatial resolution "Atlas one" detection results;

[0069] 如图8所示,分光瞳激光共焦拉曼光谱探测装置包括产生激发光束的光源系统1、测量物镜2、照明光瞳3、收集光瞳4、被测样品5、陷波滤波器6、拉曼光谱探测系统7、分光瞳激光共焦测量系统11、三维扫描系统15、位移传感器16以及数据处理单元17 ;其中,将测量物镜2的光瞳面分割为照明光瞳3和收集光瞳4,测量物镜2和照明光瞳3放置在光源系统I的光束出射方向上,照明光瞳3与激发光束同轴,陷波滤波器6放置在收集光瞳4之后,拉曼光谱探测系统7放置在陷波滤波器6的透射方向上,分光瞳激光共焦测量系统11放置在陷波滤波器6的反射方向上,数据处理单元17用于融合并处理拉曼光谱探测系统7、分光瞳激光共焦探测系统11和位移传感器16采集到的数据;拉曼光谱探测系统7中第一聚光镜8的焦点处放置第一针孔22对杂散光进行滤除,以提高拉曼光谱探测效 [0069] As shown, the spectral confocal laser Raman spectroscopy pupil detection device 8 comprises a light source generating an excitation beam system, measurement objective 2, 3 illumination pupil, the pupil collected 4, test sample 5, a notch filter 6, Raman spectroscopy detection system 7, pupil spectral confocal laser measuring system 11, three-dimensional scanning system 15, the displacement sensor 16 and a data processing unit 17; wherein the measurement objective pupil plane is divided into two illumination pupil 3 and collection pupil 4, measurement illumination pupil of the objective lens 2 and 3 is placed in the beam exit direction of the light source system I, illumination pupil 3 and the coaxial excitation beam, the notch filter 6 is placed after collection pupil 4, Raman spectroscopy detection system 7 is placed in the transmission direction of the notch filter 6, the pupil spectral confocal laser measuring system 11 is placed in the reflection direction of the notch filter 6, the data processing unit 17 for processing and integration of the Raman spectrum detection system 7 spectroscopic data pupil detection system 11 and the laser displacement sensor 16 acquired confocal; Raman spectroscopy detection system a first placed first pinhole 22 pairs of spurs focal point of condenser lens 8 in filtered light 7, to improve the Raman spectra detection efficiency ;光源系统可以通过包括第三聚光镜24、位于第三聚光镜24焦点处的第二针孔25、以及焦点位于第二针孔25处的第四聚光镜26构成的光束调制系统23对光源系统I发出的激发光束进行调制,以提高激发光束的质量;位于图像采集系统11中第二聚光镜12焦面上的像通过图像放大物镜27放大后到达CCD13的探测面上,以便于探测器进行分割焦斑探测,提高探测精度。 ; Beam modulation system may be located at the light source system 25 by the second pinhole comprises a third condenser lens 24, the pinhole 25 located at the second focal point 24 at the third condenser lens, and a focus lens 26 constituting a fourth condenser 23 emits the light source system I the excitation beam is modulated in order to improve the quality of the excitation beam; after image acquisition system 11 is located in the second focal plane of the condenser lens 12 of the objective lens 27 as an enlarged image enlarged by the arrival detection CCD13 surface so dividing the focal spot to the detector detection, improve detection accuracy.

Claims (7)

1.一种分光瞳激光共焦拉曼光谱测试方法,其特征在于,包括以下步骤: 在测量物镜⑵的光瞳面上放置照明光瞳⑶和收集光瞳⑷;光源系统⑴发出能够激发出拉曼光谱的激发光束;激发光束依次透过照明光瞳(3)、测量物镜(2)后,聚焦在被测样品(5)上,激发出载有被测样品光谱特性的拉曼散射光,同时反射出瑞利光;拉曼散射光和瑞利光依次经测量物镜(2)和收集光瞳(4)后到达二向色分光系统(6),二向色分光系统(6)对拉曼散射光和瑞利光进行无损分离; 经二向色分光系统(6)反射的瑞利光进入分光瞳激光共焦探测系统(11);分光瞳激光共焦探测系统(11)利用探测器横向偏移可使分光瞳共焦显微系统的轴向响应特性曲线产生相移的特性,实现对被测样品微区几何位置的探测;对接收瑞利光的分光瞳激光共焦探测系统(11)获得的探测光斑进行处理,得到探测区域, A laser confocal Raman spectroscopic pupil spectroscopy method comprising the steps of: placing an illumination pupil and the collection pupil ⑶ ⑷ pupil plane of the measurement objective ⑵; ⑴ system emits light to stimulate the Raman spectra of the excitation beam; excitation beam sequentially transmitted through the illumination pupil (3), (2) measurement of the objective lens is focused on the test sample (5), containing the test sample excited Raman scattered light spectral characteristics while reflecting the Rayleigh light; Raman scattered light and the Rayleigh light sequentially through measurement objective (2) and the collection pupil (4) reaches the dichroic splitting system (6), a dichroic beam splitter system (6) Raman scattered light and the Rayleigh light nondestructive separation; after two reflected by the dichroic splitting system (6) Rayleigh light enters the pupil spectral confocal laser detection system (11); pupil spectral confocal laser detection system (11) using a lateral shift detector can axially spectral confocal microscopy system pupil response curve to produce a phase shift characteristic to achieve detection of the geometrical position of a micro test sample; spectroscopic pupil Rayleigh light received laser confocal detection spot detection system (11) obtained in , to give the detection zone, 得反映样品凹凸变化的分光瞳激光共焦响应I (U),即可进行高空间分辨的三维尺度层析成像探测,其中,u为轴向归一化光学坐标; 与此同时,经二向色分光系统(6)透射的拉曼散射光进入拉曼光谱探测系统(7),利用分光瞳激光共焦响应曲线的“极值点”与测量物镜(2)焦点位置精确对应这一特性,将分光瞳激光共焦响应曲线的“极值点”精确对应测量物镜(5)的焦点,通过寻找“极值点”来精确捕获激发光斑焦点位置的光谱信息,实现高空间分辨的光谱探测; 当只对接收瑞利光的分光瞳激光共焦探测系统(11)获得的探测光斑进行处理时,能够进行高空间分辨的三维尺度层析成像探测; 当只对接收拉曼散射光的拉曼光谱探测系统(7)获得的光谱信号进行处理时,能够进行高空间分辨的光谱探测; 当同时对接收瑞利光的分光瞳激光共焦探测系统(11)获得的探 Samples have irregularities reflect changes in pupil confocal laser spectral response I (U), can be performed in three dimensions with high spatial resolution tomography probe, wherein, u is a normalized optical coordinate axis; At the same time, by the dichroic a color splitting system (6) transmitted into the Raman spectroscopy Raman scattered light detection system (7), using a confocal laser pupil spectral response curve of "extreme point" and the measurement objective (2) corresponds exactly to the focal position of this feature, the laser confocal spectral response pupil "extreme point" curve corresponds precisely focus measurement objective (5) and searching the "extreme point" capture spectral information to accurately position the focal spot of the excitation, to achieve spectral detection of high spatial resolution; when the probe only the spectral flare pupil confocal laser Rayleigh light receiving detection system (11) was subjected to treatment, it is possible to detect three-dimensional tomographic scale of high spatial resolution; when only Raman scattered light receiving Raman spectrum spectrum signal detection system (7) obtained when the processed spectral detection can be performed with high spatial resolution; the same time, when the spectroscopic probe pupil confocal laser Rayleigh light receiving detection system (11) obtained in 光斑和接收拉曼散射光的拉曼光谱探测系统(7)获得的光谱信号进行处理时,能够进行高空间分辨的微区图谱层析成像,实现对被测样品“图谱合一”的分光瞳激光共焦拉曼光谱高空间分辨成像与探测。 When processed, the signal spectrum can be received Raman scattered light spot and a Raman spectrum detection system (7) obtained in high spatial resolution imaging of micro chromatographic pattern, achieve a test sample "Atlas one" spectral pupil confocal Raman Spectroscopy high spatial resolution imaging and detection.
2.根据权利要求1所述的分光瞳激光共焦拉曼光谱测试方法,其特征在于:照明光瞳(3)和收集光瞳(4)是圆形的或D形的。 The spectroscopic pupil laser according to claim 1 confocal Raman spectroscopy method, comprising: an illumination pupil (3) and the collection pupil (4) is circular or D-shaped.
3.根据权利要求1所述的分光瞳激光共焦拉曼光谱测试方法,其特征在于:激发光束是线偏振光束或圆偏振光束,偏振光束与光瞳滤波技术的结合压缩测量聚焦光斑尺寸,提高分光瞳激光共焦探测系统的横向分辨力。 The laser of claim 1 pupil spectral confocal Raman spectroscopy method as claimed in claim, characterized in that: the excitation beam is a linearly polarized beam or a circularly polarized beam, the polarization beam combined with the compression pupil filtering techniques focus measurement spot size, confocal spectral pupil improve lateral resolution of the detection system.
4.一种分光瞳激光共焦拉曼光谱测试装置,包括光源系统(I)、测量物镜(2)、二向色分光系统(6)、拉曼光谱探测系统(7)、分光瞳激光共焦探测系统(11)、位移传感器(16)、数据处理单元(17)、三维扫描系统(15)和计算机处理系统(21);其特征在于:所述测试装置还包括照明光瞳⑶和收集光瞳(4),其中,照明光瞳(3)和收集光瞳⑷放置在测量物镜(2)的光瞳面上;照明光瞳(3)和测量物镜(2)依次位于光源系统(I)的激发光束出射方向上,照明光瞳(3)与激发光束同轴;二向色分光系统(6)位于收集光瞳(4)之后;被测样品(5)固定在三维扫描系统(15)载物台上; 拉曼光谱探测系统(7)位于二向色分光系统(6)的透射方向上;拉曼光谱探测系统(7)包括第一聚光镜(8)、光谱探测器(9)和探测器(10);其中,光谱探测器(9)的探测面位于第一聚光镜⑶的焦点处,探测器(10)位于光谱探测器(9)之后;探 A laser confocal Raman spectroscopic spectrum pupil test device comprising a light source system (the I), measurement objective (2), a dichroic beam splitter system (6), Raman spectroscopy detection system (7), the pupil spectral confocal laser focus detection system (11), a displacement sensor (16), the data processing unit (17), a three-dimensional scanning system (15) and a computer processing system (21); characterized in that: said test means further comprises illumination pupil and the collection ⑶ pupil (4), wherein the illumination pupil (3) and the collection pupil ⑷ placed on the objective lens pupil plane measurement (2); a pupil illumination (3) and the measurement objective (2) successively positioned light source system (I ) in the emission direction of the excitation beam, the illumination pupil (3) coaxially with the excitation beam; dichroic splitting system (6) after collection pupil (4) is located; the test sample (5) fixed to the three-dimensional scanning system (15 ) the stage; Raman spectroscopy detection system (7) located on the two transmission directions dichroic splitting system (6); a Raman spectrum detection system (7) comprises a first condenser lens (8), the spectral detector (9) and a detector (10); wherein the spectral detector (9) located at the focal plane of the detection of the first condenser lens ⑶, the detector (10) located in the spectral detector (9) after; probe 器(10)将得到的光谱信号发送给数据处理单元(17); 分光瞳激光共焦探测系统(11)位于二向分光系统(6)的反射方向上;分光瞳激光共焦探测系统(11)包括第二聚光镜(12)和图像采集系统(13),其中,图像采集系统(13)的探测面位于第二聚光镜(12)的焦点处; 数据处理单元(17)用于处理拉曼光谱探测系统(7)和分光瞳激光共焦探测系统(11)所得到的信息,数据处理单元(17)包括分割焦斑探测模块(18)、极值运算模块(19)、数据融合模块(20);其中,分割焦斑探测模块(18)将图像采集系统(13)采集到的艾利斑进行分割并探测,得到的信号进入极值运算模块(19)进行极值求取,得到的样品测量点位置信息进入数据融合模块(20);数据融合模块(20)接收探测器(10)得到的光谱信号,并融合光谱信息和位置信息; 计算机处理系统(21)负责协调和控制整个测试装置的运作;计算机处理系统(21)与 Detection system (11 spectral pupil confocal laser; (10) the spectral signal obtained to the data processing unit (17); spectral pupil confocal laser detection system (11) located at a dichroic beam splitter system (6) in the reflection direction ) comprising a second condenser lens (12) and the image acquisition system (13), wherein the image acquisition system (13) in the second detection surface condenser lens (12) focal point; a data processing unit (17) for processing the Raman spectroscopy detection system (7) and the beam pupil confocal laser detection system (11) the information obtained, the data processing unit (17) comprises a division of the focal spot detection module (18), the arithmetic module extremum (19), data fusion module (20 ); wherein dividing the focal spot detection module (18) of the image acquisition system (13) collected and divided Airy detection signal obtained into the extrema computing module (19) for obtaining extrema, samples obtained measurement point position information into the data fusion module (20); spectral signal data fusion module (20) obtained from the detector receiving (10), and fused spectral information and position information; computer processing system (21) is responsible for coordinating and controlling the entire test apparatus operation; computer processing system (21) and 移传感器(16)、三维扫描系统(15)、数据融合模块(20)相连接;图像采集系统(13)和分割焦斑探测模块(18)相连接,数据融合模块(21)与探测器(10)相连接;其中,通过计算机处理系统(21)控制三维扫描系统(15)移动被测样品(5),使不同区域瑞利光及对应该区域的被测样品(5)的拉曼散射光通过测量物镜(2)和收集光瞳⑷;位移传感器(16)用于将被测样品(5)的实时位置反馈至计算机处理系统(21)。 Displacement sensors (16), a three-dimensional scanning system (15), data fusion module (20) is connected; image acquisition system (13) and dividing the focal spot detection module (18) is connected, data fusion module (21) and the detector ( 10) is connected; wherein the control system of three-dimensional scanning by a computer processing system (21) (15) moving the test sample (5), so that different regions of the Raman scattered light and the Rayleigh light to be a test sample (5) region by measuring the objective lens (2) and the collection pupil ⑷; displacement sensor (16) for the test sample (5) to the real-time computer processing position feedback system (21).
5.如权利要求4所述的一种分光瞳激光共焦拉曼光谱测试装置,其特征在于,所述测试装置还包括位于第一聚光镜(8)的焦点位置的第一针孔(22)。 5. The one laser spectroscopic pupil 4 of the confocal Raman spectroscopy device, characterized in that said test means further comprising a first pin hole (22) located at a first focal position of the condenser lens (8) .
6.如权利要求4所述的一种分光瞳激光共焦拉曼光谱测试装置,其特征在于,在光源系统⑴和照明光瞳⑶之间增加光束调制系统(23);光束调制系统(23)包括沿光路依次放置的第三聚光镜(24)、位于第三聚光镜(24)焦点处的第二针孔(25)、焦点位于第二针孔(25)处的第四聚光镜(26)。 6. The one laser spectroscopic 4 pupil confocal Raman spectroscopy device, characterized in that, to increase the beam modulation system (23) between the light source and the illumination system pupil ⑴ ⑶; beam modulation system (23 ) comprises a third condenser lens (24) are sequentially positioned along the optical path, a third condenser lens (24) of the second pinhole (25) at the focal point, the focal point in the fourth condenser lens (26) at a second pin hole (25).
7.如权利要求4所述的一种分光瞳激光共焦拉曼光谱测试装置,其特征在于,在分光瞳激光共焦探测系统(11)中增加图像放大系统(27),用于放大图像采集系统(13)探测到的艾利斑;图像放大系统(27)与第二聚光镜(12)共焦点,且图像采集系统(13)位于图像放大系统(27)焦点处。 7. The one laser spectroscopic 4 pupil confocal Raman spectroscopy device, characterized in that, in a spectral confocal detection system pupil laser (11) to increase the image magnification system (27) for amplifying the image acquisition system (13) detects the Airy; image magnification system (27) and a second condenser lens (12) co-focus, and the image acquisition system (13) (27) at the focal point of the image magnification system.
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