CN116839500A - Imaging system for simultaneously extracting internal fluorescent molecular distribution and surface three-dimensional structure - Google Patents
Imaging system for simultaneously extracting internal fluorescent molecular distribution and surface three-dimensional structure Download PDFInfo
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- G01B—MEASURING LENGTH, THICKNESS OR SIMILAR LINEAR DIMENSIONS; MEASURING ANGLES; MEASURING AREAS; MEASURING IRREGULARITIES OF SURFACES OR CONTOURS
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Abstract
The invention relates to an imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure, which can simultaneously realize reflective line scanning fluorescent molecular tomography and an original data collection and reconstruction method of three-dimensional surface information by utilizing line structure light scanning. According to the system, only the existing basic elements of the fluorescent molecular tomography system are utilized, under a reflective acquisition mode, fluorescent picture data for reflective FMT fluorescent reconstruction and structured light picture data for surface information reconstruction are respectively acquired at the same time by utilizing a white light camera and a fluorescent camera, and based on the images acquired at the same time, three-dimensional surface information and three-dimensional distribution of fluorescence of an object are reconstructed, so that three-dimensional high-fidelity surface information and accurate fluorescent space-time distribution can be obtained, accurate real-time observation is more facilitated, and the final imaging result of the system has extremely high real-time performance and accuracy.
Description
Technical Field
The invention relates to a three-dimensional scanning technology, in particular to an imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure.
Background
Fluorescent molecular tomography (Fluorescence Molecular Tomography, FMT) as a macroscopic optical molecular imaging technique for visualizing the temporal-spatial distribution of fluorescence generated by specific fluorescent molecular probes in biological tissues has numerous advantages over other macroscopic imaging such as Magnetic Resonance Imaging (MRI) and Computed Tomography (CT), including non-invasive, high sensitivity, no ionizing radiation, low cost, and the like, and has great potential in the fields of new drug development, early diagnosis of diseases, surgical image navigation, and the like.
The fluorescent molecular tomography system is divided into a contact type system and a non-contact type system, wherein the non-contact type fluorescent molecular tomography system mainly comprises an excitation light source, a scanning device, an objective table, a fluorescent signal receiving device and a white light signal receiving device. The traditional non-contact system data acquisition process is to inject a targeted fluorescent reagent into a small animal body in advance, then perform point-by-point raster scanning on an imaging area by using an excitation light source and a scanning device, enable the reagent to emit fluorescence longer than the excitation light wavelength, and take an emitted light picture by using a fluorescence camera.
In a fluorescent molecular tomography reconstruction algorithm, three-dimensional surface information of an object is a key part of the object, and the addition of structural information in reconstruction is beneficial to improving the accuracy of image reconstruction; the lack of accurate structural references to well-registered contours or anatomical images due to the molecular signals in the FMT, the three-dimensional surface information also facilitates interpretation of reconstructed fluorescence or bioluminescence signals; in addition, the extraction of surface information can monitor structural changes of the tissue. Existing fluorescent molecular tomography systems with surface acquisition function are divided into two categories: one is to combine the system with other modes to obtain surface information, such as combining MRI, CT, etc.; another category is to use a separate surface information extraction module, such as a structured light depth camera, to obtain the surface information. Both types of systems present challenges in registration between different modalities, increased system complexity, and increased system costs.
The problems of the conventional non-contact system are mainly classified into the following points:
1. when data are collected, because point grid scanning is used for collecting fluorescent pictures, the collecting speed is slow, and in long time of collecting, the moving error of an object is difficult to control, so that the accuracy of reconstruction is reduced.
2. The surface information extraction and the fluorescence tomography data are acquired separately, and the organisms can move during the two acquisition, so that the surface information extraction is inaccurate, and the reconstruction accuracy is reduced.
3. The size of a focusing light spot used for scanning by most non-contact systems is more than 1mm, and the scanning precision is not improved.
Disclosure of Invention
Aiming at the problems of long acquisition time, unreliable data quality during long-time acquisition and lack of a surface information extraction device in the existing fluorescent molecular tomography system, the imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure is provided, and the method can simultaneously realize reflective line scanning fluorescent molecular tomography and an original data collection and reconstruction method of three-dimensional surface information by utilizing line structure light scanning. The system only utilizes the existing basic elements of the fluorescent molecular tomography system, and utilizes a white light camera and a fluorescent camera to simultaneously and respectively acquire fluorescent picture data for reflective FMT fluorescent reconstruction and structured light picture data for surface information reconstruction in a reflective acquisition mode, and reconstructs high-precision three-dimensional surface information of an object and three-dimensional distribution of fluorescence based on the simultaneously acquired pictures. The final imaging result of the system has extremely high real-time performance and accuracy.
The technical scheme of the invention is as follows: an imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure comprises a light source module, a scanning module, a detection module and a control module,
the light source module comprises a laser, a coupler, a multimode optical fiber and a collimator, wherein laser emitted by the laser is coupled into one end of the multimode optical fiber through the coupler, and then is input into the collimator from the other end of the optical fiber for collimation and then projected into a scanning vibrating mirror of the scanning module;
the scanning module comprises a scanning vibrating mirror, a scanning field lens and a reflecting mirror, wherein the scanning vibrating mirror focuses a collimated light beam into light spots with the size of less than 1 millimeter through the scanning field lens, the light spots are converted into light rays through the scanning vibrating mirror, and the linear illumination structure of the upper surface of a measured object on the imaging objective table is scanned strip by strip after passing through the reflecting mirror;
the detection module comprises a fluorescence camera and a white light camera, wherein the fluorescence camera and the white light camera are respectively arranged right above and at the side of the imaging object table, the fluorescence camera and the white light camera are at the same height, the fluorescence camera and the white light camera can both see the upper surface of a sample on the imaging object table in the visual field, the fluorescence camera collects fluorescence information of the sample reflected by excitation light or excited by transmission through a filter in a filter wheel, the white light camera directly collects the fluorescence information of the excitation light reflected by the excitation light incident sample, and therefore, a measured object is simultaneously photographed through the fluorescence camera and the white light camera, and fluorescence data reconstructed by fluorescence molecular tomography FMT fluorescent line scanning and the excitation light number used for surface line structural light reconstruction are obtainedAccording toAnd transmitting the data to a computer in the control module;
the control module comprises a computer and a USB wire, wherein the computer is respectively connected with the laser, the scanning galvanometer, the fluorescent camera and the white light camera through the USB wire, and transmits instructions to control data scanning and acquisition.
Preferably, the filter wheel is provided with band-pass filters with different center wavelengths, and the band-pass filters are arranged between the fluorescent camera and the measured object and filter light except the projection wavelength.
A method for simultaneously extracting internal fluorescent molecular distribution and obtaining reconstruction data by an imaging system of a surface three-dimensional structure includes the steps of firstly, placing a sample on an objective table, and shooting a bright field white light image of the sample by a fluorescent camera under a bright field condition; setting exposure time required by shooting, power of a laser and positions of a band-pass filter in a filter wheel for a white light camera and a fluorescent camera, and starting and setting a scanning mode of a scanning galvanometer as line scanning; and setting the number and the range of the scanning lines, pre-scanning to verify whether the scanning lines and the range meet expectations, resetting the number and the range of the scanning lines if the scanning lines and the range do not meet expectations, turning off bright field illumination if the scanning lines and the range do not meet expectations, simultaneously and respectively acquiring tomographic reconstruction data of line scanning and original data of surface reconstruction by using a fluorescent camera and a white light camera, turning off a laser after the simultaneous acquisition is finished, and respectively acquiring ambient background light by using the fluorescent camera and the white light camera under the conditions of same exposure time, the position of a band-pass filter and darkness when the reconstruction data are shot.
The method for reconstructing surface information of an imaging system by simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure comprises the steps of firstly extracting a light bar center pixel coordinate of line structure light in surface reconstruction original data collected by a white light camera, determining a light plane equation of laser by utilizing the center coordinate of a pixel and a space position of a vibrating mirror relative to the white light camera, then calculating a three-dimensional point cloud of a sample surface by utilizing the light plane equation and the internal parameters of the camera, and generating a grid mesh based on the three-dimensional point cloud.
The method for reconstructing internal fluorescent molecular distribution information of an imaging system by simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure comprises the steps of firstly guiding tomographic reconstruction fluorescent data of line scanning acquired by a fluorescent camera, generating grid mesh by utilizing surface information, setting reconstruction related parameters, generating a finite element grid and calculating a system matrix according to the finite element grid and the related parameters; then carrying out noise processing on the emission light image of the scanning module, and generating a weight matrix according to the processed tomographic reconstruction fluorescence data of the line scanning and a system matrix; and then selecting a corresponding parameter method to solve the inverse problem, and if the reconstructed result accords with the reconstruction expectation, ending the reconstruction, and if the reconstructed result does not accord with the reconstruction expectation, returning to the first step to acquire the data again.
The invention has the beneficial effects that: the invention simultaneously extracts the internal fluorescent molecular distribution and the imaging system of the surface three-dimensional structure, creates a line scanning fluorescent tomography system on the basis of the traditional point scanning fluorescent tomography system, and realizes faster reconstruction data acquisition and reconstruction results close to point scanning; the white light camera and the fluorescent camera are utilized to respectively acquire fluorescent picture data for reflective FMT fluorescent reconstruction and structured light picture data for surface information reconstruction, so that the acquired data quality is more reliable, the three-dimensional high-fidelity surface information and accurate fluorescent space-time distribution can be obtained, and the accurate real-time observation is more facilitated; the scanning light spot is focused to be less than 1mm, so that the scanning precision with higher resolution is realized.
Drawings
FIG. 1 is a schematic diagram of an imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure;
FIG. 2 is a flow chart of raw data acquisition of the system of the present invention;
FIG. 3 is a flow chart of FMT reconstruction of the system of the present invention;
FIG. 4 is a flow chart of the reconstruction of surface information of the system of the present invention;
fig. 5 is a schematic diagram of the instrument control circuit of the system of the present invention.
Detailed Description
The invention will now be described in detail with reference to the drawings and specific examples. The present embodiment is implemented on the premise of the technical scheme of the present invention, and a detailed implementation manner and a specific operation process are given, but the protection scope of the present invention is not limited to the following examples.
As shown in fig. 1, which is a schematic structural diagram of an imaging system for simultaneously extracting the internal fluorescent molecular distribution and the surface three-dimensional structure, the system is divided into four modules, namely: the device comprises a light source module, a scanning module, a detection module and a control module.
The light source module comprises a laser 101, a coupler 102, a multimode fiber 103 and a collimator 104; the scanning module comprises a scanning galvanometer 201, a scanning field lens 202 and reflecting lenses 203 and 204; the detection module includes a stage 301, a filter 303, a filter wheel 304, a fluorescence camera 305, and a white light camera 306. The control module includes a computer 401 and USB control lines 402, 403, 404.
Laser light emitted by a laser 101 in the light source module is coupled into one end of a multimode optical fiber 103 through a coupler 102, and then is input into a collimator 104 from the other end of the optical fiber 103 for collimation, and then is projected into a scanning galvanometer 201 of the scanning module.
The scanning module focuses the collimated light beam projected into the scanning galvanometer 201 into a light spot below 1 millimeter through the scanning field mirror 202, the scanning galvanometer 201 converts the light spot into light, the scanning light path of the light is that the scanning galvanometer 201 focuses the light beam onto the reflecting mirror 203 to the reflecting mirror 204 through the scanning field mirror 202, then the light beam reaches the objective table 301, and the sample 302 on the objective table 301 is subjected to reflective line scanning.
The detection module is arranged right above and beside the imaging object stage 301 by using the fluorescence camera 305 and the white light camera 306, the fluorescence camera 305 and the white light camera 306 are at the same height, and can both see the upper surface of the sample 302 in the visual field, the fluorescence camera 305 collects fluorescence information excited in the sample through the filter 303 in the filter wheel 304, and the white light camera 306 directly collects reflected excitation light information of excitation light on the upper surface of the sample, so that fluorescent data for FMT fluorescent line scanning reconstruction and excitation light data for surface line structure light reconstruction are respectively and simultaneously shot by the fluorescence camera 305 and the white light camera 306. The data collection mode is reflective collection, light scans from the upper surface of the sample 302, and fluorescence excited during each scan is captured and collected by the fluorescence camera 305 from the upper surface of the sample 302.
A filter wheel 304 interposed between the fluoroscopic camera 305 and the imaging stage 301 is used to switch the filter 303 of different wavelengths required for data acquisition.
The control module uses a general control computer 401 to transmit instructions by using USB data lines 402, 403 and 404 to control overall data acquisition, and the reconstruction of FMT and three-dimensional surface information is performed on the computer.
The line scanning reconstruction data acquisition process comprises the following steps: firstly, placing a sample 302 on a stage 301, and shooting a bright field white light image of the sample 302 by using a fluorescence camera 305 under bright field conditions; then setting exposure time required by shooting by a white light camera 306 and a fluorescence camera 305, power of the laser 101 and position of a band-pass filter 303 in a filter wheel 304, and starting and setting a scanning mode of the scanning galvanometer 201 as line scanning; and then setting the number and the range of scanning lines, pre-scanning to verify whether the scanning lines and the range meet the expectations, resetting the number and the range of the scanning lines if the scanning lines and the range do not meet the expectations, finally turning off bright field illumination, simultaneously and respectively acquiring tomographic reconstruction data of line scanning and original data of surface reconstruction by using a fluorescence camera 305 and a white light camera 306, turning off a laser after the simultaneous acquisition is finished, and respectively acquiring ambient background light by using the fluorescence camera 305 and the white light camera 306 under the conditions of the same exposure time as the process of shooting reconstruction data, the position of a band-pass filter 303 and light-off darkness.
The surface information reconstruction flow of the system is shown in fig. 3. The process comprises the following steps: first, the center pixel coordinates of the light bar of the line structure light in the surface reconstruction raw data collected by the white light camera 306 are extracted, and the light plane equation of the laser is determined by using the center coordinates of the pixels and the spatial position of the galvanometer relative to the white light camera, then the three-dimensional point cloud of the sample surface is calculated by using the light plane equation and the camera internal parameters, and the grid mesh generated based on the three-dimensional point cloud can be obtained.
The line scan FMT reconstruction flow of the system is shown in fig. 4. The process comprises the following steps: firstly introducing line scanning fluorescence data acquired by a fluorescence camera 305 and used for FMT reconstruction and grid mesh generated by using surface information, setting reconstructed related parameters, generating a finite element grid and calculating a system matrix according to the finite element grid and the related parameters; then carrying out noise processing on the emission light image of the scanning module, and generating a weight matrix according to the processed tomographic reconstruction fluorescence data of the line scanning and a system matrix; and then selecting a corresponding parameter method to solve the inverse problem, and if the reconstructed result accords with the reconstruction expectation, ending the reconstruction, and if the reconstructed result does not accord with the reconstruction expectation, returning to the first step to acquire the data again.
In one embodiment: the light source module is used for shaping the laser beam. The shaping light path is as follows: the laser is emitted from the laser 101, coupled into the multimode optical fiber 103 through the coupler 102, and connected into the collimator 104 at the other end of the optical fiber 103 to obtain a collimated beam, and finally the beam is focused into a spot with the focal length of less than 1mm through the galvanometer 201 and the field lens 202.
In one embodiment: when the laser beam is shaped into a collimated beam from the light source module and projected into the galvanometer 201, the optical path of the reflection type optical path is: the beam is first emitted from the scanning galvanometer 201, through the mirror 203 to the mirror 204, then to the stage 301, and finally to the upper surface of the imaging target 302 on the imaging stage 301.
In one embodiment: the imaging stage 301 functions to position the test sample 302. The filter wheel 304 houses band pass filters 303 of different center wavelengths and is positioned between the fluoroscopic camera 305 and the imaging target 302. The purpose of filter wheel 304 and filter 303 is to filter out excitation light from laser 101, allowing fluorescence to pass through and into imaging detector fluorescence camera 305.
In one embodiment: using 630nm excitation light source 101, data acquisition mode is set to reflective mode, galvanometer 201 is set to line scan mode, 680nm filter 303 is placed in filter wheel 304, and fluorescence camera 305 exposure time is set to 1000ms. The light source projects into the scanning galvanometer 201 from the light source module, reflected scanning light rays from the scanning galvanometer 201 are transmitted from the reflecting mirror 203 to the reflecting mirror 204, then the reflected scanning light rays are transmitted to the object stage 301 to irradiate the upper surface of the sample 302, 20 light rays are respectively scanned on the surface of the sample along the horizontal direction and the vertical direction of the sample, fluorescence is excited by excitation light, 680nm fluorescence is emitted, finally the fluorescence is collected by the fluorescence camera 305 through the filter wheel, and excitation light on the upper surface of the sample is collected by the white light camera 306. And after the data acquisition is finished, reconstructing the FMT and the surface point cloud by using a reconstruction algorithm flow.
In one embodiment: the processor is a computer 401 as a control layer, and a fluorescence camera 305, a white light camera 306, a filter wheel 304, a scanning galvanometer 201 and a laser 101 are used as field layer devices. The method comprises the following steps: the computer 401 controls the on and off of the laser 101, controls the scanning mode (point scanning or line scanning) of the galvanometer 201, the number of scanning points, the conversion of the position of the inner filter 303 of the filter wheel 304, and the shooting of the camera and the storage of pictures. All communication interfaces are connected by USB data lines. The control schematic is shown in fig. 5.
The above examples illustrate only a few embodiments of the invention, which are described in detail and are not to be construed as limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention. Accordingly, the scope of protection of the present invention is to be determined by the appended claims.
Claims (5)
1. An imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure is characterized by comprising a light source module, a scanning module, a detection module and a control module,
the light source module comprises a laser, a coupler, a multimode optical fiber and a collimator, wherein laser emitted by the laser is coupled into one end of the multimode optical fiber through the coupler, and then is input into the collimator from the other end of the optical fiber for collimation and then projected into a scanning vibrating mirror of the scanning module;
the scanning module comprises a scanning vibrating mirror, a scanning field lens and a reflecting mirror, wherein the scanning vibrating mirror focuses a collimated light beam into light spots with the size of less than 1 millimeter through the scanning field lens, the light spots are converted into light rays through the scanning vibrating mirror, and the linear illumination structure of the upper surface of a measured object on the imaging objective table is scanned strip by strip after passing through the reflecting mirror;
the detection module comprises a fluorescence camera and a white light camera, wherein the fluorescence camera and the white light camera are respectively arranged right above and at the side of the imaging object table, the fluorescence camera and the white light camera are at the same height, the fluorescence camera and the white light camera can both see the upper surface of a sample on the imaging object table in the visual field, the fluorescence camera collects fluorescence information of the sample reflected by excitation light or excited by transmission through a filter in a filter wheel, the white light camera directly collects the fluorescence information of the excitation light reflected by the excitation light incident sample, and therefore, a measured object is simultaneously photographed through the fluorescence camera and the white light camera, and fluorescence data reconstructed by fluorescence molecular tomography FMT fluorescent line scanning and the excitation light number used for surface line structural light reconstruction are obtainedAccording toSending the data to a computer in a control module;
the control module comprises a computer and a USB wire, wherein the computer is respectively connected with the laser, the scanning galvanometer, the fluorescent camera and the white light camera through the USB wire, and transmits instructions to control data scanning and acquisition.
2. The imaging system of claim 1, wherein the filter wheel is placed with bandpass filters of different center wavelengths and placed between the fluorescence camera and the object under test, the bandpass filters filtering light other than the projected wavelength.
3. A method for acquiring reconstruction data by an imaging system for simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure is characterized in that a sample is firstly placed on an objective table, and a bright field white light image of the sample is shot by a fluorescent camera under a bright field condition; setting exposure time required by shooting, power of a laser and positions of a band-pass filter in a filter wheel for a white light camera and a fluorescent camera, and starting and setting a scanning mode of a scanning galvanometer as line scanning; and setting the number and the range of the scanning lines, pre-scanning to verify whether the scanning lines and the range meet expectations, resetting the number and the range of the scanning lines if the scanning lines and the range do not meet expectations, turning off bright field illumination if the scanning lines and the range do not meet expectations, simultaneously and respectively acquiring tomographic reconstruction data of line scanning and original data of surface reconstruction by using a fluorescent camera and a white light camera, turning off a laser after the simultaneous acquisition is finished, and respectively acquiring ambient background light by using the fluorescent camera and the white light camera under the conditions of same exposure time, the position of a band-pass filter and darkness when the reconstruction data are shot.
4. A method for reconstructing surface information of imaging system by extracting internal fluorescent molecular distribution and surface three-dimensional structure includes extracting central pixel coordinate of light bar of line structure light in original data of surface reconstruction collected by white light camera, determining light plane equation of laser by central coordinate of pixel and space position of vibrating mirror relative to white light camera, calculating three-dimensional point cloud of sample surface by light plane equation and camera, and generating grid mesh based on three-dimensional point cloud.
5. A method for reconstructing internal fluorescent molecular distribution information of an imaging system by simultaneously extracting internal fluorescent molecular distribution and a surface three-dimensional structure is characterized in that tomographic reconstruction fluorescent data of line scanning acquired by a fluorescent camera and grid mesh generated by utilizing surface information are first introduced, then related parameters of reconstruction are set, a finite element grid is generated, and a system matrix is calculated according to the finite element grid and the related parameters; then carrying out noise processing on the emission light image of the scanning module, and generating a weight matrix according to the processed tomographic reconstruction fluorescence data of the line scanning and a system matrix; and then selecting a corresponding parameter method to solve the inverse problem, and if the reconstructed result accords with the reconstruction expectation, ending the reconstruction, and if the reconstructed result does not accord with the reconstruction expectation, returning to the first step to acquire the data again.
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