CN113433074A - Circular dichroism microscopic imaging system based on discrete modulation - Google Patents
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- 238000003384 imaging method Methods 0.000 title claims abstract description 18
- 238000002983 circular dichroism Methods 0.000 title claims abstract description 16
- 230000010287 polarization Effects 0.000 claims abstract description 30
- 238000012634 optical imaging Methods 0.000 claims abstract description 6
- 230000003287 optical effect Effects 0.000 claims description 39
- 239000011521 glass Substances 0.000 claims description 13
- 238000000386 microscopy Methods 0.000 claims description 8
- 230000000737 periodic effect Effects 0.000 claims description 3
- 230000009466 transformation Effects 0.000 claims 1
- 239000007787 solid Substances 0.000 abstract description 10
- 238000001142 circular dichroism spectrum Methods 0.000 abstract description 6
- 238000010521 absorption reaction Methods 0.000 abstract description 2
- 229910021532 Calcite Inorganic materials 0.000 abstract 1
- 238000005259 measurement Methods 0.000 abstract 1
- 239000002245 particle Substances 0.000 abstract 1
- CPBQJMYROZQQJC-UHFFFAOYSA-N helium neon Chemical compound [He].[Ne] CPBQJMYROZQQJC-UHFFFAOYSA-N 0.000 description 3
- 238000002267 linear dichroism spectroscopy Methods 0.000 description 3
- 238000000862 absorption spectrum Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
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- 238000000978 circular dichroism spectroscopy Methods 0.000 description 1
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- 239000012535 impurity Substances 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
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- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/19—Dichroism
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- G02B21/0092—Polarisation microscopes
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
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Abstract
A circular dichroism microscopic imaging system based on discrete modulation belongs to the field of optical imaging. Commercial CD spectrometers have difficulty measuring solid chirality, and their measured CD spectra are necessarily accompanied by artifact signals. The invention solves the problem of artifact signals when the existing CD spectrometer measures the chirality of a solid. According to the invention, linearly polarized light with alternating vertical and horizontal polarization directions is obtained by utilizing a calcite beam shifter and a chopper for discrete modulation, pure left-handed circularly polarized light and pure right-handed circularly polarized light are obtained by adopting an 1/4 wave plate and are irradiated onto a sample, and CD signals are formed by different absorption of the sample on the left-handed circularly polarized light and the right-handed circularly polarized light. Single particle measurement is realized through a confocal mode, and scanning imaging of a sample is realized through control over a nanometer mobile platform. The invention is suitable for chiral solid microscopic imaging.
Description
Technical Field
The invention relates to a chiral solid microscopic optical imaging system, and belongs to the field of optical imaging.
Background
The circular dichroism spectrum is the difference between the absorption spectrum of a sample under irradiation of left-handed circularly polarized light and the absorption spectrum thereof under irradiation of right-handed circularly polarized light. A substance is said to have circular dichroism (hereinafter CD) if it absorbs left-circularly polarized light differently from right-circularly polarized light. CD spectroscopy is an important tool for studying the structure of chiral molecules and can help provide useful information about the conformation and handedness of the constituent molecules and their interactions with solvents, typically measured in solution.
In recent years, solid chemistry has been vigorously developed as a leading field of chemical research. Solid state spectroscopy provides valuable information about the structure of the solid phase and the properties of supramolecules, which cannot be obtained from the solution phase. However, it has been very difficult to measure the chirality of solids so far using commercial CD spectrometers. The CD spectrum is necessarily accompanied by artifacts (anisotropic signals related to the linear polarization component) which originate on the one hand from the macroscopic anisotropy of the sample, which is characteristic of solids, and on the other hand from the non-ideal properties of the polarization modulation instrument. To obtain a true CD spectrum, we must remove artifacts from the observed CD spectrum.
Conventional CD spectrometers are typically polarization modulated using a photoelastic modulator, which inevitably results in anisotropic signals associated with the linearly polarized components, such as Linear Dichroism (LD) and Linear Birefringence (LB) artifacts, during the modulation process. In practice, the LD and LB signals are 2 to 3 orders of magnitude greater than the CD and CB signals, so we have difficulty in measuring the true CD spectrum of a typical solid sample.
Disclosure of Invention
The invention aims to solve the problem that an artifact signal exists when the existing commercial CD spectrometer measures the solid chirality, and provides a circular dichroism microscopic imaging system based on discrete modulation.
The invention relates to a circular dichroism microscopic imaging system based on discrete modulation, which comprises a linear light source, an 1/2 wave plate, a polaroid, a first reflector, a first beam shifter, an optical chopper, a second beam shifter, a 1/4 wave plate, a first objective and a sample objective table, wherein the linear light source is arranged on the linear light source;
linearly polarized light emitted by a linear light source is incident to an 1/2 wave plate, the polarization direction of the linearly polarized light is adjusted by a 1/2 wave plate and then is incident to a polarizing plate, the polarizing plate converts the incident light into 45-degree linearly polarized light, the 45-degree linearly polarized light is reflected by a first reflector and then is incident to a first light beam shifter, the first light beam shifter separates the 45-degree linearly polarized light into two linearly polarized light beams with the same intensity and mutually vertical polarization directions, the two linearly polarized light with the same intensity and mutually vertical polarization directions are emitted to a second light beam shifter in a periodically alternating conversion manner after passing through an optical chopper, the second light beam shifter synthesizes the two linearly polarized light in a periodically alternating manner into one discrete linearly polarized light, the discrete linearly polarized light is converted into discrete circularly polarized light by a 1/4 wave plate, and the discrete circularly polarized light is focused by a first objective lens and then is irradiated onto a sample to be measured, the sample to be detected is positioned on a glass slide, and the glass slide is fixed on a sample objective table; and obtaining an optical image of the sample to be detected on the glass slide.
Further, the invention also comprises a white light source, a computer, a nanometer moving platform, a second reflector and a camera;
when the white light optical imaging is carried out, a white light source is adopted to emit white light to be incident on the sample irradiated by the scattered circularly polarized light through the first objective lens, the second reflector reflects the white light passing through the sample glass slide to a camera of the camera, and an optical image of the sample is obtained;
the sample stage is fixed on the nanometer moving platform, the computer receives the optical image of the sample obtained by the camera, and the position of the sample in the optical image is adjusted by controlling the nanometer moving platform.
Furthermore, the invention also comprises a photoelectric detector, a phase-locked amplifier and a controller;
the photoelectric detector is used for detecting left-handed circularly polarized light and right-handed circularly polarized light absorbed by a sample to be detected, converting an optical signal into an electric signal and transmitting the converted electric signal to the signal input end of the phase-locked amplifier;
the controller is used for controlling the rotation of blades of the optical chopper, along with the rotation of the blades, two beams of linearly polarized light with the same intensity and mutually perpendicular polarization directions are periodically and alternately emitted to realize discrete modulation, the reference signal output end of the controller is connected with the reference signal input end of the phase-locked amplifier, the phase-locked amplifier demodulates the received signals and transmits the demodulated signals to the computer.
Further, in the present invention, the polarizing plate is a 45-degree polarizing plate.
Further, in the present invention, the first beam shifter separates the 45-degree polarized light into two linearly polarized lights having horizontal and vertical polarization directions.
Further, the invention also comprises a second objective, wherein the second objective is used for focusing the transmitted light passing through the sample to be detected and then emitting the focused transmitted light to the second mirror.
Further, the invention also comprises a white light source which is arranged in front of the first objective lens.
Furthermore, the invention also comprises a convex lens and a pinhole, wherein the pinhole is placed at the focus of the convex lens and attached to the front side of the acquisition surface of the photoelectric detector.
The invention obtains linearly polarized light with alternating vertical and horizontal polarization directions by utilizing the beam shifter and the chopper to carry out discrete modulation, obtains pure left-handed circularly polarized light and pure right-handed circularly polarized light by adopting the 1/4 wave plate, irradiates the pure left-handed circularly polarized light and the pure right-handed circularly polarized light on a sample, forms CD signals by different absorption of the sample on the left-handed circularly polarized light and the right-handed circularly polarized light, obtains pure circularly polarized light by carrying out circular polarization modulation, does not participate in modulation by a linear polarization component, and fundamentally prevents the generation of artifact signals.
Drawings
FIG. 1 is a schematic diagram of a camera for capturing images under a white light source according to the system of the present invention;
fig. 2 is a schematic structural diagram of the system of the present invention when a photoelectric detector is used to collect images.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
It should be noted that the embodiments and features of the embodiments may be combined with each other without conflict.
The first embodiment is as follows: the following describes the present embodiment with reference to fig. 1, and the present embodiment describes a circular dichroism microscopy imaging system based on discrete modulation, which includes a linear light source 1, an 1/2 wave plate 2, a polarizer 3, a first mirror 4, a first beam shifter 5, an optical chopper 6, a second beam shifter 7, a 1/4 wave plate 8, a first objective lens 10 and a sample stage 12;
linearly polarized light emitted by a linear light source 1 is incident to an 1/2 wave plate 2, the polarization direction of the linearly polarized light is adjusted by a 1/2 wave plate 2 and then is incident to a polarizing plate 3, the polarizing plate 3 converts the incident light into 45-degree linearly polarized light, the 45-degree linearly polarized light is reflected by a first reflecting mirror 4 and then is incident to a first light beam shifter 5, the first light beam shifter 5 separates the 45-degree linearly polarized light into two linearly polarized light beams with the same intensity and mutually perpendicular polarization directions, the two linearly polarized light beams with the same intensity and mutually perpendicular polarization directions are emitted to a second light beam shifter 7 in a periodic alternating manner after passing through an optical chopper 6, the two linearly polarized light beams with the same intensity and mutually perpendicular polarization directions are synthesized into one discrete linearly polarized light beam by the second light beam shifter 7, the discrete linearly polarized light beam is converted into discrete circularly polarized light by a 1/4 wave plate 8, and the discrete circularly polarized light is focused by a first objective lens 10 and then is irradiated onto a sample to be detected, the sample to be detected is positioned on a glass slide, and the glass slide is fixed on a sample object stage 12; and obtaining an optical image of the sample to be detected on the glass slide.
Further, as described with reference to fig. 1, the present embodiment further includes a white light source 9, a computer 19, a nano-scale moving platform 11, a second reflector 14, and a camera 18;
when the white light optical imaging is carried out, a white light source 9 is adopted to emit white light to be incident on the sample irradiated by the scattered circularly polarized light through a first objective lens 10, and a second reflector 14 reflects the white light passing through the sample glass slide to a camera of a camera 18 to obtain an optical image of the sample;
the sample stage 12 is fixed on the nano moving platform 11, the computer 19 receives the optical image of the sample obtained by the camera 18, and the position of the sample in the optical image is adjusted by controlling the nano moving platform.
The second embodiment is as follows: the present embodiment is described below with reference to fig. 2, and the present embodiment differs from the first embodiment in that it further includes a photodetector 17, a lock-in amplifier 20, and a controller 21;
the photoelectric detector 17 is used for detecting left-handed circularly polarized light and right-handed circularly polarized light absorbed by a sample to be detected, converting an optical signal into an electrical signal, and transmitting the converted electrical signal to a signal input end of the phase-locked amplifier 20;
the controller 21 is used for controlling the rotation of the blades of the optical chopper 6, along with the rotation of the blades, two beams of linearly polarized light with the same intensity and mutually perpendicular polarization directions periodically and alternately emit out to realize discrete modulation, the reference signal output end of the controller 21 is connected with the reference signal input end of the phase-locked amplifier 20, the phase-locked amplifier 20 demodulates the received signals, and transmits the demodulated signals to the computer 19.
In this embodiment, the controller 21 is configured to control the rotation of the blades of the optical chopper 6, and along with the rotation of the blades, two beams of linearly polarized light with the same intensity and mutually perpendicular polarization directions are periodically and alternately emitted to realize discrete modulation, a reference signal output end of the controller 21 is connected to a reference signal input end of the lock-in amplifier 20, and the lock-in amplifier 20 demodulates the received signal and transmits the demodulated signal to the computer 19. At this time, the camera does not acquire a signal and is in an off state.
Further, in the first and second embodiments, the polarizing plate 3 is a 45-degree polarizing plate.
Further, in the first embodiment and the second embodiment, the first beam shifter 5 separates the 45-degree polarized light into two linearly polarized lights having horizontal and vertical polarization directions.
When the embodiment is used, the removable white light source and the second reflector are used for imaging according to actual conditions, and then the removable white light source and the second reflector are removed to collect and receive optical signals passing through a sample by the photoelectric detector 17.
Further, in the first embodiment and the second embodiment, the second mirror 13 is further included, and the second mirror 13 is configured to focus the transmitted light passing through the sample to be measured and emit the focused transmitted light to the second mirror 14.
Further, in the first embodiment and the second embodiment, a white light source 9 is further included, and the white light source 9 is placed in front of the first objective lens 10.
Further, in the first embodiment and the second embodiment, the optical device further includes a convex lens 15 and a pinhole 16, and the pinhole 16 is placed at the focus of the convex lens 15 and attached to the front side of the collecting surface of the photodetector.
In this embodiment, the first objective lens 10, the second objective lens 13, the convex lens 15 and the pinhole 16 are used to form a confocal spot on the sample, so as to effectively prevent the impurity signal, reduce the intensity of the background signal, and improve the spatial resolution of the system.
The embodiment of the invention comprises the following steps:
a helium-neon laser is adopted as a light source of the embodiment, monochromatic light with the wavelength of 633nm emitted by the helium-neon laser passes through an 1/2 wave plate and a 45-degree polarizing plate, the function of the helium-neon laser is to adjust the polarization direction and change incident light into linearly polarized light in the 45-degree direction, the obtained linearly polarized light in the 45-degree direction is obtained, and the linearly polarized light in the 45-degree direction is reflected to a subsequent optical system through a first reflecting mirror; linearly polarized light reflected by the first reflector is separated into two beams of linearly polarized light (in the horizontal direction and the vertical direction) with the same intensity and mutually vertical polarization directions by the first beam shifter; the optical chopper periodically and alternately emits two linearly polarized lights with the same intensity and mutually vertical polarization directions to a second light beam shifter, and the optical chopper has 30 groove blades and the duty ratio is 25 percent; the second beam shifter combines two beams of linearly polarized light which are periodically and alternately incident into one beam of linearly polarized light which is subjected to discrete polarization modulation; then the circularly polarized light is converted into circularly polarized light modulated by discrete polarization after passing through 1/4 wave plates.
The nano moving platform controls the sample stage to move, so that a white light optical image and a laser scanning image of the sample can be obtained; wherein the removable white light source and the second reflector are used only when an optical image is obtained on the sample, and the white light source and the second reflector can be added in the light path to reflect the white light to the camera through the second reflector to obtain the optical image of the sample; when the white light source and the second reflector are removed, discretely modulated circularly polarized light emitted by the 1/4 wave plate is focused by the first objective lens (20x, n.a. ═ 0.4) and then irradiated onto a sample, the sample is positioned on the glass slide and fixed on the sample stage, and the nano moving platform is used for controlling the sample stage to move, so that an optical image of the sample under the white light source and a scanning image under the laser can be obtained.
When the white light source is removed, monochromatic light emitted from the second objective lens (50x, n.a. ═ 0.75) passes through a convex lens with a focal length of 100mm, and the light is converged at the focal point, and then a pinhole with a size of 100 μm is arranged at the focal point of the convex lens, and finally reaches a photodetector (here, the photodetector uses an avalanche photodiode APD). The nano moving platform used in the system is an XY piezoelectric ceramic nano positioner of P-733.2CD, the moving range is 100 x 100 μm, and the moving range is controlled by an E761 digital piezoelectric controller. Meanwhile, a LabVIEW program capable of positioning the nano mobile platform is written, so that when the sample is scanned and imaged, a scanning point can be positioned at the center of the scanning platform or other determined positions, and the appearance of the sample can be scanned.
When the nano moving platform controls the sample to move, the discretely modulated circular dichroism microscopic imaging system detects the left circularly polarized light and the right circularly polarized light absorbed by the sample to be detected at the avalanche photodiode, converts an optical signal into an electric signal and transmits the converted electric signal to the signal input end of the phase-locked amplifier SR 830; the optical chopper is placed in a light path, an output signal of the controller controls the rotating blade to rotate, two linearly polarized light beams with the same intensity and mutually vertical polarization directions periodically and alternately pass along with the rotation of the blade to realize discrete modulation, a reference signal output end of the controller is connected with a reference signal input end of a phase-locked amplifier, the phase-locked amplifier demodulates received signals and transmits the demodulated signals to a computer, and the computer is used for collecting and storing the received and demodulated signals.
The former first beam shifter, the second beam shifter, the 1/4 wave plate, the nanometer moving platform, the chopper controller, the camera for obtaining the optical image of the sample and the lock-in amplifier are all controlled by a computer, thereby improving the control precision of the system. And the camera is used for receiving the white light to obtain an optical image of the sample, and the optical image of the sample obtained by shooting can be transmitted to the electronic terminal.
Finally, the signals are demodulated by using a phase-locked amplifier, and a required CD image is obtained under the combined action of the chopper and the 1/4 wave plate. When the spatial resolution is high enough, the laser is applied to a single point on the sample, and the corresponding CD signal is obtained by controlling 1/4 wave plate (rotating the wave plate).
Although the invention herein has been described with reference to particular embodiments, it is to be understood that these embodiments are merely illustrative of the principles and applications of the present invention. It is therefore to be understood that numerous modifications may be made to the illustrative embodiments and that other arrangements may be devised without departing from the spirit and scope of the present invention as defined by the appended claims. It should be understood that features described in different dependent claims and herein may be combined in ways different from those described in the original claims. It is also to be understood that features described in connection with individual embodiments may be used in other described embodiments.
Claims (8)
1. The circular dichroism microscopic imaging system based on discrete modulation is characterized by comprising a linear light source (1), an 1/2 wave plate (2), a polarizing plate (3), a first reflecting mirror (4), a first beam shifter (5), an optical chopper (6), a second beam shifter (7), a 1/4 wave plate (8), a first objective lens (10) and a sample objective table (12);
linearly polarized light emitted by a linear light source (1) is incident to an 1/2 wave plate (2), the polarization direction of the linearly polarized light is adjusted by a 1/2 wave plate (2) and then is incident to a polarizing plate (3), the polarizing plate (3) converts the incident light into 45-degree linearly polarized light, the 45-degree linearly polarized light is reflected by a first reflecting mirror (4) and then is incident to a first light beam shifter (5), the first light beam shifter (5) separates the 45-degree linearly polarized light into two linearly polarized light beams with the same intensity and mutually vertical polarization directions, the two linearly polarized light beams with the same intensity and mutually vertical polarization directions are subjected to periodic alternating transformation after passing through an optical chopper (6) and then are emitted to a second light beam shifter (7), the two linearly polarized light beams which are in periodic alternation are synthesized into one beam of discrete linearly polarized light by the second light beam shifter (7), and the discrete linearly polarized light is converted into discrete circularly polarized light by a 1/4 wave, the scattered circularly polarized light is focused by a first objective lens (10) and then irradiated onto a sample to be detected, the sample to be detected is positioned on a glass slide, and the glass slide is fixed on a sample object stage (12); and obtaining an optical image of the sample to be detected on the glass slide.
2. The discrete modulation-based circular dichroism microscopy imaging system of claim 1, further comprising a white light source (9), a computer (19), a nano-moving platform (11), a second mirror (14), and a camera (18);
when the white light optical imaging is carried out, a white light source (9) is adopted to emit white light to be incident on the sample irradiated by the scattered circularly polarized light through a first objective lens (10), and a second reflector (14) reflects the white light passing through the sample glass slide to a camera of a camera (18) to obtain an optical image of the sample;
the sample object stage (12) is fixed on the nano moving platform (11), the computer (19) receives the sample optical image obtained by the camera (18), and the position of the sample in the optical image is adjusted by controlling the nano moving platform.
3. The discrete modulation-based circular dichroism microscopy imaging system of claim 1, further comprising a photodetector (17), a lock-in amplifier (20), and a controller (21);
the photoelectric detector (17) is used for detecting left-handed circularly polarized light and right-handed circularly polarized light absorbed by a sample to be detected, converting an optical signal into an electric signal and transmitting the converted electric signal to a signal input end of the phase-locked amplifier (20);
the controller (21) is used for controlling the rotation of blades of the optical chopper (6), along with the rotation of the blades, two beams of linearly polarized light with the same intensity and mutually perpendicular polarization directions periodically and alternately emit out to realize discrete modulation, the reference signal output end of the controller (21) is connected with the reference signal input end of the phase-locked amplifier (20), the phase-locked amplifier (20) demodulates received signals, and the demodulated signals are transmitted to the computer (19).
4. Circular dichroism microscopy imaging system based on discrete modulation according to claim 1, 2 or 3, characterized in that the polarizer (3) is a 45 degree oriented polarizer.
5. Circular dichroism microscopy imaging system based on discrete modulation according to claim 1, 2 or 3, characterized in that the first beam shifter (5) separates the 45 degree polarized light into two linearly polarized light beams with horizontal and vertical polarization directions.
6. The circular dichroism microscopy imaging system based on discrete modulation as claimed in claim 1, 2 or 3, characterized in that the system further comprises a second objective (13), wherein the second objective (13) is used for focusing and emitting the transmitted light passing through the sample to be measured to the second mirror (14).
7. The discrete modulation-based circular dichroism microscopy imaging system according to claim 1, characterized in that the white light source (9) is placed in front of the first objective (10).
8. The discrete modulation-based circular dichroism microscopy imaging system according to claim 3, characterized in that, further comprises a convex lens (15) and a pinhole (16), the pinhole (16) is placed at the focus of the convex lens (15) and is attached to the front side of the photodetector acquisition surface.
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| CN116953460A (en) * | 2023-04-03 | 2023-10-27 | 中国矿业大学 | Method for measuring circular dichroism photocurrent microscopic imaging of micro-nano device |
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