CN115825014A - Surface interface in-situ spectrum and imaging test system coupled by plasmon - Google Patents

Abstract

The invention provides a plasmon coupling surface interface in-situ spectrum and imaging test system, which records reflection light intensity or spectrum while changing a laser incidence angle, excites surface plasmon polaritons of a silver film in an optical near-field range in a prism coupling mode when the incident laser angle is just equal to the incident laser angle, observes that the intensity of reflected light reaches the lowest point, namely surface plasmon resonance, keeps incident light exciting a sample at a resonance angle, and then a target molecule system on the surface of the silver film can scatter and absorb electromagnetic surface waves bound in the near-field range and re-emits optical signals to a far field. The system realizes the in-situ combination of technologies such as surface plasmon resonance, surface enhanced Raman scattering spectrum, total internal reflection fluorescence spectrum and imaging by collecting the optical signals.

Description

A plasmon-coupled surface-interface in-situ spectroscopy and imaging test system

technical field

The invention belongs to the field of surface-interface spectral analysis, in particular to a surface-interface in-situ spectrum and imaging test system coupled with plasmons.

Background technique

Studying and understanding the physical and chemical processes of surfaces and interfaces is of great significance to promote many disciplines including photoelectrocatalysis, sensing and detection, integrated optoelectronic devices, etc. A variety of technical means have been established in the process of scientific exploration of surfaces and interfaces, such as electron microscopy, scanning probe microscopy, photoelectron spectroscopy, and electrochemical techniques, which provide information about surfaces and interfaces at different levels. As an important research method, spectroscopy has the advantages of low damage, in situ, relaxed operating environment, and the ability to respond to material structure information, and has been extensively and deeply developed in surface and interface science.

Raman spectroscopy and fluorescence spectroscopy are the two most widely used spectroscopic techniques in surface and interface analysis. As a fingerprint spectrum, Raman spectroscopy is related to the vibrational modes of molecules in the chemical environment. By analyzing the peak position, peak intensity and Information such as half-peak width can obtain the structure and orientation changes of surface interface molecules. And with the development of surface-enhanced Raman spectroscopy technology, the sensitivity of Raman spectroscopy has been significantly improved. By constructing a suitable Raman-enhanced substrate and selecting efficient excitation and acquisition methods, Raman analysis and detection at the single-molecule level has become a reality. Fluorescence spectroscopy and imaging are very universal techniques in the study of physical and chemical processes at surfaces and interfaces. On the one hand, fluorescence spectroscopy reflects the excited state information of substances, and indirect evidence of intermolecular interactions can be obtained by analyzing fluorescence spectroscopy. On the other hand, fluorescent labeling methods can be used to detect and track highly sensitive physical and chemical processes on the surface and interface, such as obtaining the distribution of target molecules in space, detecting binding events between molecules and substrates, and so on.

The characteristics of the surface-interface problem put forward high requirements for the analysis method. First, the surface-interface research focuses on the area in the micro-nano scale near the interface. In order to shield the interference from signals or background noise outside the interface, the selected technology Should have high spatial resolution in the direction perpendicular to the interface. On the other hand, since the analysis object in the surface-interface problem is often a thin film composed of a single layer or a few layers of molecules, the analysis technology must have a high enough sensitivity to realize the acquisition of low-concentration target substance signals. The physical and chemical processes that finally occur at the surface interface are highly complex. In order to analyze such problems, a variety of analytical techniques are required to be used together to achieve the characterization of the target in different dimensions.

Surface plasmons provide an effective solution to overcome the above problems. First, the electromagnetic field has a small penetration depth in the direction perpendicular to the metal-medium interface, thus effectively limiting the spectral detection range to the interface region. On the other hand, due to the optical field control ability of plasmons, the excitation field energy can be highly concentrated in the hotspot area, and this electromagnetic field effect can significantly enhance the intensity of Raman and fluorescence spectral signals.

Patent CN1657914A and academic article (Liu, Yu, et al.Review ofscientific instruments 81.3 (2010): 036105.) in the prior art, set up a kind of SPR-SERS in situ test device in these two works, the present invention and its The difference lies in three aspects, 1. Use the reflective side of the prism instead of the air-transmissive side to collect Raman signals, so as to effectively use the directional emission effect of plasmons. 2. Adding a polarization beam splitter prism to the reflection arm can simultaneously measure the optical signals of S and P polarizations, which can be applied to the research related to the molecular orientation of the interface. 3. In addition to being applied to testing SPR and SERS, the present invention can also perform total internal reflection fluorescence spectroscopy and imaging testing, realizing the coordinated use of the three technologies of SPR-SERS-TIRF. 4. In the present invention, the excitation light is coupled into the system through an optical fiber, instead of directly fixing the laser on the incident arm, so that the excitation wavelength can be switched conveniently.

Contents of the invention

In order to solve the problems existing in the surface-interface spectral analysis technology, combined with the advantages of surface plasmon resonance, surface-enhanced Raman spectroscopy, and total internal reflection fluorescence microscopy imaging, the present invention constructs a plasmon that can be controlled by incident angle In-situ test platform for coupled surface-interface fluorescence and Raman spectroscopy. This platform records the reflected light intensity or spectrum while changing the incident angle of the laser. When the incident laser angle is just in the way of prism coupling, the silver film (or The surface plasmon polaritons of the gold film) will observe that the reflected light intensity reaches the lowest point, which is called surface plasmon resonance. Keep the incident light at the resonance angle to excite the sample. At this time, the target molecular system modified on the surface of the silver film will scatter and absorb the electromagnetic surface wave bound in the near field range, and re-emit the optical signal to the far field. The inventive device realizes the in-situ combination of surface plasmon resonance, surface-enhanced Raman scattering spectroscopy, total internal reflection fluorescence spectroscopy and imaging by collecting these optical signals.

A plasmon-coupled surface-interface in-situ spectroscopy and imaging test system of the present invention includes a laser coaxial adjustment and fiber coupling system, a frame and a rotating arm drive device, a variable-angle incident arm and a light excitation system, and can A variable-angle reflection arm, an optical signal acquisition system, a sample stage and a prism mounting frame, and a microscopic spectrum collection and imaging system; a cylindrical prism is installed on the sample stage and the prism mounting frame;

The laser coaxial adjustment and fiber coupling system includes: a long-pass dichroic mirror, a group of lasers with different wavelengths, a fiber coupler, an optical fiber and a reflector, and the long-pass dichroic mirror combines a group of lasers with different wavelengths into a One beam, after being reflected by the mirror, is coupled into the optical fiber by the fiber coupler, and finally transmitted to the variable-angle incident arm and the optical excitation system;

The variable-angle incident arm and optical excitation system include: fiber flange, aspheric lens, linear polarizer, half-wave plate, adjustable diaphragm, and focusing objective lens; the light beam coupled to the free space by the optical fiber is first collimated by the aspheric lens Straight, and then through the linear polarizer and the broadband half-wave plate in turn to generate linearly polarized light with adjustable polarization direction, the linearly polarized light is spatially filtered through the adjustable diaphragm, and finally focused by the focusing objective lens into the cylindrical prism;

The reflective arm of the variable angle and the optical signal acquisition system include: collecting objective lens, optical filter, first focusing lens, first optical fiber connector, polarization beam splitting prism, second focusing lens, second optical fiber connector; The reflected light passes through the filter and enters the polarization beam splitter prism after being collected by the collecting objective lens. The S polarized light is reflected while the P polarized light passes through the polarization beam splitter prism. The reflected light and the transmitted light pass through the first focusing lens and the second focusing lens respectively. The focusing lens is focused to the center of the photodiode or optical fiber fixed at the first optical fiber joint and the second optical fiber joint;

The micro-spectrum acquisition and imaging system includes component imaging objective lens, mirror, long-pass filter, the third focusing lens, camera, 1:1 beam splitter, the fourth focusing lens, spectrometer; imaging objective lens collects the sample at the interface Raman and fluorescence signals, the optical signal is first reflected by the mirror, and then the excitation light is filtered out by the long-pass filter, and then the optical signal is divided into reflection and transmission by the 1:1 beam splitter, and the reflection light passes through the third The focusing lens is imaged on the image plane of the camera, and the transmitted light is focused to the slit of the spectrometer through the fourth focusing lens for spectrum collection;

The variable-angle incident arm, optical excitation system, variable-angle reflective arm and optical signal acquisition system are symmetrically distributed on both sides of the sample stage and the prism mounting frame, and the surface plasmon resonance angle is scanned and measured at the resonance angle. Raman scattering and fluorescence spectroscopy;

The microscopic imaging device is located at the bottom of the sample stage and the prism mounting frame, and is used for total internal reflection fluorescence imaging and spectrum collection.

Further, the fluorescence and Raman spectrum signals collected by the microscopic spectrum collection and imaging system are excited by the surface plasmon polaritons of the metal film, and the signals are entirely from the sample at the surface interface of the metal film.

Further, the surface plasmon polaritons of the metal film are used to excite the fluorescence and Raman spectrum signals of the molecules, and the penetration depth of the surface plasmon polaritons is passed through the rotation variable angle incident arm and the optical excitation The system adjusts.

Further, the spectral signal is collected by the variable-angle reflective arm and the optical signal collection system on the side of the cylindrical prism or the microscopic spectrum collection and imaging system on the air side.

Further, the variable-angle reflecting arm and the polarization beam splitting prism in the optical signal collection system simultaneously measure the reflected light intensity or Raman spectrum in two orthogonal polarization states.

Description of drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings that need to be used in the description of the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present invention. For those skilled in the art, other drawings can also be obtained based on these drawings without any creative effort.

Figure 1 is a schematic diagram of the laser coaxial adjustment and fiber coupling system;

Fig. 2 is a schematic diagram of the in-situ spectroscopy and imaging test system of the plasmon coupling surface interface of the present invention;

Figure 3 is the total internal reflection fluorescence imaging collected when the critical angle is excited;

Fig. 4 is the total internal reflection fluorescence spectrum collected when the critical angle is excited;

Fig. 5 is the SPR reflection spectrum of silver film, and this figure abscissa is angle, and vertical axis is normalized light intensity;

Fig. 6 is the Raman spectrum of the 4-MBA probe molecule collected under the SPR resonance angle;

Among them, 1 frame and rotating arm driving device; 2 variable angle incident arm and light excitation system, including: 2-1 fiber optic flange, 2-2 aspherical lens, 2-3 linear polarizer, 2-4 half-wave film, 2-5 adjustable diaphragm, 2-6 focusing objective lens, 3 variable angle reflection arms and optical signal acquisition system, including: 3-1 collecting objective lens, 3-2 optical filter, 3-3 polarizing beam splitting prism, 3-4 the first focusing lens, 3-5 the first optical fiber interface, 3-6 the second focusing lens, 3-7 the second optical fiber interface; 4 sample stage and prism mounting frame; 5 microscopic spectrum acquisition and imaging system, including Components 5-1 imaging objective lens, 5-2 mirror, 5-3 long-pass filter, 5-4 third focusing lens, 5-5 camera, 5-6 1:1 beam splitter, 5-7 fourth Focusing lens, 5-8 spectrometer.

Detailed ways

In order to make the purposes, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments It is a part of the embodiments of this application, not all of them. Based on the embodiments in this application, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the scope of protection of this application.

In the accompanying drawings of specific embodiments of the present invention, in order to better and more clearly describe the working principle of each component in the system, the connection relationship of each part in the device is shown, and only the relative positions between the components are clearly distinguished. The relationship does not constitute a limitation on the signal transmission direction, connection sequence, and the size, size, and shape of each part of the component or structure.

The specific structure of the device of the present invention mainly includes the laser coaxial adjustment and fiber coupling system as shown in Figure 1, and the schematic diagram of the surface interface in-situ spectroscopy and imaging test system for plasmon coupling as shown in Figure 2, including a machine Frame and rotating arm driving device; 2 variable-angle incident arm and light excitation system; 3 variable-angle reflective arm and optical signal acquisition system; 4 sample stage and prism mounting frame; 5 microscopic spectrum acquisition and imaging system.

The laser coaxial adjustment and fiber coupling system is shown in Figure 1. A group of lasers with different wavelengths are combined into one beam through a long-pass dichroic mirror. in the light path of the arm. The wavelength of the laser used in the present invention can be 375nm, 405nm, 473nm, 532nm, 633nm and so on. The specific wavelength of the laser can be changed as needed.

As shown in Figure 2, the optical excitation system and the optical signal acquisition system are respectively fixed on the frame and the variable-angle incident arm and variable-angle reflection arm of the rotating arm drive device, and can be moved along the vertical direction of the rotating arm through the translation platform to achieve Realize the collimation adjustment of the initial optical path. The bottom microscopic imaging and spectrum acquisition device is fixed on the horizontal slide rail, and its whole can move independently on the X-Y plane relative to the rotary table, so as to realize the coincidence of the collection range and the excitation area.

The specific working principle of the present invention is: as shown in Figure 1, the laser beams emitted by a group of lasers with different wavelengths are coupled into the optical fiber after being combined by a dichroic mirror, and the output end of the optical fiber is connected to the optical fiber as shown in Figure 2 On the fiber flange 2-1 of the optical excitation system, the light beam coupled from the fiber to free space is first collimated by an aspheric lens 2-2, and then polarized by a linear polarizer 2-3 and a broadband half-wave plate 2-4 in turn. Linearly polarized light with adjustable direction, the linearly polarized light is spatially filtered by the adjustable diaphragm 2-5, and finally focused by the focusing objective lens 2-6 into the cylindrical prism whose center coincides with the rotation axis (in the case of sufficient laser power density It is not necessary to focus, just remove 2-6 at this time).

After the laser light undergoes total internal reflection on the lower surface of the cylindrical prism fixed at the sample stage and the prism mounting frame 4, a surface electromagnetic mode called the evanescent field will be formed. Surface plasmon polaritons are excited at the interface formed by the film and air. Thereafter, the detection optical path of the optical signal is divided into two paths:

One of them is fixed on the variable-angle reflection arm and optical signal acquisition system 3 on the right side of Figure 1 to collect reflected light and Raman scattered light. The optical signal is first collected by the collection objective lens 3-1, and then passed through the detachable The filter 3-2 enters the polarizing beam splitting prism 3-3, the S polarized light is reflected by the polarizing beam splitting prism 3-3 and the P polarized light passes through the polarizing beam splitting prism. Reflected light and transmitted light are respectively focused to the center of the optical detector or optical fiber fixed at the first optical fiber interface 3-5 and the second optical fiber interface 3-7 through the first focusing lens 3-4 and the second focusing lens 3-6 . When using the system to collect reflected light intensity, a photodetector is installed at the optical fiber interface, and the current signal is transmitted to the computer after passing through the transimpedance amplifier, low-pass filter and data acquisition card in turn. When the device works in the Raman spectrum acquisition mode, an optical fiber is installed at the collecting end, and the Raman scattering signal is coupled into the spectrometer 5-8 through the optical fiber.

The other path is on the air side below the prism, and the fluorescence of the interface sample is collected through the imaging objective lens 5-1. The fluorescence signal is first reflected by the mirror 5-2, and then the excitation light is filtered out by the long-pass filter 5-3. After that, the fluorescence signal is filtered out. The 1:1 beam splitter 5-6 is equally divided into reflection and transmission. The reflected light is imaged to the image plane of the camera 5-5 through the third focusing lens 5-4, while the transmitted light passes through the fourth focusing lens 5-7. Focus to the slits of the spectrometer 5-8 for spectrum acquisition.

In Example 1, fluorescence spectrum and imaging of silica fluorescent microspheres coated with RBITC (rhodamine B isocyanate).

The laser wavelength used in this embodiment is 532nm (selected according to the excitation spectrum characteristics of the fluorophore).

On the BK7 glass with a size of 10mm*10mm*1mm sputtered with a 45nm silver film, an ethanol solution of silica microspheres wrapped with a fluorescent dye RBITC was spin-coated. The plates and cylindrical prisms are index matched by cedar oil.

Connect the photodetector to the receiving end of the variable-angle reflective arm signal acquisition system, such as the polarization beam splitter 3-5 or fiber coupler or photodiode 3-7, and scan the variable-angle incident arm in the opposite direction at the same speed And the optical excitation system 2, the variable-angle reflector arm and the optical signal acquisition system 3, to ensure that the direction of the reflector arm is always kept at the reflection angle. When the minimum value appears in the reflection intensity curve, it means that the surface plasmon resonance has been realized At this time, the fluorescence imaging is performed through the optical path at the bottom of the prism to obtain Figure 3, and the fluorescence spectrum is collected at the same time to obtain Figure 4. The bright part in Figure 3 is the silica sphere containing the fluorescent dye, because in the experiment the fluorescent dye is excited by the evanescent field limited on the surface of the silver film, so the imaging shows good contrast without the interference of the fluorescent background. The peak at 560-600nm shown in the fluorescence spectrum in Figure 4 is the fluorescence emission peak of the dye RBITC.

Example 2: Surface Plasmon Resonance Enhanced Raman Scattering Spectrum of Silver Film Surface Modified 4-Mercaptobenzoic Acid.

The laser wavelength used in this embodiment is 532nm, and the optical power density is 10mW/cm ² .

The Raman spectrum signal collected in this embodiment comes from the Raman probe molecule 4-mercaptobenzoic acid.

First, magnetron sputtering a chromium layer with a thickness of 5nm on the BK7 glass with a size of 10mm*10mm*1mm is used to enhance the adhesion of the silver film to the substrate, and then continue to magnetron sputter a 50nm silver layer on the surface of the chromium layer. The obtained BK7/chromium/silver substrate was soaked in an ethanol solution of 4-mercaptobenzoic acid at a concentration of 10uM/L to modify the Raman probe molecule 4-mercaptobenzoic acid on the surface of the silver film. Finally, the resulting BK7/chrome/silver/4-MBA was bonded to a lenticular prism with cedar oil coated between the substrate and the lenticular prism to match the refractive index.

Connect the photodetector to the receiving end of the variable-angle reflective arm signal acquisition system, such as the polarization beam splitter 3-5 or the fiber coupler or photodiode 3-7, and use the computer control software to drive the variable-angle incident arm to rotate to Change the incident angle of the laser, and at the same time, the variable-angle reflection arm rotates with the incident arm, and record the reflected light intensity at the reflection angle, and the SPR reflection spectrum as shown in Figure 5 can be obtained. The reflectivity presents the minimum value at 44.3 degrees, and the incident The energy of the light is coupled into the surface plasmon polaritons of the silver film, and this electromagnetic field pattern bound to the surface of the silver film can be further scattered by the molecules into the far field and detected.

Scan the variable-angle incident arm to the SPR resonance angle, replace the photodetector on the first optical fiber interface 3-5 or the second optical fiber interface 3-7 with an optical fiber, and connect the output end of the optical fiber to the spectrometer. The results shown in Figure 6 can be obtained by collecting the Raman spectrum. The peaks at the Raman shifts of 1150 and 1600 wavenumbers belong to the characteristic peaks of the Raman probe molecule 4-mercaptobenzoic acid.

In the above embodiments, all or part of them may be implemented by software, hardware, firmware or any combination thereof. When implemented using software, it may be implemented in whole or in part in the form of a computer program product. The computer program product includes one or more computer instructions. When the computer program instructions are loaded and executed on the computer, the processes or functions according to the embodiments of the present application will be generated in whole or in part. The computer can be a general purpose computer, a special purpose computer, a computer network, or other programmable devices. The computer instructions may be stored in or transmitted via a computer-readable storage medium. The computer-readable storage medium may be any available medium that can be accessed by a computer, or a data storage device such as a server or a data center integrated with one or more available media. The available medium may be a magnetic medium (for example, a floppy disk, a hard disk, or a magnetic tape), an optical medium (for example, DVD), or a semiconductor medium (for example, a solid state disk (solid state disk, SSD)) and the like.

The above is only a specific embodiment of the application, but the scope of protection of the application is not limited thereto. Any person familiar with the technical field can easily think of various equivalents within the scope of the technology disclosed in the application. Modifications or replacements, these modifications or replacements shall be covered within the scope of protection of this application. Therefore, the protection scope of the present application should be based on the protection scope of the claims.

Claims (5)

1. A plasmon-coupled surface-interface in-situ spectroscopy and imaging test system, characterized in that it includes a laser coaxial adjustment and fiber coupling system, a frame and a rotating arm drive device, a variable-angle incident arm, and optical excitation system, a variable-angle reflector arm, an optical signal acquisition system, a sample stage and a prism mounting frame, and a microscopic spectrum collection and imaging system; a cylindrical prism is installed on the sample stage and the prism mounting frame; The laser coaxial adjustment and fiber coupling system includes: a long-pass dichroic mirror, a group of lasers with different wavelengths, a fiber coupler, an optical fiber and a reflector, and the long-pass dichroic mirror combines a group of lasers with different wavelengths into a One beam, after being reflected by the mirror, is coupled into the optical fiber by the fiber coupler, and finally transmitted to the variable-angle incident arm and the optical excitation system; The variable-angle incident arm and optical excitation system include: fiber flange, aspheric lens, linear polarizer, half-wave plate, adjustable diaphragm, and focusing objective lens; the light beam coupled to the free space by the optical fiber is first collimated by the aspheric lens Straight, and then through the linear polarizer and the broadband half-wave plate in turn to generate linearly polarized light with adjustable polarization direction, the linearly polarized light is spatially filtered through the adjustable diaphragm, and finally focused by the focusing objective lens into the cylindrical prism; The reflective arm of the variable angle and the optical signal acquisition system include: collecting objective lens, optical filter, first focusing lens, first optical fiber connector, polarization beam splitting prism, second focusing lens, second optical fiber connector; The reflected light passes through the filter and enters the polarization beam splitter prism after being collected by the collecting objective lens. The S polarized light is reflected while the P polarized light passes through the polarization beam splitter prism. The reflected light and the transmitted light pass through the first focusing lens and the second focusing lens respectively. The focusing lens is focused to the center of the photodiode or optical fiber fixed at the first optical fiber joint and the second optical fiber joint; The micro-spectrum acquisition and imaging system includes component imaging objective lens, mirror, long-pass filter, the third focusing lens, camera, 1:1 beam splitter, the fourth focusing lens, spectrometer; imaging objective lens collects the sample at the interface Raman and fluorescence signals, the optical signal is first reflected by the mirror, and then the excitation light is filtered out by the long-pass filter, and then the optical signal is divided into reflection and transmission by the 1:1 beam splitter, and the reflection light passes through the third The focusing lens is imaged on the image plane of the camera, and the transmitted light is focused to the slit of the spectrometer through the fourth focusing lens for spectrum collection; The variable-angle incident arm, optical excitation system, variable-angle reflective arm and optical signal acquisition system are symmetrically distributed on both sides of the sample stage and the prism mounting frame, and the surface plasmon resonance angle is scanned and measured at the resonance angle. Raman scattering and fluorescence spectroscopy; The microscopic imaging device is located at the bottom of the sample stage and the prism mounting frame, and is used for total internal reflection fluorescence imaging and spectrum collection. 2. according to claim 1, the in-situ spectroscopy and imaging test system of a kind of plasmon coupling surface interface is characterized in that, the fluorescence and the Raman spectrum signal collected by the microscopic spectrum acquisition and imaging system are collected by The surface plasmon polaritons of the metal film are excited, and the signal comes entirely from the sample at the surface interface of the metal film. 3. The surface-interface in-situ spectroscopy and imaging test system of a plasmon coupling according to claim 2, characterized in that the surface plasmon polaritons of the metal film are used to excite the fluorescence and Raman of molecules For spectral signals, the penetration depth of the surface plasmon polaritons is adjusted through the rotating variable-angle incident arm and the light excitation system. 4. according to claim 1, a kind of plasmon coupled surface interface in-situ spectroscopy and imaging test system is characterized in that, the spectral signal passes through the variable-angle reflector arm and the variable-angle reflector on the cylindrical prism side. Optical signal acquisition system or air-side microscopic spectrum acquisition and imaging system collection. 5. according to claim 1, a kind of surface-interface in-situ spectroscopy and imaging test system of plasmon coupling, is characterized in that, the polarization beam splitting prism in described variable-angle reflection arm and optical signal acquisition system, Simultaneously measure reflected light intensity or Raman spectra in two orthogonal polarization states.

Images