CN206876950U - A kind of quick efficient self-adapted optical imagery compensation system based on interference enhancing - Google Patents

Abstract

The utility model discloses a fast and efficient adaptive optical imaging compensation system based on interference enhancement. The beam emitted by the laser is transmitted through the optical fiber and then enters the digital micromirror device through the collimating lens. There is a phase compensation module next to the digital micromirror device, and a beam splitter and a beam shrinker module are placed on the front side. The phase compensation module is located in the beam splitter. At the side of the side, the side reflected beam is reflected to the beam splitter after passing through the phase compensation module, and the positive side reflected beam is directly reflected to the beam splitter; the beam is reflected by the dichroic mirror and enters the microscope objective lens to focus through the scanning module, and the experimental sample The internally excited fluorescence passes through the microscope objective lens and the scanning module, and then is received by the light intensity detection module through the dichroic mirror. The utility model starts from the principle of optical interference and phase compensation, improves the adaptive focusing speed and focusing quality of the imaging beam deep inside the scattering medium, and provides a new technology for the field of non-invasive optogenetics and living deep high-resolution microscopic imaging system.

Description

A Fast and Efficient Adaptive Optical Imaging Compensation System Based on Interference Enhancement

technical field

The utility model belongs to the field of optogenetics and optical microscopic imaging, and particularly relates to a fast and efficient adaptive optical imaging compensation system based on interference enhancement, and is applied to non-invasive optogenetic light stimulation and deep penetrating optical microscopy imaging.

Background technique

In the field of biomedical optics, optical scattering is the main factor restricting the quality of optical imaging. Most optical techniques for deep tissue imaging (eg, confocal laser imaging, two-photon microscopy, and optical coherence tomography) primarily utilize non-scattered photons (ie, ballistic photons) for imaging. The number of ballistic photons decays exponentially with depth, thus limiting the optical focus to a depth of 1 mm.

Optogenetic technology requires photostimulation of specific neurons to study its neural circuit mechanism. However, the traditional optical fiber implantable optogenetics has serious damage to living organisms, which is not conducive to long-term research. The adaptive optics technology, which was previously applied in astronomy, provides new technical support for the realization of light stimulation and imaging of deep biological tissues.

The existing non-invasive adaptive optogenetics technology is based on the precise phase correction technology of adaptive optics, or the phase compensation based on coherent light adaptive technology. The precise phase correction technology divides the spatial light modulator into several partitions, changes the additional phase of the beam in the partitions at equal intervals, and detects the optimal beam focusing phase. Each partition is cyclically iterated in turn to obtain the final corrected phase, and phase compensation is performed on the incident beam to correct its distorted phase and form a good beam focus.

However, the above precise phase correction needs to consume a lot of time. In order to obtain better optical correction, it is necessary to divide more partitions and finer phase intervals. Optical corrections are time consuming due to the low image refresh rate of the spatial light modulator.

Coherent light adaptive technology is also to divide the spatial light modulator into several partitions, and use block deformable mirrors or digital micromirror devices to perform rapid intensity modulation on the beams incident on the spatial light modulator, so as to use the light intensity obtained by coherence of each beam Calculate the compensation phase required by the beam corresponding to different partitions, and then load the phase to the spatial light modulator for beam compensation.

Although the use of coherent light adaptive technology can effectively shorten the time for correcting the beam phase, it still cannot overcome the time consumed by loading the phase of the spatial light modulator for correction, which is not conducive to real-time optogenetic stimulation and high-resolution imaging in living organisms. Research has restricted the promotion of adaptive optogenetic technology.

In order to be able to extend non-invasive adaptive optogenetics to the research application of living biological tissues, it is necessary to achieve deeper optical stimulation with lower incident light power while increasing the focusing speed. This is also an urgent problem to be solved in the application of adaptive optics in biology.

Utility model content

In order to solve the problems in the background technology, the purpose of this utility model is to propose a fast and efficient adaptive optical imaging compensation system based on interference enhancement. The problem that the spatial light modulator takes a long time.

The utility model utilizes the fast refresh rate of the digital micromirror device, proceeds from the principle of optical interference, divides the light beam into several areas, learns from the fast compensation phase detection technology of coherent light adaptive technology, and distinguishes the interference effect of different phases on the focus center of the light beam , the beam partitions with interference constructive in the focus center will be reserved, and the beam partition with interference destructive in the focus center will be added with a phase difference π to make them meet the interference constructive conditions and be reintroduced into the optical path. By converting the beam components that are unfavorable to beam focusing into components that enhance beam focusing, the utilization rate of incident light is greatly improved, and the quality of beam focusing is improved. The utility model saves the time consumed by using the spatial light modulator to load the compensation phase, shortens the correction time of the adaptive light stimulation, and makes the incident light be compensated to the focus center to participate in the optical stimulation as much as possible, from the perspective of time and space Optimized for adaptive optogenetics.

In order to achieve the above object, the technical solution of the utility model comprises the following steps:

The system includes a laser, a first optical fiber, a collimating lens, a digital micromirror device, a phase compensation module, a beam splitter, a beam shrinking module, a scanning module, a dichroic mirror, a microscope objective lens, an experimental sample and a light intensity detection module. The first optical fiber and collimating lens are arranged behind the laser. The beam emitted by the laser is transmitted through the first optical fiber and then enters the digital micromirror device through the collimating lens. A phase compensation module is placed in front of the output end of the digital micromirror device. A beam splitter and a beam reduction module are installed at the exit end of the front side of the mirror device, and the phase compensation module is located at the side of the beam splitter. The side reflected beam of the digital micromirror device is reflected into the beam splitter after passing through the phase compensation module The front side reflection beam of the digital micromirror device is directly reflected into the beam splitter; a dichroic mirror is arranged in front of the beam reduction module; the beam is reflected by the dichroic mirror and then enters the microscope objective lens to focus through the scanning module, and the experimental sample is located in the microscope On the focal plane of the objective lens; the fluorescence excited in the experimental sample passes through the microscope objective lens and the scanning module, and then is received by the light intensity detection module through the dichroic mirror for light intensity detection.

The phase compensation module includes, but is not limited to, an optical delay line, a spatial light modulator, a phase retarder, and the like.

The beam shrinking module includes a front shrinking module lens and a rear shrinking module lens; the front shrinking module lens and the rear shrinking module lens are sequentially arranged in parallel on the front side of the beam splitter, and the light beams reflected by the digital micromirror device are sequentially passed through The beam splitter, the front reducer module lens, and the rear reducer module lens are incident on the oblique side of the dichroic mirror after being reduced in parallel.

The scanning module includes a front scanning vibrating mirror, a front beam collimating lens, a rear beam collimating lens, a rear scanning vibrating mirror, a front scanning module lens and a rear scanning module lens; the front scanning vibrating mirror, the front beam collimating lens, the rear The beam collimating lens, the rear scanning galvanometer, the front scanning module lens and the rear scanning module lens are sequentially arranged on the front side of the dichroic mirror, and the beam reflected by the dichroic mirror passes through the front scanning galvanometer, the front beam collimating lens, The rear beam collimating lens, the rear scanning galvanometer, the front scanning module lens and the rear scanning module lens are incident on the microscope objective lens.

The light intensity detection module includes an optical filter, a collimating focusing lens, a second optical fiber, a focusing lens and a photomultiplier tube; the optical filter, a collimating focusing lens, a second optical fiber, a focusing lens and a photomultiplier tube are arranged in sequence On the rear side of the dichroic mirror, the fluorescence emitted by the experimental sample passes through the microscope objective lens, the rear scanning module lens, the front scanning module lens, the rear scanning galvanometer, the rear beam collimating lens, the front beam collimating lens, and the front scanning galvanometer , a dichroic mirror, an optical filter, a collimating focusing lens, a second optical fiber and a focusing lens enter into a photomultiplier tube for light intensity detection.

The beneficial effects of the utility model are:

The utility model utilizes the digital micromirror device to realize fast and parallel self-adaptive beam focus compensation, utilizes the fast image refresh rate of the digital micromirror device, overcomes the problem of slow speed when using the spatial light modulator to perform phase correction in the past, and improves the beam Speed of focusing versus optical stimulation.

The utility model uses a digital micromirror device to perform self-adaptive optical focus compensation, replaces an expensive spatial light modulator, reduces experiment cost while ensuring beam focus, and is more conducive to the application of the system in research experiments.

Moreover, the utility model can be easily combined with various existing microscopic imaging technologies, and can be used in the research and development of non-invasive optogenetic light stimulation technology to realize synchronous light stimulation and microscopic imaging, which is beneficial to brain science research further development.

Description of drawings

Fig. 1 is the structural representation of the utility model system;

Fig. 2 is the scattered focus spot image when no digital micromirror device is judged in the embodiment;

Fig. 3 is a schematic diagram of the modulation frequency applied by the first part of the digital micromirror device in the embodiment;

4 is a schematic diagram of the modulation frequency applied by the second part of the digital micromirror device in the embodiment;

Fig. 5 is the partial fluorescent light intensity value after the intensity modulation that photomultiplier tube detects in the embodiment;

Fig. 6 is the partial frequency-amplitude diagram after the Fourier transform of the fluorescent light intensity value in the embodiment;

Fig. 7 is the compensation phase value of some partitions calculated from the fluorescent light intensity value in the embodiment;

Fig. 8 is the distribution map of the partitions required for compensation in the first part of the digital micromirror device partition in the embodiment;

Fig. 9 is the distribution diagram of the partitions required for compensation in the second partition of the digital micromirror device in the embodiment;

Fig. 10 is the partition phase distribution figure that needs to be compensated after the judgment of the digital micromirror device in the embodiment;

FIG. 11 is a focus spot image after phase compensation is performed on all subregions that need to be compensated in the embodiment.

detailed description

The following embodiments of fast and efficient adaptive optics imaging compensation based on interference enhancement can illustrate the utility model in more detail, but the utility model is not limited in any form.

As shown in Figure 1, the utility model system includes a laser 1, a first optical fiber 2, a collimating lens 3, a digital micromirror device 4, a phase compensation module 5, a beam splitter 6, a front shrinking module lens 7 and a rear shrinking beam Module lens 8, dichroic mirror 9, front scanning vibrating mirror 10, front beam collimating lens 11, rear beam collimating lens 12, rear scanning vibrating mirror 13, front scanning module lens 14 and rear scanning module lens 15, microscope Objective lens 16 , experimental sample 17 , optical filter 18 , collimator focusing lens 19 , second optical fiber 20 , focusing lens 21 and photomultiplier tube 22 .

The optical imaging compensation process of the present utility model is:

1) no experimental sample is placed at the focal plane of the objective lens, the beam is focused with a digital micromirror device without loading partitions, an ideal focused spot is obtained at the focal plane of the objective lens, and the focal center position O _f of the ideal focused spot is recorded;

Beam focusing is specifically that the laser emits a beam, which is reflected on the digital micromirror device after being collimated and expanded, and then focused by the objective lens.

2) Place the experimental sample at the focal plane of the objective lens, use a digital micromirror device without loading partitions to detect the light intensity, and record and obtain the light intensity value at the focus center position O _f ;

Light intensity detection is specifically that the laser beam emits a beam, which is reflected on the digital micromirror device after being collimated and expanded, and then focused on the sample through the objective lens. The sample contains a fluorescent material, which generates a distorted scattering spot in the sample and excites fluorescence. After the excited fluorescence is focused by the lens, the photomultiplier tube and the galvanometer cooperate to scan and detect to obtain the fluorescence image excited by the scattered spot, and record the light intensity value of the focus center position O _f .

3) Divide the reflective surface of the digital micromirror device into multiple areas, and divide all areas into two parts; specifically, divide the micromirror pixel elements of the digital micromirror device into n×n evenly, so that the quasi- The incident beam after direct beam expansion is divided into corresponding n×n beam units.

4) Perform light intensity detection in an intensity modulation manner for each area of each part, obtain a light intensity value corresponding to the recorded focus center position O _f of each part, and process to obtain the compensation phase value corresponding to each area in each part.

Each area in each part can be connected or disconnected, that is, a checkerboard-like equal division.

Specifically, multiple regions are divided into two parts with no common partition, and the appropriate modulation frequency range for each part is selected according to the Nyquist sampling law, and each partition in each part is set to have a different intensity modulation frequency , the intensity modulation frequencies corresponding to all partitions of each part can cover the modulation frequency range of this part.

Then, when the light intensity is detected, the reflective surfaces of each area in the first part are kept unchanged, and the reflective surfaces of each area in the second part are deflected back and forth at different frequencies at the same time, so that the beams generated by the reflective surfaces of each area are set at The frequency deflects back and forth between the positive side light path and the side light path. Different regions have different frequencies of back and forth deflection, and the beams of different partitions are modulated at different frequencies by continuous deflection back and forth for a long time, and the light intensity of the modulated beams is detected.

Finally, perform Fourier transform on the detected light intensity value and draw a frequency-amplitude diagram, and convert and calculate the compensation phase from the light intensity amplitude corresponding to the modulation frequency in different regions in the frequency-amplitude diagram, so as to obtain the second Compensation phase values required for different areas within the section.

On the contrary, keeping the reflective surfaces of each area in the second part unchanged, and deflecting the reflective surfaces of each area in the first part at different frequencies at the same time, the compensation phase required by different areas in the first part can be calculated.

5) Compare the compensation phase value of each area with π, and use the following method to obtain the judgment result:

If the compensation phase value is less than or equal to π, the area corresponding to the light intensity value is an area that does not require phase compensation;

If the compensation phase value is greater than π, the area corresponding to the light intensity value is the area requiring phase compensation;

6) Load the partition image adjusted according to the judgment result to the digital micromirror device. After the incident beam is collimated and expanded, the beam corresponding to the partition that needs to be compensated is reflected to the side optical path and makes its phase compensation and then returns to the optical path , phase compensation is to add a phase difference of π to the original phase of the beam. The light beams corresponding to the unchanged partitions are focused by the objective lens to form the final optical focus compensation spot in the experimental sample, and stimulate stronger fluorescence.

Adjusting the partition according to the judgment result specifically refers to: when passing through the digital micromirror device, after the judgment, the phase compensation area needs to adjust the reflection angle of the reflected beam generated after receiving the incident beam, so that the reflected beam goes to the side optical path for phase compensation and then returns to In the reflection optical path before the reflection angle is adjusted, at the same time, the reflection angle of the reflection beam is kept the same as the reflection angle of the reflection beam in the step 2) when the light intensity is detected in the area where phase compensation is not required after the judgment.

The fluorescent material includes fluorescent protein, fluorescent beads or fluorescent dyes.

The experimental samples are, but not limited to, living biological tissues, isolated biological tissues, agar blocks containing pellets, and the like.

Embodiment of the present utility model and concrete process thereof are as follows:

(1) The light beam emitted by the laser 1 passes through the first optical fiber 2 and the collimating lens 3 in sequence, and then irradiates the digital micromirror device 4 . When the digital micromirror device 4 is not loaded with an image and is not loaded with an experimental sample 17, the light beam is reflected through the beam splitter 6, the front shrinking module lens 7 and the rear shrinking module lens 8, and the side of the dichroic mirror 9 is irradiated. Reflection, the reflected light beam enters the microscope objective lens 16 after passing through the front scanning vibrating mirror 10, the front beam collimating lens 11, the rear beam collimating lens 12, the rear scanning vibrating mirror 13, the front scanning module lens 14 and the rear scanning module lens 15, and An ideal focused spot is formed at the focal plane, and the position of the center of the focused spot at the focal plane is O _f .

(2) When the experimental sample 17 is loaded and the digital micromirror device 4 is not loaded with an image, the light beam sent by the laser 1 passes through the first optical fiber 2 and the collimating lens 3 in sequence, and then irradiates on the digital micromirror device 4, and the light beam is reflected through the splitter. The beam device 6, the front beam shrinker module lens 7 and the rear beam shrinker module lens 8 are reflected on the side of the dichroic mirror 9, and the reflected beam passes through the front scanning galvanometer 10, the front beam collimator lens 11, and the rear beam collimator. The lens 12, the rear scanning galvanometer 13, the front scanning module lens 14 and the rear scanning module lens 15 enter the microscope objective lens 16, and form a distorted scattered focus spot at the inner focal plane of the experimental sample 17, and excite fluorescence.

(3) Fluorescence enters the microscope objective lens 16 from the experimental sample 17, and then passes through the rear scanning module lens 15, the front scanning module lens 14, the rear scanning galvanometer 13, the rear beam collimating lens 12, and the front beam collimating lens 11 , the front scanning galvanometer 10, the dichroic mirror 9, the optical filter 18, the focusing lens 19 and the second optical fiber 20, and the focusing lens 21 enter the photomultiplier tube 22 for light intensity detection, and obtain the judgment of the non-digital micromirror device by scanning The scattered and focused spot image at the time is shown in Fig. 2. Record the light intensity value at the position corresponding to O _f at this time. In this specific implementation case, due to the offset of the point with the strongest light intensity, the light intensity value at the corresponding position of O _f is 11.35.

(4) In this specific implementation case, the micromirror pixel element of the digital micromirror device is evenly divided into 1024 areas in the form of 32×32, so that the collimated and expanded incident beam is divided into corresponding 1024 beam units. The 1024 partitions are equally divided into two parts with no common partitions, and the appropriate modulation frequency is selected according to the Nyquist sampling law, so that each partition in each part has a different intensity modulation frequency. When the intensity modulation of the beam is performed, the side optical path does not change the phase of the beam and only acts as a beam blocking effect, keeping the partitions of the second part unchanged, and the micromirrors in each partition in the first part vibrate at the corresponding different frequencies at the same time, so that The light beam is deflected back and forth between the subsequent light path and the side light path at the corresponding frequency, so that the intensity modulation of the light beam in different partitions is performed at different frequencies. The fluorescence light intensity value at O _f was recorded in real time by the photomultiplier tube. In this specific implementation case, the 1024 partitions are equally divided as shown in Figure 3 and Figure 4; the maximum modulation frequency used is 204.8 Hz, and the frequency interval is 0.2 Hz; the intensity-modulated part detected by the photomultiplier tube The fluorescence intensity values are shown in Figure 5.

(5) Perform Fourier transform on the light intensity value obtained in step (4), and the frequency-amplitude diagram drawn after Fourier transform is shown in Figure 6, and the corresponding frequency coordinates are obtained on the frequency coordinates corresponding to the modulation frequencies of each partition. compensation phase. Keep the partitions whose phase value is less than or equal to π unchanged and mark the partitions whose phase value is greater than π to obtain the marked partition image. In this specific implementation case, the partial partition compensation phase values calculated from the fluorescent light intensity values are shown in Figure 7; the partition distribution diagram of the phase compensation required for the first part of the partition is shown in Figure 8; Figure 9 shows the distribution of partitions that require phase compensation.

(6) The marked partition image is loaded onto the digital micromirror device 4, and the incident light beam is irradiated on the digital micromirror device 4 after the first optical fiber 2 and the collimating lens 3, and the unchanged partitioned light beam is captured by the digital micromirror The device 4 reflects and reaches the beam splitter 6, and the beam corresponding to the marked partition is reflected to the phase compensation module 5 of the side optical path, so that the phase difference of π changes, and returns to the beam splitter 6 to correspond to the unchanged partition The light beam passes through the front shrinking module lens 7 and the rear shrinking module lens 8 together, and is reflected on the side of the dichroic mirror 9, and the reflected beam passes through the front scanning galvanometer 10, the front beam collimating lens 11, and the back beam collimation After the lens 12, the rear scanning galvanometer 13, the front scanning module lens 14 and the rear scanning module lens 15, enter the microscope objective lens 16, and form a final adaptive optics focus-compensated spot at the inner focal plane of the experimental sample 17, and excite Stronger fluorescence. The phase compensation module used in this specific implementation case is an optical delay line, and the phase distribution diagram of the partitions that need to be compensated after the digital micromirror device is judged is shown in FIG. 10 .

(7) Fluorescence excited after optical focus compensation is emitted from the experimental sample 17 into the microscope objective lens 16, and then passes through the rear scanning module lens 15, the front scanning module lens 14, the rear scanning galvanometer 13, and the rear beam collimating lens 12 , front beam collimator lens 11, front scanning vibrating mirror 10, dichroic mirror 9, optical filter 18, focusing lens 19 and second optical fiber 20, after focusing lens 21 enter photomultiplier tube 22, obtain digital micromirror by scanning The device 4 loads the focused spot image after compensation by the marked partition, as shown in FIG. 11 , and the light intensity value corresponding to the position O _f of the image is 1195 in this specific implementation case.

Claims (5)

1. A fast and efficient adaptive optical imaging compensation system based on interference enhancement, characterized in that: comprising laser (1), first optical fiber (2), collimating lens (3), digital micromirror device (4), phase Compensation module (5), beam splitter (6), beam reduction module, scanning module, dichroic mirror (9), microscope objective lens (16), experimental sample (17) and light intensity detection module; The first optical fiber ( 2) and the collimating lens (3) are arranged behind the laser (1), the laser emits a light beam that is transmitted through the first optical fiber (2) and then enters the digital micromirror device (4) through the collimating lens (3), and the digital micromirror A phase compensation module (5) is arranged at the outgoing end of the side of the device (4), a beam splitter (6) and a beam reduction module are arranged at the outgoing end of the digital micromirror device (4), and a dichroic mirror (9) is arranged at the front. ); the light beam is reflected by the dichroic mirror (9) and enters the microscopic objective lens (16) to focus through the scanning module, and the experimental sample (17) is located on the focal plane of the microscopic objective lens (16); Fluorescence is received by the light intensity detection module through the dichroic mirror (9) after passing through the microscope objective lens (16) and the scanning module. 2. A fast and efficient adaptive optical imaging compensation system based on interference enhancement according to claim 1, characterized in that: said phase compensation module (5) includes but not limited to optical delay line, spatial light modulator, Phase retarder. 3. A fast and efficient adaptive optics imaging compensation system based on interference enhancement according to claim 1, characterized in that: the beam shrinking module includes a front shrinking module lens (7) and a rear shrinking module lens ( 8); the front shrinking module lens (7) and the rear shrinking module lens (8) are arranged in parallel on the front side of the beam splitter (6) successively, and the light beam reflected by the digital micromirror device (4) passes through the beam splitter ( 6), the front shrinking module lens (7) and the rear shrinking module lens (8) are incident on the side of the dichroic mirror (9) after shrinking in parallel. 4. A fast and efficient adaptive optical imaging compensation system based on interference enhancement according to claim 1, characterized in that: said scanning module includes a front scanning galvanometer (10), a front beam collimating lens (11) , rear beam collimating lens (12), rear scanning galvanometer (13), front scanning module lens (14) and rear scanning module lens (15); front scanning galvanometer (10), front beam collimating lens (11) , the rear beam collimating lens (12), the rear scanning galvanometer (13), the front scanning module lens (14) and the rear scanning module lens (15) are sequentially arranged on the front side of the dichroic mirror (9), and the dichroic The beam reflected by the mirror (9) passes through the front scanning galvanometer (10), the front beam collimating lens (11), the rear beam collimating lens (12), the rear scanning galvanometer (13), and the front scanning module lens (14) in sequence And the rear scanning module lens (15) is incident to the microscope objective lens (16). 5. A fast and efficient adaptive optical imaging compensation system based on interference enhancement according to claim 1, characterized in that: the light intensity detection module includes an optical filter (18), a collimating focusing lens (19) , second optical fiber (20), focusing lens (21) and photomultiplier tube (22); optical filter (18), collimating focusing lens (19), second optical fiber (20), focusing lens (21) and photoelectric The multiplier tube (22) is sequentially arranged on the rear side of the dichroic mirror (9), and the fluorescence emitted by the experimental sample (17) passes through the microscope objective lens (16), the rear scanning module lens (15), and the front scanning module lens (14) successively. ), rear scanning galvanometer (13), rear beam collimating lens (12), front beam collimating lens (11), front scanning galvanometer (10), dichroic mirror (9), optical filter (18) , the collimating focusing lens (19), the second optical fiber (20) and the focusing lens (21) enter the photomultiplier tube (22) for light intensity detection.