WO2017161535A1

WO2017161535A1 - Fluorescent scattering optical imaging system and method

Info

Publication number: WO2017161535A1
Application number: PCT/CN2016/077225
Authority: WO
Inventors: 童潇; 陈昳丽; 付楠; 朱艳春; 李荣茂; 余绍德; 陈鸣闽; 谢耀钦
Original assignee: 中国科学院深圳先进技术研究院
Priority date: 2016-03-24
Filing date: 2016-03-24
Publication date: 2017-09-28
Also published as: CN105873501A; CN105873501B

Abstract

A fluorescent scattering optical imaging system and method. The system comprises a laser (101), a micro-displacement table (102), an object table (103), at least one planar reflective mirror (104), a filter (105), a CCD camera (106) and a processor (107). An optical fiber head of the laser (101) is placed on the micro-displacement table (102). A reflecting surface of the planar reflective mirror (104) faces an object to be tested on the object table (103). The micro-displacement table (102) and the CCD camera (106) are electrically connected with the processor (107). The micro-displacement table (102) moves within a two-dimensional plane according to control signals of the processor (107). The laser (101) scans an area to be tested of the object to be tested which contains a fluorescent material to excite fluorescence. The CCD camera (106) is used for obtaining fluorescent images and laser images directly or on the basis of the reflection of the planar reflective mirror (104). The processor (107) is used for obtaining position information of the CCD camera (106), position information of the optical fiber head, position information of the planar reflective mirror (104), CT images or MRI images, fluorescent images and laser images of the object to be tested, so as to generate a three-dimensional fluorescent image of the area to be tested. The present invention can improve imaging quality and streamline system structure.

Description

Fluorescence scattering optical imaging system and method

Technical field

The present invention relates to the field of medical imaging technology, and in particular, to a fluorescence scattering optical imaging system and method.

Background technique

Fluorescence Diffuse Optical Tomography (FDOT) works by implanting tumors and corresponding targeted fluorescent reagents in small animals in advance, using a laser to scan in a certain plane in the area where small animals are located. Fluorescent reagents It is excited by laser, emits near-infrared light, and then obtains a picture of the excitation light through the detector. Finally, the position and distribution of the tumor in the animal body are determined by three-dimensional reconstruction. Compared with Magnetic Resonance Imaging (MRI), Computed Tomography (CT), and Positron Emission Tomography (PET), FDOT imaging has low cost, easy operation, no radiation, etc. Advantages, often used for small animal living imaging.

Existing studies have shown that FDOT/PET dual-modality imaging combined with FDOT and PET can provide information on the different physiological processes of experimental objects. This combination can help to increase the molecular level information provided at a given point in time. Comparing the characteristics of the two modal imaging systems, the PET system gantry records the acquired signals from all possible angles. However, this FDOT/PET dual-modality imaging system is complicated in structure and high in cost.

In the existing single-angle imaging FDOT system, a CCD (Charge-coupled Device) camera is fixed on the top of the object to be tested, and the excitation light source performs plane scanning on the opposite side of the CCD camera, so the existing common FDOT system A single angle imaging acquisition structure. Compared with PET, the single-angle imaging FDOT system has fewer acquisition modes, less information acquisition, longer acquisition period, and more difficult reconstruction. In addition, the single-angle acquisition of the image makes the FDOT imaging inferior to the longitudinal tomographic imaging along the line of the light source-CCD camera. Under normal circumstances, the geometry of the image acquisition system will in turn affect the quality of the reconstruction of the output image.

Summary of the invention

The invention provides a fluorescence scattering optical imaging system and method for improving the quality of fluorescence scattering optical tomography and shortening the imaging time.

The invention provides a fluorescence scattering optical imaging system, comprising: a laser, a micro-displacement stage, a stage, at least one plane mirror, a filter, a CCD camera and a processor; the fiber head of the laser is mounted on the micro a reflecting surface of the plane mirror facing the object to be tested on the stage; the micro-displacement stage and the CCD The camera is respectively electrically connected to the processor; the micro-displacement station is configured to move in a set plane area below the stage according to a control signal of the processor; the laser is used for scanning a built-in fluorescent substance The area to be tested of the object to be tested is used to excite fluorescence; the CCD camera is configured to obtain a fluorescence image and a laser image from above the stage, and the obtaining manner comprises: directly collecting from the object to be tested and reflecting on the plane based on the object Mirror reflection acquisition; the processor is configured to acquire position information of the CCD camera, position information of the fiber tip, position information of the plane mirror, CT image or MRI image of the object to be tested, the fluorescence image and the laser image, and This generates a three-dimensional fluorescence image of the region to be tested.

In one embodiment, an edge of the planar mirror is attached to the stage.

In one embodiment, the system includes two of the planar mirrors; two sides of each of the planar mirrors that are in contact with the stage are parallel to each other, and two of the planar mirrors and the carrier The angle of the object is the same.

In one embodiment, the filter comprises a fluorescent filter for filtering fluorescence and a laser filter for filtering laser light; the fluorescent filter is a 488 nm narrow band pass filter, the laser The filter is a long pass filter of 600 nm or more.

The invention also provides a fluorescence scattering optical imaging method, comprising: a micro-displacement stage driving a fiber head of a laser mounted thereon according to a control signal of a processor to move in a set plane region below the stage; the laser is to be tested The two-dimensional laser scanning of the object to be tested is performed to induce fluorescence of the fluorescent substance in the area to be measured; the CCD camera collects the composite fluorescent image and the composite laser image from above the stage, and the collection method includes: directly from the test Collecting and collecting based on the reflection of the plane mirror; the processor acquires position information of the CCD camera, position information of the fiber tip, position information of the plane mirror, CT image or MRI image of the object to be tested, and the composite A fluorescent image and a composite laser image are used to generate a three-dimensional fluorescence image of the region to be tested.

In one embodiment, the processor acquires position information of the CCD camera, position information of the fiber tip, position information of the plane mirror, CT image or MRI image of the object to be tested, the composite fluorescence image, and the composite laser image, and thereby Generating the three-dimensional fluorescence image of the area to be tested, comprising: trimming the composite laser image and the composite fluorescence image into a plurality of single laser images and a plurality of single fluorescent images respectively; according to the position information of the optical fiber head Position information of the CCD camera, position information of the plane mirror, CT or MRI image of the object to be tested, the single laser image, and the single fluorescence image are generated by the three-dimensional reconstruction software Three-dimensional fluorescence image of the area.

In one embodiment, the method further includes: setting a fluorescent filter in front of the CCD camera to filter out fluorescence emitted by the fluorescent substance; and a CCD camera acquiring a composite laser image from above the stage, including: the CCD The camera directly collects the laser light emitted by the laser fiber head and passes through the object to be tested, generates a first laser image, and simultaneously collects laser light that passes through the object to be tested and is reflected by the plane mirror to generate a second A laser image, the first laser image and the second laser image constitute the composite laser image.

In one embodiment, the method further includes: setting a laser filter in front of the CCD camera to filter out laser light emitted by the laser fiber tip; and collecting, by the CCD camera, a composite fluorescent image from above the stage, comprising: The CCD camera directly collects fluorescence emitted by the fluorescent substance in the area to be detected, generates a first fluorescent image, and simultaneously collects fluorescence emitted by the fluorescent substance in the area to be measured and reflected by the planar mirror to generate a second fluorescent image, the first fluorescent image and the second fluorescent image forming the composite fluorescent image.

In one embodiment, according to the position information of the fiber head, the position information of the CCD camera, the position information of the plane mirror, the CT or MRI image of the object to be tested, the single laser image and the The three-dimensional fluorescence image is generated by the three-dimensional reconstruction software, and the CT image or the MRI image is meshed by the volume mesh generation software to generate the body of the to-be-tested area. Surface mesh data; calculating position information of the CCD camera image in the plane mirror by using a specular reflection principle according to position information of the CCD camera and position information of the plane mirror; and positioning the fiber head Information, location information of the CCD camera, position information of the CCD camera image, the single laser image, the single fluorescence image, and the body surface mesh data are input to the three-dimensional reconstruction software, and are calculated The three-dimensional fluorescence image is obtained.

In one embodiment, the position information of the optical fiber head, the position information of the CCD camera, the position information of the CCD camera image, the single laser image, the single fluorescent image, and the body surface network Inputting the data into the three-dimensional reconstruction software, calculating the three-dimensional fluorescence image, comprising: scaling and matching the laser image and the fluorescence image to an actual size of the to-be-measured area; Laser image, zoom-matched fluorescent image, position information of the fiber tip, position information of the CCD camera, position information of the CCD camera image, and the body surface mesh data are input to the three-dimensional reconstruction In the software, the three-dimensional fluorescence image is calculated.

In one embodiment, one side of the planar mirror is attached to the stage; the composite laser image and the composite fluorescent image are respectively cut into a plurality of single laser images and a plurality of single fluorescent images, including : cutting the composite laser image into a plurality of the single laser images along an intersection of a plane of the plane mirror and a plane of the stage; along a plane of the plane mirror and the carrier An intersection line of the plane of the stage cuts the composite fluorescent image into a plurality of said single fluorescent images.

The fluorescence scattering optical imaging system and method of the embodiment of the invention can reflect the laser and the fluorescence from the object to be tested by using the plane mirror to reflect the laser and the fluorescence, thereby obtaining a richer object to be tested. The two-dimensional fluorescence image and the two-dimensional laser image information can improve the image reconstruction accuracy and improve the intensity of the reconstructed signal, and can obtain a three-dimensional fluorescence image with higher imaging quality than the existing single-angle FDOT system. The imaging system of the embodiment of the invention can realize multi-angle shooting and multi-angle imaging with only one real CCD camera Compared with the FDOT system, the system has the advantages of low equipment cost, and the real CCD camera and the at least one CCD camera image the laser image and the fluorescence image of the object to be tested at the same time, and the imaging system of the present invention has a faster imaging speed.

DRAWINGS

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the embodiments or the description of the prior art will be briefly described below. Obviously, the drawings in the following description are only It is a certain embodiment of the present invention, and other drawings can be obtained from those skilled in the art without any creative work. In the drawing:

1 is a schematic structural view of a fluorescence scattering optical imaging system according to an embodiment of the present invention;

2 is a schematic view showing imaging of a CCD camera in a plane mirror according to an embodiment of the present invention;

3 is a schematic diagram of a composite fluorescent image and a cut generated by the fluorescence scattering optical imaging system shown in FIG. 1;

4 is a schematic view showing the position setting of a plane mirror according to an embodiment of the present invention;

FIG. 5 is a schematic flow chart of a fluorescence scattering optical imaging method according to an embodiment of the present invention; FIG.

6 is a schematic flow chart of a method for imaging a three-dimensional fluorescence image according to an embodiment of the present invention;

7 is a flow chart showing a method of cropping a composite image into a single image according to an embodiment of the present invention;

8 is a schematic flow chart of a method for imaging a three-dimensional fluorescence image according to an embodiment of the present invention;

9 is a flow chart showing a method of performing three-dimensional fluorescence image imaging in an embodiment of the present invention.

detailed description

The embodiments of the present invention will be further described in detail below with reference to the accompanying drawings. The illustrative embodiments of the present invention and the description thereof are intended to explain the present invention, but are not intended to limit the invention.

The existing single-angle imaging FDOT system has the advantages of low cost and no radiation, but has the disadvantages of less acquisition mode, less information collection, difficulty in reconstruction, and longitudinal deterioration of quality. In order to take advantage of the advantages of the current FDOT system, the inventors have considered the influence of the geometry of the existing FDOT system on the quality of its output image, and through creative labor, improved the geometric structure of the existing single-angle imaging FDOT system to ensure imaging. Improve image quality while speed.

1 is a schematic view showing the structure of a fluorescence scattering optical imaging system according to an embodiment of the present invention. As shown in FIG. 1, a fluorescence scattering optical imaging system according to an embodiment of the present invention may include a laser 101, a micro-displacement stage 102, a stage 103, at least one planar mirror 104, a filter 105, a CCD camera 106, and a processor. 107. Fiber head of laser 101 It is mounted on the micro-stage 102. The reflecting surface of the plane mirror 104 faces the object to be tested on the stage 103. The micro-displacement stage 102 and the CCD camera 103 are electrically connected to the processor 107, respectively.

The laser 101 is used to scan a region to be tested of a test substance having a built-in fluorescent substance to excite fluorescence. The test object may be a living small animal, and the test area may be a tissue or an organ of a small animal, such as a tumor area. The laser light emitted by the laser 101 can be a near-infrared laser. As shown in FIG. 1, the laser 101 emits laser light from the bottom to the top, and the laser induces fluorescence of the fluorescent substance in the object to be tested on the stage 103, and the laser light and the fluorescence can be received by the CCD camera 106. In other embodiments, the laser 101 can illuminate the object to be tested from top to bottom, and accordingly, the CCD camera 106 can receive laser and fluorescence under the object to be tested.

The micro-displacement stage 102 is configured to move within a set plane area below the stage 103 according to a control signal of the processor 107. Under the driving of the micro-stage 102, for example, the fiber head is clamped on the micro-displacement stage, and the fiber head of the laser 101 can be moved along the set path in the xy plane for two-dimensional laser scanning. For example, the position of the laser scanning may be a position moving every other set distance along the x-axis, moving a total of N times, moving one position every other set distance along the y-axis, and moving a total of N times to form a laser (N+ 1) *(N+1) array. For example, the laser scanning may be centered on a certain set point, moved one position at a set angle along the circumferential direction, and moved M times to form a laser array.

Since the above-mentioned laser and fluorescent light are usually present at the same time, when the laser image is separately collected or the fluorescence image is separately collected, it is necessary to filter the unnecessary light by the filter 105 and then collect it by the CCD camera 106. The filter 105 may include a fluorescence filter for filtering out fluorescence and a laser filter for filtering laser light. The fluorescent filter can be a 488 nm narrow band pass filter, which can only acquire 488 nm light from the CCD camera 106, and is suitable for collecting laser light of the corresponding wavelength. The laser filter can be a long pass filter of 600 nm or more, and only the CCD camera 106 can collect light of 600 nm or more, and since the object to be tested can generally emit fluorescence of 600 to 700 nm under the excitation of the 488 nm laser, the The long pass filter of 600 nm or more can collect fluorescence well.

The CCD camera 106 is configured to acquire a fluorescence image and a laser image from above the stage 103. The acquisition method includes: directly collecting from the object to be tested and collecting the reflection based on the plane mirror 104. Wherein, when the fluorescence image and the laser image are directly collected from the object to be tested, the laser and the fluorescence do not pass through the plane mirror 104, specifically, the laser light passing through the object to be tested directly enters the CCD camera 106, and is controlled by the object to be tested. The fluorescence emitted by the fluorescent material directly enters the CCD camera 106. The fundamental difference between the two acquisition methods is that the path from the laser and the fluorescence that is directly emitted from the object to be tested to the CCD camera 106 is not changed, so the present invention is not limited to directly collecting images from the object to be tested. There are other elements between the object to be tested and the CCD camera 106 that do not change the optical path.

In one embodiment, the CCD camera 106 can be an Electro-Multiplying CCD (EMCCD) camera or a liquid-cooled CCD, which can have a more image acquisition effect.

The processor 107 is configured to acquire position information of the CCD camera 106, position information of the fiber head of the laser 101, position information of the plane mirror 104, CT image or MRI image of the object to be tested on the stage 103, fluorescence image, and laser image. And generating a three-dimensional fluorescence image of the above-mentioned test area. The processor 107 may be various devices capable of calculating the above-described three-dimensional fluorescent image based on the input information described above, such as a computer. The position information of the plane mirror 104 can be manually input to the processor 107 by hand. The position information of the CCD camera image in the plane mirror 104 can be calculated by using the position information of the plane mirror 104 and the position information of the CCD camera 106. The position information of the CCD camera image can be used to calculate the object to be tested. Three-dimensional fluorescence image of the area to be tested. The CT image or the MRI image of the object to be tested on the stage 103 may be a three-dimensional image of the area to be tested of the object to be tested, and may be acquired in advance by a corresponding device.

2 is a schematic view of a CCD camera imaged in a plane mirror in accordance with an embodiment of the present invention. As shown in FIG. 2, the planar mirror 104 is used to reflect laser light and fluorescence to a real CCD camera 106 to form

CCD camera images

106a, 106b, and a CCD camera image from a different angle than that of the real CCD camera 106. The shooting angles of 106a, 106b capture two images of a two-dimensional laser image and a two-dimensional fluorescent image of the area to be tested. As is clear from Fig. 2, the CCD camera 106 can be a CCD camera image in each of the plane mirrors 104. In a fluorescence scattering optical imaging system provided with a plurality of planar mirrors 104, a plurality of CCD camera images can be correspondingly formed, and each CCD camera image collects fluorescence and laser angles from the region to be tested of the object to be tested on the stage 103. Differently, the two-dimensional fluorescence image and the two-dimensional laser image of the to-be-measured area photographed from at least two different angles can be obtained by the above-described CCD camera 106 and each CCD camera image.

As shown in Figures 1 and 2, two planar mirrors 104 are provided in the system. This configuration is equivalent to three CCD cameras of the same optical characteristics (the CCD camera 106 on the top of the object and the virtual CCD on the left side of the object). The camera 106b and the right virtual CCD camera 106a) simultaneously record fluorescent signals emitted within the object at three locations. Compared with the existing multi-angle FDOT system, the multi-angle imaging system formed by the plane mirror greatly simplifies the system structure, and since it retains the original single-angle system structure, it can still be applied to the conventional FDOT image acquisition scheme. wide range.

In the fluorescence scattering optical imaging system of the embodiment of the present invention, there may be only one real CCD camera 106, so that each two-dimensional fluorescent image will be on a composite fluorescent image, and each two-dimensional laser image will be in a composite laser image. on.

3 is a schematic diagram of a composite fluorescent image and a cut generated by the fluorescence scattering optical imaging system shown in FIG. 1. As shown in FIG. 1 and FIG. 3, in the fluorescence scattering optical imaging system of this embodiment, two planar mirrors 104 are disposed, and two plane mirrors 106 disposed on both sides of the mouse can be used to obtain one frame. A composite fluorescence image 200 of a two-dimensional fluorescence image. The composite fluorescent image 200 includes a fluorescent image 201 captured on the top of the mouse by the CCD camera 106 on the top of the mouse (subject) on the stage 103, and a CCD camera in the two planar mirrors 106 of the CCD camera 106. The

fluorescent images

202 and 203 on the side of the mouse were photographed separately.

4 is a schematic view showing the position setting of a plane mirror in an embodiment of the present invention. As shown in FIG. 4, the plane mirror 104 can be disposed in various ways, and can be located at various positions, and its angle with the horizontal plane (for example, the angles α, β) can be various angles, and the plane mirror 104 and the to-be-tested The distance L between the objects may be various values as long as the plane mirror 104 can reflect the laser light and fluorescence from the region to be tested of the object to be tested to the CCD camera 106.

In one embodiment, the plane mirror 104 can be disposed on the stage 103. Specifically, an edge of the plane mirror 104 can be attached to the stage 103. Thus, the plane mirror 104 is conveniently disposed. And an image of the side of the object to be tested can be collected.

In one embodiment, the fluorescence scattering optical imaging system can include two planar mirrors 104. The sides of the two planar mirrors 104 that are respectively attached to the stage 103 may be parallel to each other, that is, the intersection of the planes of the two plane mirrors 104 and the plane of the stage 103 may be parallel to each other. The angle between the two plane mirrors 104 and the stage 103 can be the same, that is, in the case where the plane mirror 104 faces the object to be tested, the plane of the two plane mirrors 104 and the plane of the stage 103 are located. The angle of the angle can be the same. As shown in FIG. 1, the angles α and β of the two plane mirrors 104 and the stage 103 may be the same. In other embodiments, the angles α and β may be different. The magnitudes of the angles α and β can be various values. The inventors have found that the angles of the angles α and β are 30° by calculating the angle of the CCD camera image in the plane mirror 104. In the range of ~40°, in this way, more image information of the object to be tested can be obtained.

In the fluorescence scattering optical imaging system of the embodiment of the invention, the laser and the fluorescence are reflected by the plane mirror, and the laser and the fluorescence from the object to be tested can be collected from the angle different from the real CCD camera, thereby obtaining a richer object to be tested. Dimensional fluorescence image and two-dimensional laser image information, according to which a three-dimensional fluorescence image with higher imaging quality than the existing single-angle FDOT system can be obtained. The imaging system of the embodiment of the invention can realize multi-angle shooting only by a real CCD camera, and has the advantage of low equipment cost compared with the FDOT system of multi-angle imaging, and the real CCD camera and at least one CCD camera image are simultaneously taken. The laser image and the fluorescence image of the object are measured, and the imaging system of the present invention has a faster imaging speed.

Based on the same inventive concept as the fluorescence scattering optical imaging system shown in FIG. 1, the embodiment of the present application also provides a fluorescence scattering optical imaging method, as described in the following embodiments. Since the principle of solving the problem by the fluorescence scattering optical imaging method is similar to that of the fluorescence scattering optical imaging system, the implementation of the fluorescence scattering optical imaging method can be referred to the implementation of the fluorescent scattering optical imaging system, and the repetition will not be repeated.

FIG. 5 is a schematic flow chart of a fluorescence scattering optical imaging method according to an embodiment of the present invention. As shown in FIG. 5, the fluorescence scattering optical imaging method of the embodiment of the present invention may include the following steps:

S310: The micro-displacement station drives the fiber head of the laser mounted thereon to move in a set plane region below the stage according to a control signal of the processor;

S320: performing a two-dimensional laser scanning on the to-be-measured area of the object to be measured by the laser to induce fluorescence of the fluorescent substance in the area to be tested;

S330: The CCD camera collects the composite fluorescent image and the composite laser image from above the above-mentioned stage, and the collecting manner includes: directly collecting from the object to be tested and collecting based on the reflection of the plane mirror;

S340: The processor acquires position information of the CCD camera, position information of the fiber head, position information of the plane mirror, CT image or MRI image of the object to be tested, the composite fluorescent image and the composite laser image, and generates the above-mentioned test Three-dimensional fluorescence image of the area.

In the above step S310, under the driving of the micro-stage, for example, the fiber head is clamped on the micro-displacement stage, and the fiber head of the laser can be moved along the set path in the set two-dimensional plane for two-dimensional laser scanning. . For example, as described above, the position of the laser scanning may be a position moving every other set distance along the x-axis, moving a total of N times, moving one position every other set distance along the y-axis, and moving a total of N times to form a laser. (N+1)*(N+1) array. For example, the laser scanning may be centered on a certain set point, moved one position at a set angle along the circumferential direction, and moved M times to form a laser array. The laser emitted by the laser can be a near-infrared laser.

In the above step S320, the object to be tested may be a living small animal, and the test area may be a tissue or an organ of a small animal, such as a tumor area. For example, a laser having a wavelength of 488 nm is used to illuminate the analyte, and the fluorescent material in the analyte can emit fluorescence of 600 to 700 nm.

In the above step S330, when the composite fluorescence image and the composite laser image are directly collected from the object to be tested, the laser and the fluorescence are not reflected by the plane mirror, specifically, the laser light passing through the object to be tested directly enters the CCD camera. The fluorescence emitted by the fluorescent substance in the analyte directly enters the CCD camera. The fundamental difference between the two acquisition methods is that the path from the laser and the fluorescence that is emitted from the object to be tested directly to the CCD camera is not changed. The composite laser image may include multiple two-dimensional laser images, and the composite fluorescent image may include multiple two-dimensional fluorescence images. As shown in FIG. 3, the composite fluorescent image 200 includes three two-dimensional

fluorescent images

201, 202, and 203. .

In the above step S340, the CT image or the MRI image of the object to be tested may be a three-dimensional image of the to-be-measured area of the object to be tested, and may be acquired in advance by a corresponding device.

In the embodiment of the present invention, the image acquired by the FDOT imaging system may be merged with the CT image or merged with the MRI image, so that the functional image provided by the FDOT can be compared and processed in the process of comparing with the CT image or the MRI image. The spatial structure provided by the CT image or the MRI image is more intuitively and accurately presented.

The fluorescence scattering optical imaging system of the embodiment of the present invention can reflect the laser and the fluorescence from the object to be tested by using a plane mirror to reflect the laser and the fluorescence, thereby obtaining a richer two-dimensionality of the object to be tested. The fluorescence image and the two-dimensional laser image information can be used to obtain a three-dimensional fluorescence image with higher imaging quality than the existing single-angle FDOT system.

6 is a flow chart showing a method of imaging a three-dimensional fluorescence image according to an embodiment of the present invention. As shown in FIG. 6, in the above step S340, the processor acquires position information of the CCD camera, position information of the optical fiber head, position information of the plane mirror, CT image or MRI image of the object to be tested, the composite fluorescent image and the composite The laser image, and the method for generating the three-dimensional fluorescent image of the above-mentioned test area, may include the following steps:

S341: The composite laser image and the composite fluorescent image are respectively cut into a plurality of single laser images and a plurality of single fluorescent images;

S342: Perform three-dimensional reconstruction according to the position information of the fiber head, the position information of the CCD camera, the position information of the plane mirror, the CT or MRI image of the object to be tested, the single laser image, and the single fluorescence image. The software generates a three-dimensional fluorescence image of the above-mentioned area to be tested.

In the above step S341, under the driving of the micro-displacement stage, when the laser scans the object to be tested from different positions, the CCD image is collected and saved as a laser-excited laser image sequence and a fluorescence image sequence, and the composite laser image and the composite laser image are When the composite fluorescent image is separately cut into a plurality of single laser images and a plurality of single fluorescent images, the laser image sequence and the fluorescent image sequence are trimmed. The single laser image and the single fluorescent image may be images of the area to be tested.

In this embodiment, the composite laser image and the composite fluorescent image are respectively cut into a plurality of single laser images and a plurality of single fluorescent images, and only the image of the area to be tested can be retained, and when the three-dimensional fluorescent image is generated, only for the test The reconstruction of the image of the area eliminates the need to reconstruct the non-target imaging area, which helps to save the reconstruction time of the three-dimensional fluorescence image, thereby improving the imaging speed.

7 is a flow chart showing a method of cropping a composite image into a single image in an embodiment of the present invention. One side of the plane mirror is attached to the stage, and as shown in FIG. 7, in the above step S341, the above The method for respectively cutting a composite laser image and a composite fluorescent image into a plurality of single laser images and a plurality of single fluorescent images may include the following steps:

S3411: cutting the composite laser image into a plurality of the single laser images along a line connecting the plane of the plane mirror and the plane of the stage;

S3412: cutting the composite fluorescent image into a plurality of the single fluorescent images along a line connecting the plane of the plane mirror and the plane of the stage.

In this embodiment, the composite image (composite laser image, composite fluorescent image) is cut into a plurality of single images (single laser image, single fluorescent image) along the intersection of the plane of the plane mirror and the plane of the stage. . In one embodiment, as shown in FIG. 3, the composite fluorescent image 200 is cropped into three single

fluorescent images

201, 202, and 203 along the

intersection lines

2021, 2031. The clipping area 2022 of the single fluorescent image 202 and the clipping area 2032 of the single fluorescent image 203 can be selected according to requirements. For example, the

square areas

2022 and 2032 as shown in FIG. 3 can be cut out, and then the object to be tested is obtained. The image area corresponding to the area to be tested, or the two-dimensional fluorescence image and the laser image of the area to be tested are cut out only by one process, and can be selected as needed.

In this embodiment, by cutting the composite image into a single image along the intersection of the plane of the plane mirror and the plane of the stage, a complete single image can be easily obtained, and the clipping error is not easy to occur.

In other embodiments, the composite laser image and the composite fluorescence image obtained by the method shown in FIG. 7 can be further cut into the laser of the region to be tested before being used to generate the three-dimensional fluorescence image. The image is cut into a fluorescent image of the area to be tested to reduce the data processing amount of the processor and improve the imaging time of the three-dimensional fluorescent image.

In one embodiment, a fluorescent filter is disposed in front of the CCD camera to filter out fluorescence emitted by the fluorescent substance. For example, the analyte is excited at 488 nm, and emits fluorescence of 600-700 nm. The first filter is 488 nm. The narrow bandpass (passband 10nm) filter only allows the CCD to capture 488nm light. At this time, in the above step S330, the method for the CCD camera to collect the composite laser image from above the above-mentioned stage may include the following steps:

S331: The CCD camera directly collects the laser light emitted from the laser fiber head and passes through the object to be tested, generates a first laser image, and simultaneously collects the laser light that passes through the object to be tested and is reflected by the plane mirror to generate a second In the laser image, the first laser image and the second laser image constitute the composite laser image.

In one embodiment, a laser filter is disposed in front of the CCD camera to filter out the laser light emitted by the laser fiber head. Specifically, for example, the object to be tested is excited at 488 nm, emitting fluorescence of 600-700 nm, and changing the filter. The filter is a long pass filter of 600 nm or more, and the CCD camera collects a fluorescent image. At this time, the method for the CCD camera to collect the composite fluorescent image from above the above stage may include the following steps:

S332: The CCD camera directly collects fluorescence emitted by the fluorescent substance in the area to be detected, generates a first fluorescent image, and simultaneously collects fluorescence emitted by the fluorescent substance in the area to be measured and reflected by the planar mirror, to generate In the second fluorescent image, the first fluorescent image and the second fluorescent image constitute the composite fluorescent image.

FIG. 8 is a flow chart showing a method of imaging a three-dimensional fluorescence image according to an embodiment of the present invention. As shown in FIG. 8, in the above step S342, based on the position information of the optical fiber head, the position information of the CCD camera, the position information of the plane mirror, the CT or MRI image of the object to be tested, and the single laser image. And the method for generating the three-dimensional fluorescence image of the to-be-tested area by using the three-dimensional reconstruction software, and the method includes the following steps:

S3421: meshing the CT image or the MRI image by using a volume mesh generation software to generate body surface mesh data of the to-be-tested area;

S3422: calculating, according to the position information of the CCD camera and the position information of the plane mirror, the position information of the CCD camera image in the plane mirror by using a specular reflection principle;

S3423: input position information of the optical fiber head, position information of the CCD camera, position information of the CCD camera image, the single laser image, the single fluorescence image, and the body surface mesh data into the three-dimensional reconstruction software. The above three-dimensional fluorescence image is calculated.

In the above step S3421, the volume mesh generation software may be a plurality of different meshing software, such as iso2mesh software. In the above step S3421, the three-dimensional reconstruction software may be a plurality of different reconstruction software, such as toast software.

When reconstructing a three-dimensional fluorescence image by using three-dimensional reconstruction software, firstly, a finite element reference iterative algorithm is used to generate a fluorescence image, wherein the reference iterative algorithm is based on a coupling diffusion equation describing excitation light propagation and scattered light in the region to be tested:

Where r is the positional variable, φ _x is the photon density of the excitation light x, and φ _m is the photon density of the scattered light m,

Is the diffusion coefficient of the excitation light x,

Is the diffusion coefficient,

Is the absorption coefficient of the excitation light x,

Is the absorption coefficient of the scattered light m,

Is the scattering coefficient of the attenuation of the excitation light x,

Is the attenuation coefficient of the scattered light m, a is the boundary correlation coefficient of the internal reflection, S _x (r)=S ₀ δ(rr ₀ ) is the excitation source term of the excitation point x point source, and S ₀ represents the intensity of the point source , δ(rr ₀ ) is the Dirac-delta function centered on the point source of position r ₀ , and η is the edge diffusion coefficient.

In order to excite the absorption coefficient of light

Or the absorption coefficient of scattered light

The equation matrix obtained by the finite element discrete relation is shown in equations (1) and (2), and further obtains a series of equations for solving the inverse problem:

[A _x,m ]{φ _x,m }={b _x,m }, (3)

Wherein, the parameters of the matrix [A _x,m ] and the terms in the column vector {b _x,m } can represent a Lagrangian basis function by a set of spatial variations; J _x,m is derived from φ _x,m The object observes the Jacobian matrix of χ at each boundary; Δχ is the optical and fluorescence property distribution update vector; I is the identity matrix; λ can be a scale or diagonal matrix;

Is the transposed matrix of the matrix J _x,m ; χ is the fluorescence characteristic distribution vector, expressing D _x ,

or

φ _x,m is the photon density of the excitation light x or the scattered light m;

Is the observed photon density of the excitation light x or the scattered light m;

It is the calculated photon density of the excitation light x or the scattered light m. The laser image and the fluorescence image are formed by iteratively solved by the equations (3) to (5), and the optical fluorescence characteristic distribution is updated from the properties of the properties.

9 is a flow chart showing a method of performing three-dimensional fluorescence image imaging in an embodiment of the present invention. As shown in FIG. 9, in the above step S3423, the position information of the optical fiber head, the position information of the CCD camera, the position information of the CCD camera image, the single laser image, the single fluorescent image, and the body surface The grid data is input into the above three-dimensional reconstruction software, and the method for calculating the three-dimensional fluorescence image is calculated, which may include the following steps:

S34231: scaling and matching the above laser image and the fluorescent image to an actual size of the to-be-measured area;

S34232: input, in the above, the laser image after the scaling matching, the fluorescence image after the scaling matching, the position information of the optical fiber head, the position information of the CCD camera, the position information of the CCD camera image, and the body surface mesh data. In the three-dimensional reconstruction software, the above three-dimensional fluorescence image is calculated.

In one embodiment, the fluorescence scattering optical imaging method comprises the steps of:

1) placing an object to be reconstructed on the stage, the object containing a fluorescent substance capable of exciting fluorescence under the corresponding excitation light source, adjusting the field of view of the CCD camera to cover the entire object;

2) Place a filter under the CCD camera to filter out the fluorescence emitted by the object. For example, an object is excited at a 488 nm laser to emit fluorescence at a wavelength of 600-700 nm. The first filter placed was a 488 nm narrow bandpass (passband 10 nm) filter that only allowed the CCD to acquire 488 nm light.

3) operating the two-dimensional micro-displacement table to move according to the programmed program in the processor, that is, moving the position of the laser fiber head to emit laser light at different positions;

4) simultaneously acquiring CCD images at different scanning positions of the laser, and storing the image sequences excited by the laser;

5) Change the filter. For example, when the object is excited by a 488 nm laser and emits fluorescence of 600-700 nm, the filter with a long pass of 600 nm or more can be changed, so that the CCD collects a fluorescent image;

6) The composite laser image and the composite fluorescent image captured by the CCD camera are cut, and the image of the object on the front side and the sides is cut, and the intersection of the plane mirror and the horizontal plane can be cut for the dividing line.

7) scanning a CT image of the object, and generating a surface volume mesh data mesh of the object by software (for example, iso2mesh) of the generated mesh;

8) The laser image and the fluorescence image, as well as the CCD camera (the image of the real CCD camera and CCD camera in the plane mirror) and the position information of the laser source, the mesh information of the object as the input information of the reconstruction software, through the 3D reconstruction software (eg toast), generating three-dimensional data containing a fluorescence distribution.

In step 8), the fluorescent image includes a fluorescent image of the front side of the segmented object and a fluorescent image of the side surface. The specific trimming step may include: selecting a region of interest ROI region of the object, such as a mouse lung (for example, a size of 1.2 cm*1.2 cm), and combining the laser image and the fluorescence image after the selected ROI region The actual dimensions of the objects are matched, then the ROI area is clipped and the Jacobian matrix is generated by the 3D reconstruction software. The position information includes: position information of the CCD (for example, the position information of the CCD camera is "12 14 40 0 0 -1", which sequentially represents the coordinates of the x-axis, the y-axis, and the z-axis, and the unit is mm, and 0 0 -1 indicates that the CCD is Under the acquisition), the position information of the laser source (for example, the laser source position information is, for example, "12 14 - 5 0 0 1", which sequentially represents the coordinates of the x-axis, the y-axis, and the z-axis, and the unit is mm, and 0 0 1 represents the laser source. Upward excitation, mesh information (ie, three-dimensional coordinates of the body surface mesh, for example, 25.595, 60.6565, 20.565, representing the coordinates of the x-axis, y-axis, and z-axis, respectively), through three-dimensional reconstruction software (for example, calling toastMapSolToMesh, toastSolutionMask, IWT2_P0, FDOTAdj0p or tostQvec, etc.) to reconstruct the distribution information of the fluorescent substance in the object, that is, the three-dimensional fluorescence image.

The fluorescence scattering optical imaging method of the embodiment of the invention can reflect the laser and the fluorescence from the object to be tested from the angle different from the real CCD camera by reflecting the laser and the fluorescence by the plane mirror, thereby obtaining a richer object to be tested. The dimensional fluorescence image and the two-dimensional laser image information can improve the image reconstruction accuracy and improve the intensity of the reconstructed signal, and obtain a three-dimensional fluorescence image with higher imaging quality than the existing single-angle FDOT system. The imaging system of the embodiment of the invention can realize multi-angle shooting only by a real CCD camera, and has the advantage of low equipment cost compared with the FDOT system of multi-angle imaging, and the real CCD camera and at least one CCD camera image are simultaneously taken. The laser image and the fluorescence image of the object are measured, and the imaging system of the present invention has a faster imaging speed.

In the description of the specification, reference is made to the terms "one embodiment", "one embodiment", "some embodiments", "for example", "example", "specific example", or "some examples" Combined with the real Particular features, structures, materials or features described in the examples or examples are included in at least one embodiment or example of the invention. In the present specification, the schematic representation of the above terms does not necessarily mean the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in a suitable manner in any one or more embodiments or examples.

Those skilled in the art will appreciate that embodiments of the present invention can be provided as a method, system, or computer program product. Accordingly, the present invention may take the form of an entirely hardware embodiment, an entirely software embodiment, or a combination of software and hardware. Moreover, the invention can take the form of a computer program product embodied on one or more computer-usable storage media (including but not limited to disk storage, CD-ROM, optical storage, etc.) including computer usable program code.

The present invention has been described with reference to flowchart illustrations and/or block diagrams of methods, apparatus (system), and computer program products according to embodiments of the invention. It will be understood that each flow and/or block of the flowchart illustrations and/or FIG. These computer program instructions can be provided to a processor of a general purpose computer, special purpose computer, embedded processor, or other programmable data processing device to produce a machine for the execution of instructions for execution by a processor of a computer or other programmable data processing device. Means for implementing the functions specified in one or more of the flow or in a block or blocks of the flow chart.

The computer program instructions can also be stored in a computer readable memory that can direct a computer or other programmable data processing device to operate in a particular manner, such that the instructions stored in the computer readable memory produce an article of manufacture comprising the instruction device. The apparatus implements the functions specified in one or more blocks of a flow or a flow and/or block diagram of the flowchart.

These computer program instructions can also be loaded onto a computer or other programmable data processing device such that a series of operational steps are performed on a computer or other programmable device to produce computer-implemented processing for execution on a computer or other programmable device. The instructions provide steps for implementing the functions specified in one or more of the flow or in a block or blocks of a flow diagram.

The above described specific embodiments of the present invention are further described in detail, and are intended to be illustrative of the embodiments of the present invention. All modifications, equivalent substitutions, improvements, etc., made within the spirit and scope of the invention are intended to be included within the scope of the invention.

Claims

A fluorescence scattering optical imaging system, comprising: a laser, a micro-displacement stage, a stage, at least one planar mirror, a filter, a CCD camera, and a processor;

a fiber head of the laser is mounted on the micro-displacement stage; a reflective surface of the planar mirror faces an object to be tested on the stage; the micro-displacement stage and the CCD camera respectively perform the processing Electrical connection

The micro-displacement stage is configured to move in a set plane area below the stage according to a control signal of the processor;

The laser is configured to scan a region to be tested of the analyte with a built-in fluorescent substance to excite fluorescence;

The CCD camera is configured to acquire a fluorescence image and a laser image from above the stage, and the acquiring manner comprises: directly collecting from the object to be tested and collecting the reflection based on the plane mirror;

The processor is configured to acquire position information of a CCD camera, position information of a fiber tip, position information of a plane mirror, a CT image or an MRI image of the object to be tested, the fluorescence image, and a laser image, and generate the Three-dimensional fluorescence image of the survey area.
A fluorescence scattering optical imaging system according to claim 1 wherein an edge of said planar mirror is attached to said stage.
A fluorescence scattering optical imaging system according to claim 2, wherein said system comprises two of said planar mirrors; and said two planar mirrors are each parallel to the side of said stage, And the angle between the two plane mirrors and the stage is the same.
The fluorescence scattering optical imaging system according to claim 1, wherein said filter comprises a fluorescence filter for filtering fluorescence and a laser filter for filtering laser light; said fluorescent filter The sheet is a 488 nm narrow band pass filter, and the laser filter is a long pass filter of 600 nm or more.
A fluorescence scattering optical imaging method, comprising:

The micro-displacement stage drives the fiber head of the laser mounted thereon to move in a set plane region below the stage according to a control signal of the processor;

Performing a two-dimensional laser scanning on the area to be tested of the laser to induce fluorescence of the fluorescent substance in the area to be tested;

The CCD camera collects the composite fluorescent image and the composite laser image from above the stage, and the collecting manner includes: directly collecting from the object to be tested and collecting based on the reflection of the plane mirror;

The processor acquires position information of the CCD camera, position information of the fiber tip, position information of the plane mirror, CT image or MRI image of the object to be tested, the composite fluorescence image, and the composite laser image, and generates the to-be-tested Three-dimensional fluorescence image of the area.
The fluorescence scattering optical imaging method according to claim 5, wherein the processor acquires position information of the CCD camera, position information of the fiber tip, position information of the plane mirror, CT image or MRI image of the object to be tested, and The composite fluorescent image and the composite laser image are used to generate a three-dimensional fluorescent image of the region to be tested, including:

The composite laser image and the composite fluorescent image are respectively cut into a plurality of single laser images and a plurality of single fluorescent images;

And according to position information of the fiber head, position information of the CCD camera, position information of the plane mirror, CT or MRI image of the object to be tested, the single laser image, and the single fluorescence image And generating a three-dimensional fluorescence image of the to-be-measured area by using three-dimensional reconstruction software.
The fluorescence scattering optical imaging method according to claim 5, further comprising: arranging a fluorescent filter in front of the CCD camera to filter out fluorescence emitted by the fluorescent substance; and a CCD camera from above the stage Collect composite laser images, including:

The CCD camera directly collects laser light emitted by the laser fiber head and passes through the object to be tested, generates a first laser image, and simultaneously collects laser light that passes through the object to be tested and is reflected by the plane mirror. A second laser image is generated, the first laser image and the second laser image constituting the composite laser image.
The fluorescence scattering optical imaging method according to claim 5, further comprising: providing a laser filter in front of the CCD camera to filter out laser light emitted from the laser fiber head; and a CCD camera from the stage Collect composite fluorescent images above, including:

The CCD camera directly collects fluorescence emitted by the fluorescent substance in the area to be detected, generates a first fluorescent image, and simultaneously collects fluorescence emitted by the fluorescent substance in the area to be measured and reflected by the planar mirror And generating a second fluorescent image, the first fluorescent image and the second fluorescent image forming the composite fluorescent image.
The fluorescence scattering optical imaging method according to claim 6, wherein the position information of the optical fiber head, the position information of the CCD camera, the position information of the planar mirror, and the CT of the object to be tested are Or the MRI image, the single laser image, and the single fluorescence image, and the three-dimensional fluorescence image of the to-be-measured area is generated by the three-dimensional reconstruction software, including:

The CT image or the MRI image is meshed by the volume mesh generation software to generate body surface mesh data of the to-be-tested area;

Calculating position information of the CCD camera image in the plane mirror by using a specular reflection principle according to position information of the CCD camera and position information of the plane mirror;

Inputting the position information of the optical fiber head, the position information of the CCD camera, the position information of the CCD camera image, the single laser image, the single fluorescent image, and the body surface mesh data to the In the three-dimensional reconstruction software, the three-dimensional fluorescence image is calculated.
The fluorescence scattering optical imaging method according to claim 9, wherein position information of the optical fiber head, position information of the CCD camera, position information of the CCD camera image, the single laser image, The single fluorescent image and the body surface mesh data are input into the three-dimensional reconstruction software, and the three-dimensional fluorescent image is calculated, including:

Scaling and matching the laser image and the fluorescent image to an actual size of the area to be tested;

And scaling the matched laser image, scaling the matched fluorescence image, position information of the fiber tip, position information of the CCD camera, position information of the CCD camera image, and the body surface mesh Data is input to the three-dimensional reconstruction software, and the three-dimensional fluorescence image is calculated.
The fluorescence scattering optical imaging method according to claim 6, wherein one side of the plane mirror is attached to the stage;

The composite laser image and the composite fluorescent image are respectively cut into a plurality of single laser images and a plurality of single fluorescent images, including:

Cutting the composite laser image into a plurality of the single laser images along an intersection of a plane of the plane mirror and a plane of the stage;

The composite fluorescent image is clipped into a plurality of the single fluorescent images along an intersection of a plane of the planar mirror and a plane of the stage.