CN117030621A - Marking-free laser histological imaging system and imaging method - Google Patents
Marking-free laser histological imaging system and imaging method Download PDFInfo
- Publication number
- CN117030621A CN117030621A CN202311174028.2A CN202311174028A CN117030621A CN 117030621 A CN117030621 A CN 117030621A CN 202311174028 A CN202311174028 A CN 202311174028A CN 117030621 A CN117030621 A CN 117030621A
- Authority
- CN
- China
- Prior art keywords
- laser
- sample
- dichroic mirror
- imaging
- objective lens
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000003384 imaging method Methods 0.000 title claims abstract description 104
- 238000001514 detection method Methods 0.000 claims abstract description 45
- 238000012545 processing Methods 0.000 claims abstract description 33
- 230000005540 biological transmission Effects 0.000 claims abstract description 25
- 238000006073 displacement reaction Methods 0.000 claims abstract description 18
- 238000000034 method Methods 0.000 claims description 20
- 238000001914 filtration Methods 0.000 claims description 14
- 230000008878 coupling Effects 0.000 claims description 13
- 238000010168 coupling process Methods 0.000 claims description 13
- 238000005859 coupling reaction Methods 0.000 claims description 13
- 230000003287 optical effect Effects 0.000 claims description 11
- 238000011144 upstream manufacturing Methods 0.000 claims description 7
- 238000004891 communication Methods 0.000 claims description 4
- 230000001808 coupling effect Effects 0.000 claims description 4
- 239000008367 deionised water Substances 0.000 claims description 4
- 229910021641 deionized water Inorganic materials 0.000 claims description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 4
- 238000005516 engineering process Methods 0.000 abstract description 6
- 238000000799 fluorescence microscopy Methods 0.000 abstract description 2
- 239000000523 sample Substances 0.000 description 74
- 210000001519 tissue Anatomy 0.000 description 27
- 210000000805 cytoplasm Anatomy 0.000 description 20
- 210000003855 cell nucleus Anatomy 0.000 description 15
- 230000000694 effects Effects 0.000 description 11
- 238000010186 staining Methods 0.000 description 10
- 230000005284 excitation Effects 0.000 description 9
- 210000004940 nucleus Anatomy 0.000 description 8
- 238000010586 diagram Methods 0.000 description 7
- 230000001575 pathological effect Effects 0.000 description 7
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 6
- 239000000975 dye Substances 0.000 description 6
- 210000004556 brain Anatomy 0.000 description 5
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 5
- 108020004414 DNA Proteins 0.000 description 4
- 206010028980 Neoplasm Diseases 0.000 description 4
- 238000010827 pathological analysis Methods 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 238000004043 dyeing Methods 0.000 description 3
- YQGOJNYOYNNSMM-UHFFFAOYSA-N eosin Chemical compound [Na+].OC(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C(O)=C(Br)C=C21 YQGOJNYOYNNSMM-UHFFFAOYSA-N 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 2
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 2
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 2
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 2
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000000980 acid dye Substances 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 239000002131 composite material Substances 0.000 description 2
- 238000007405 data analysis Methods 0.000 description 2
- 238000013480 data collection Methods 0.000 description 2
- 210000002744 extracellular matrix Anatomy 0.000 description 2
- 238000002073 fluorescence micrograph Methods 0.000 description 2
- 230000000877 morphologic effect Effects 0.000 description 2
- 230000000269 nucleophilic effect Effects 0.000 description 2
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 1
- 108091093105 Nuclear DNA Proteins 0.000 description 1
- SJEYSFABYSGQBG-UHFFFAOYSA-M Patent blue Chemical compound [Na+].C1=CC(N(CC)CC)=CC=C1C(C=1C(=CC(=CC=1)S([O-])(=O)=O)S([O-])(=O)=O)=C1C=CC(=[N+](CC)CC)C=C1 SJEYSFABYSGQBG-UHFFFAOYSA-M 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000012472 biological sample Substances 0.000 description 1
- 238000001574 biopsy Methods 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 238000004624 confocal microscopy Methods 0.000 description 1
- 230000001054 cortical effect Effects 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000002497 edematous effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000001917 fluorescence detection Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 238000012333 histopathological diagnosis Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000011164 ossification Effects 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 238000005424 photoluminescence Methods 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
- 238000010882 preoperative diagnosis Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 230000001052 transient effect Effects 0.000 description 1
- 238000013519 translation Methods 0.000 description 1
- 238000010200 validation analysis Methods 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/1702—Systems in which incident light is modified in accordance with the properties of the material investigated with opto-acoustic detection, e.g. for gases or analysing solids
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N21/6456—Spatial resolved fluorescence measurements; Imaging
- G01N21/6458—Fluorescence microscopy
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/1702—Systems in which incident light is modified in accordance with the properties of the material investigated with opto-acoustic detection, e.g. for gases or analysing solids
- G01N2021/1706—Systems in which incident light is modified in accordance with the properties of the material investigated with opto-acoustic detection, e.g. for gases or analysing solids in solids
Landscapes
- Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- Life Sciences & Earth Sciences (AREA)
- Immunology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Pathology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Optics & Photonics (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The application discloses a label-free laser histological imaging system and an imaging method, which combine fluorescence imaging and photoacoustic imaging technologies, wherein the imaging system comprises a laser, a filter assembly, a dichroic mirror, an objective lens, a container, an annular ultrasonic transducer, a three-dimensional displacement platform, a transmission assembly, a photoelectric detector, a data acquisition card and an imaging processing terminal. Compared with the prior art, the laser device generates laser through the laser, the dichroic mirror reflects the incident laser to the objective lens, and the laser emitted by the objective lens passes through the through hole arranged in the annular ultrasonic transducer and irradiates the sample; the sample is irradiated by laser to generate fluorescence and penetrates through the dichroic mirror, then is emitted into the photoelectric detector through the transmission component, the data acquisition card respectively transmits ultrasonic signals and fluorescence signals to the imaging processing terminal to generate a sample image, and the tissue sample image can be obtained simultaneously on the premise of no slicing and no marking by combining fluorescence and photoacoustic detection, so that the imaging efficiency is greatly improved.
Description
Technical Field
The application relates to the technical field of microscopic imaging, in particular to a label-free laser histological imaging system and an imaging method.
Background
Histopathological examination is one of the methods for disease diagnosis by pathologists, and plays an important role in preoperative diagnosis, surgical decision and planning, surgical boundary judgment, postoperative validation and assessment. The pathologist realizes accurate study and judgment of tissues such as tumor, infection, inflammation, injury and the like mainly through histomorphology and cell microstructure change in pathological sections. However, conventional pathology examinations often require complex and time-consuming tissue sample preparation, mainly involving the following aspects: specimen collection fixation, tissue processing, paraffin embedding, slice preparation, H & E staining and sealing, which typically take hours to days, make it difficult to quickly identify the edges of cancer tissue during tumor surgery execution, and do not make accurate intra-operative decisions. In contrast, frozen sections can provide rapid histological analysis. However, the freezing process adopted in the frozen section affects the structures of edematous tissues and tissues with high fat content, and generates artifacts, so that the quality of pathological images is poor, and the requirements of pathological diagnosis are difficult to meet. Moreover, because of the hard ossification effect, frozen sections are difficult to apply to hard tissues (e.g., cortical bone and calcified tumors).
Aiming at the problems, a method for considering efficiency and accuracy is urgently needed to be developed, and the pathological features of the fresh tissue sample can be studied and judged in the operation, so that rapid clinical guidance is provided for a doctor to formulate an operation scheme. The nucleus and the cytoplasm are two types of factors which are the most concerned by the histopathological diagnosis, and the morphology, the proportion, the spatial structure and the like of the two types of factors provide information of the pathological diagnosis. In the prior art methods, in order to clearly image the nuclei and cytoplasm in a tissue sample, the tissue sample is usually stained with a stain, and the stain is combined with the nuclear DNA in the tissue sample, thereby showing the microstructure characteristics of the cells. For example, in H & E staining, hematoxylin dye binds to DNA of the nucleus in the form of nucleophilic dye, giving the nucleus a deep blue or purple color; eosin dyes stain the cytoplasm and extracellular matrix in the form of acid dyes, making them pink. However, the existing imaging method needs to perform pathological section and frozen section, and the tissue sample obtained based on the section is dyed, and the conventional pathological section and frozen section all need a series of tedious processes to form an H & E image for a doctor to study and judge, so that the timeliness is low, and the requirements of the guidance in operation are difficult to be met. Therefore, the technical method for imaging a tissue sample in the prior art method has the problem of low imaging speed.
Disclosure of Invention
The application provides a label-free laser histology imaging system and an imaging method, and aims to solve the problems of complex sample preparation, low timeliness, low imaging speed and the like in the traditional pathological section technology. The existing single-mode photoacoustic and fluorescence technology can only acquire certain side information of the cell nucleus or the cytoplasm, and the histopathological examination depends on the composite information of the cell nucleus and the cytoplasm, so that the photoacoustic or fluorescence technology cannot meet the requirements of histological detection. The application adopts the short pulse laser with the wavelength of 266nm as a light source, DNA/RNA rich in the cell nucleus in the sample has strong optical absorption effect on the wave band, and can be stimulated to generate a photoacoustic signal, thereby specifically presenting the form of the cell nucleus and accurately matching with a hematoxylin staining chart in a gold standard H & E imaging method; the pigment substances such as tryptophan, tyrosine and the like contained in the cytoplasm in the sample can automatically emit fluorescent signals under the irradiation of the laser with the wavelength, so that the structure of the cytoplasm is displayed with high contrast and is accurately matched with eosin staining patterns in a gold standard H & E imaging method. The application does not need to carry out complicated sample preparation process, can simultaneously obtain morphological structure images of two elements of nucleus and cytoplasm of a fresh tissue sample on the premise of no slicing and no staining, is in one-to-one correspondence with the histopathological examination gold standard H & E staining image, and has the application potential of rapid histopathological detection.
In a first aspect, embodiments of the present application provide a label-free laser histological imaging system, wherein,
the system comprises a laser, a filter assembly, a dichroic mirror, an objective lens, a container, an annular ultrasonic transducer, a three-dimensional displacement platform, a transmission assembly, a photoelectric detector, a data acquisition card and an imaging processing terminal;
the laser is arranged at the upstream of the dichroic mirror, and the filter component is arranged between the laser and the dichroic mirror;
the annular ultrasonic transducer is arranged in the container, a coupling solution is placed in the container, and a sample is placed in the container; the three-dimensional displacement platform is fixedly connected with the container; the connecting line between the objective lens and the sample passes through a through hole arranged in the annular ultrasonic transducer;
the objective lens is arranged above the container, and the objective lens is positioned between the dichroic mirror and the container; the photoelectric detector is arranged on the other side of the dichroic mirror opposite to the objective lens, and the transmission component is arranged between the photoelectric detector and the dichroic mirror;
the laser generated by the laser is emitted into the dichroic mirror through the filtering component; the dichroic mirror reflects the incident laser to the objective lens, and the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample;
the sample is irradiated by laser to generate fluorescence and penetrates through the dichroic mirror, and then is injected into the photoelectric detector through the transmission component;
the data acquisition card is respectively in communication connection with the annular ultrasonic transducer, the photoelectric detector and the imaging processing terminal;
the data acquisition card respectively acquires ultrasonic signals obtained by detection of the annular ultrasonic transducer and fluorescent signals obtained by detection of the photoelectric detector and outputs the ultrasonic signals to the imaging processing terminal; the imaging processing terminal processes the ultrasonic signal and the fluorescence signal to generate a sample image.
The label-free laser histological imaging system, wherein the sample is placed at the focal point of the beam emitted by the objective lens.
The unmarked laser histological imaging system, wherein the laser is a short wave ultraviolet laser.
The unmarked laser histological imaging system comprises a filter assembly, a first lens, a second lens and an incidence baffle;
the incident baffle is arranged between the first lens and the second lens, and an incident pinhole is arranged on the incident baffle;
the laser beam emitted by the first transmission mirror is focused at the incidence needle hole and emitted to the second lens through the incidence needle hole.
The unmarked laser histological imaging system comprises a transmission assembly, a first lens, a second lens, a third lens, a light filter and a detection baffle;
the optical filter is arranged between the third lens and the detection baffle, and the third lens is positioned at the upstream of the fluorescent light beam emitted by the dichroic mirror; the detection baffle is provided with a detection pinhole;
the fluorescent light beam emitted by the optical filter is focused at the detection pinhole and is emitted to the photoelectric detector through the detection pinhole.
The label-free laser histological imaging system, wherein the optical filter is a long-pass optical filter.
The label-free laser histological imaging system, wherein the coupling solution is deionized water.
In a second aspect, an embodiment of the present application further provides a label-free laser histological imaging method, where the imaging method is applied to the imaging system in the first aspect, and the imaging method includes:
starting a laser to emit laser beams, filtering the laser beams by a filtering component, expanding the laser beams, and then injecting the laser beams into a dichroic mirror;
the dichroic mirror reflects the incident laser to the objective lens, the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample, and the sample and the coupling solution in the container are subjected to coupling action to generate an ultrasonic signal;
controlling the three-dimensional displacement platform to drive the container to move in three dimensions so as to enable laser emitted by the objective lens to scan the sample in three dimensions;
after the sample is irradiated by laser to generate fluorescence and penetrates through a dichroic mirror, the fluorescence is emitted into a photoelectric detector through a transmission component; the data acquisition card acquires ultrasonic signals obtained by detection of the annular ultrasonic transducer, and fluorescence signals obtained by detection of the photoelectric detector and transmits the signals to the imaging processing terminal in real time;
the imaging processing terminal performs imaging processing on the ultrasonic signals and the fluorescent signals at the same time to generate a sample image.
The laser emission wavelength of the laser is 250-275nm.
The marking-free laser histological imaging method comprises the steps of enabling the central wavelength of the fluorescent signal to be 290-310nm and 345-365nm.
The embodiment of the application provides a label-free laser histological imaging system and an imaging method. Compared with the prior art, the application generates laser through the laser and emits the laser into the dichroic mirror through the filter component; the dichroic mirror reflects the incident laser to the objective lens, and the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample; after the sample is irradiated by laser to generate fluorescence and penetrates through a dichroic mirror, the fluorescence is transmitted into a photoelectric detector through a transmission component, and a data acquisition card respectively acquires a photoacoustic signal obtained by detection of an annular ultrasonic transducer and a fluorescence signal obtained by detection of the photoelectric detector and outputs the photoacoustic signal and the fluorescence signal to an imaging processing terminal; the imaging processing terminal comprehensively processes the two received signals to generate a sample image, and by combining fluorescence and photoacoustic detection, tissue samples do not need to be dyed, histological images of fresh tissue samples can be obtained on the premise of no slicing and no marking, imaging efficiency is greatly improved, the histological images are in one-to-one correspondence with the histopathological examination gold standard H & E dyeing images, and matching is accurate.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for the description of the embodiments will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present application, and other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a block diagram of a device of a label-free laser histological imaging system provided by an embodiment of the application;
FIG. 2 is a method flow chart of a method for label-free laser histological imaging provided by an embodiment of the present application;
fig. 3 is an application effect diagram of a marking-free laser histological imaging method according to an embodiment of the present application.
Reference numerals: 1. a laser; 2. a filtering component; 3. a dichroic mirror; 4. an objective lens; 5. a container; 6. an annular ultrasonic transducer; 7. a three-dimensional displacement platform; 8. a transmissive assembly; 9. a photodetector; 10. a data acquisition card; 11. an imaging processing terminal; 21. a first lens; 22. a second lens; 23. an incident baffle; 81. a third lens; 82. a light filter; 83. the baffle is detected.
Detailed Description
The following description of the embodiments of the present application will be made clearly and fully with reference to the accompanying drawings, in which it is evident that the embodiments described are some, but not all embodiments of the application. All other embodiments, which can be made by those skilled in the art based on the embodiments of the application without making any inventive effort, are intended to be within the scope of the application.
It should be understood that the terms "comprises" and "comprising," when used in this specification and the appended claims, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof.
It is also to be understood that the terminology used in the description of the application is for the purpose of describing particular embodiments only and is not intended to be limiting of the application. As used in this specification and the appended claims, the singular forms "a," "an," and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise.
It should be further understood that the term "and/or" as used in the present specification and the appended claims refers to any and all possible combinations of one or more of the associated listed items, and includes such combinations.
In this embodiment, please refer to fig. 1. As shown in the figure, the embodiment of the application provides a label-free laser histological imaging system, wherein the system comprises a laser 1, a filter component 2, a dichroic mirror 3, an objective lens 4, a container 5, an annular ultrasonic transducer 6, a three-dimensional displacement platform 7, a transmission component 8, a photoelectric detector 9, a data acquisition card 10 and an imaging processing terminal 11; the laser 1 is arranged upstream of the dichroic mirror 3, and the filter assembly 2 is arranged between the laser 1 and the dichroic mirror 3; the annular ultrasonic transducer 6 is arranged in the container 5, a coupling solution is placed in the container 5, and a sample is placed in the container 5; the three-dimensional displacement platform 7 is fixedly connected with the container 5; the connecting line between the objective lens 4 and the sample passes through a through hole arranged in the annular ultrasonic transducer 6; the objective lens 4 is arranged above the container 5, and the objective lens 4 is positioned between the dichroic mirror 3 and the container 5; the photodetector 9 is arranged on the other side of the dichroic mirror 3 opposite to the objective lens 4, and the transmission component 8 is arranged between the photodetector 9 and the dichroic mirror 3; the laser light generated by the laser 1 is emitted into the dichroic mirror 3 through the filter component 2; the dichroic mirror 3 reflects the incident laser light to the objective lens 4, and the laser light emitted from the objective lens 4 passes through a through hole provided in the annular ultrasonic transducer 6 and irradiates a sample; after the sample is irradiated by laser to generate fluorescence and penetrates through the dichroic mirror 3, the fluorescence is emitted into the photodetector 9 through the transmission component 8; the data acquisition card 10 is respectively in communication connection with the annular ultrasonic transducer 6, the photoelectric detector 9 and the imaging processing terminal 11; the data acquisition card 10 acquires ultrasonic signals detected by the annular ultrasonic transducer 6 and fluorescent signals detected by the photoelectric detector 9 respectively and outputs the ultrasonic signals and the fluorescent signals to the imaging processing terminal 11; the imaging processing terminal 11 processes the ultrasonic signal and the fluorescence signal to generate a sample image.
Specifically, the laser 1 is disposed at the upstream of the imaging light path and is used as an excitation light source, the laser beam generated by the laser 1 passes through the filter component 2, the filter component 2 filters and expands the laser beam, the laser beam emitted by the filter component 2 is reflected to the objective lens 4 through the dichroic mirror 3, and the filter component 2 expands the laser beam to maximize the back aperture of the objective lens 4 filled with the laser beam. Under the irradiation of laser beams, the same position of a tissue sample simultaneously generates ultrasonic waves and fluorescence at a focal point due to the transient thermoelastic effect and the photoluminescence effect, the annular ultrasonic transducer 6 detects the generated ultrasonic signals (the coupling effect is generated between the tissue sample and the ultrasonic transducer through the coupling solution in the container 5), and the ultrasonic signals are collected by the data collection card 10 and transmitted to the imaging processing terminal 11 for storage for subsequent data analysis and image reconstruction, so that the specific form of the cell nucleus can be reflected. Meanwhile, the generated fluorescence is transmitted through the dichroic mirror 3 and then is emitted into the transmission component 8, the fluorescence is purified by the transmission component 8 and then is emitted into the photoelectric detector 9, scattered light from a non-focal plane in the fluorescence excited by the sample can be removed through purification treatment, the photoelectric detector 9 realizes detection of fluorescence signals, and the fluorescence signals are collected through the data collection card 10 and transmitted to the imaging processing terminal 11 for storage, so that the specific form of cytoplasm can be reflected for subsequent data analysis and image reconstruction.
More specifically, the three-dimensional displacement platform 7 is in communication connection with the imaging processing terminal 11. The imaging processing terminal 11 can control the movement of the three-dimensional displacement platform 7 by using the LABVIEW, the three-dimensional displacement platform 7 drives the container 5 and the tissue sample in the container 5 to move, and an imaging image of the complete sample can be obtained through single scanning of an XY axis, and the imaging image simultaneously contains composite information of cell nuclei and cytoplasm. Because the photoacoustic (ultrasonic signal) and the fluorescence signal are detected simultaneously in the process of three-dimensional movement of the sample, the problems of shift, mismatch and the like cannot occur in the process of inherent registration of cell nuclei and cytoplasm in the process of reconstructing the sample image. Wherein, for thinner tissue samples (thickness below 7 um), the XY plane only needs one translation; for thicker tissue samples, the effect of confocal microscopy is achieved by matching with Z-axis movement of the samples, and detection information of different depths in the thick samples can be obtained.
In a more specific embodiment, the sample is placed at the focal point of the beam emitted by the objective lens 4. Wherein the laser 1 is a short wave ultraviolet laser 1. Specifically, the coupling solution is deionized water.
In order to achieve a better imaging effect, a sample can be arranged at the focal point of the light beam emitted by the objective lens 4, so that the same position of the tissue sample can generate ultrasonic waves and fluorescence at the focal point at the same time, the concentration of the generated ultrasonic waves and fluorescence is improved, namely, the signal intensity of ultrasonic signals and fluorescence signals is improved, and the imaging quality of the sample is improved. Further, in order to improve the quality of fluorescence generated by the sample, the laser 1 may be a short-wave ultraviolet laser 1, and then the laser 1 may generate short-wave ultraviolet light (uv-C ultraviolet light with a wavelength between 200nm and 280 nm). In order to improve the quality of ultrasonic signals generated by coupling the samples, the coupling solution can be deionized water. Furthermore, the laser 1 may be set as a 266nm short pulse laser 1, so that the laser 1 may generate a 266nm short pulse laser beam, thereby improving the effect of the laser beam on fluorescence excitation of the sample. The wavelength of the laser beam is not limited to 266nm, but may be other wavelengths, based on the peak value of the excitation light of autofluorescence of different substances in the sample.
In a more specific embodiment, the filtering component 2 includes a first lens 21, a second lens 22, and an incident baffle 23; the incident baffle 23 is disposed between the first lens 21 and the second lens 22, and an incident pinhole is disposed on the incident baffle 23; the laser beam emitted from the first transmission mirror is focused at the incident pinhole and emitted from the incident pinhole to the second lens 22.
Specifically, a 4F system comprising the first lens 21 and the second lens 22 may be configured to perform a beam expanding operation on the excitation beam, and the incident baffle 23 is provided with an incident pinhole for filtering out the spatial stray wave, thereby improving the purity of the laser beam.
In a more specific embodiment, the transmission assembly 8 includes a third lens 81, a filter 82, and a detection baffle 83; the filter 82 is disposed between the third lens 81 and the detection baffle 83, and the third lens 81 is located upstream of the fluorescent light beam emitted from the dichroic mirror 3; the detection baffle 83 is provided with a detection pinhole; the fluorescent light beam emitted from the filter 82 is focused at the detection pinhole and emitted from the detection pinhole to the photodetector 9. Wherein the filter 82 is a long-pass filter 82.
Further, the transmission assembly 8 may be configured to include a third lens 81, an optical filter 82, and a detection baffle 83, where the third lens 81 is configured to focus fluorescence, and after the fluorescence is focused by the third lens 81, the fluorescence is filtered by the optical filter 82, so that the fluorescence beam is further purified, and the fluorescence beam passes through a detection pinhole disposed on the detection baffle 83, and the detection pinhole can reject stray light from a non-focal plane in the sample, so as to improve the fluorescence quality output to the photodetector 9. To further enhance the filtering effect of the filter 82, the filter 82 may be provided as a long-pass filter 82.
The microscopic imaging system disclosed by the application integrates two technologies of photoacoustic imaging and fluorescence imaging. The excitation and detection modes of the photoacoustic signal and the fluorescence signal generated by sample coupling are innovatively designed. Firstly, in the aspect of signal excitation, short pulse laser with the wavelength of 266nm is adopted as a light source, DNA/RNA rich in cell nuclei in a sample has strong optical absorption effect on the wave band, and photoacoustic signals can be generated by excitation, so that the cell nuclei are specifically presented; tryptophan and tyrosine contained in cytoplasm in the sample are stimulated to radiate under the laser irradiation with the wavelength to generate fluorescence signals with peak value of about 355nm and 300nm (fluorescence detection wavelength is not limited to about 300nm and 355nm, and is based on the emission peak value of autofluorescence of different substances in the sample), thereby displaying cytoplasm structure with high contrast. Secondly, in the aspect of signal detection, a reflective photoacoustic-fluorescence signal acquisition light path is established (an imaging system is not limited to reflective type, and can also be transmissive type, and the actual requirements and experimental effects are taken as targets), an annular ultrasonic transducer is adopted to detect a photoacoustic signal, and a photoacoustic image is reconstructed; the dichroic mirror meeting the system requirement of the inverse and transmission wavelength is used for separating the incident light and the stimulated fluorescence signal, the long-pass filter is used for further purifying the fluorescence signal, and then the fluorescence signal is collected by the photoelectric detector, so that a fluorescence image is reconstructed. The photoacoustic signals and the fluorescent signals are mutually noninterfere and mutually complement, the three-dimensional displacement platform does not need to repeatedly move for excitation and collection of the fluorescent signals, one-time movement and two imaging are achieved, the two images are inherently registered and correspond to the histopathological examination gold standard H & E staining images one by one, and a quick and reliable technical means is hopefully provided for pathological diagnosis in operation.
The embodiment of the application also provides a marking-free laser histological imaging method, referring to fig. 2, as shown in the figure, the imaging method comprises steps S110 to S160.
S110, starting the laser to emit laser beams, filtering the laser beams by the filtering component, expanding the laser beams, and then injecting the laser beams into the dichroic mirror.
The position of the sample to be imaged can be adjusted to be positioned at the focal point of the beam emitted by the objective lens, then the laser is started to emit laser beams, the laser beams are filtered and expanded by the filtering component, the laser beams are emitted into the dichroic mirror, the dichroic mirror reflects the beams with shorter wavelengths (such as the beams with the wavelengths of 250-275 nm) and transmits the beams with longer wavelengths (such as the beams with the wavelengths of more than 300 nm). Wherein the laser emitted by the laser has a laser wavelength of 250-275nm, and the preferred laser wavelength is 266nm.
And S120, the dichroic mirror reflects the incident laser to the objective lens, the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample, and the sample and the coupling solution in the container are subjected to coupling action to generate an ultrasonic signal.
The laser emitted by the objective lens passes through the through holes arranged in the annular ultrasonic transducer and irradiates the sample, the annular ultrasonic transducer can collect ultrasonic signals around the light spot, the annular ultrasonic transducer can improve the integrity of ultrasonic signal collection, and the laser beam and the fluorescent beam are emitted through the through holes arranged in the annular ultrasonic transducer, so that the collection of the ultrasonic signals and the emission of the laser beam and the emission of the fluorescent beam are not mutually interfered.
S130, controlling the three-dimensional displacement platform to drive the container to move in three dimensions, so that laser emitted by the objective lens scans the sample in three dimensions.
And controlling the three-dimensional displacement platform to drive the container so as to enable the container to move in three dimensions. Specifically, the container can be driven to perform two-dimensional scanning on an XY axis (horizontal plane) through the three-dimensional displacement platform; the container can be driven by the three-dimensional displacement platform to carry out three-dimensional scanning on XYZ axes.
S140, after the sample is irradiated by laser to generate fluorescence and penetrates through the dichroic mirror, the fluorescence is emitted into the photoelectric detector through the transmission component.
The photodetector is used for collecting the generated fluorescence signals, wherein the center wavelength of the fluorescence signals is 290-310nm and 345-365nm, and preferably, the center wavelength of the fluorescence signals is 300nm and 355nm respectively.
And S150, the data acquisition card simultaneously acquires ultrasonic signals obtained by detection of the annular ultrasonic transducer and fluorescent signals obtained by detection of the photoelectric detector, and transmits the ultrasonic signals to the imaging processing terminal in real time.
And S160, the imaging processing terminal performs imaging processing on the ultrasonic signals and the fluorescence signals at the same moment to generate a sample image.
In the implementation of the application, a single excitation light source is used for respectively exciting and generating ultrasonic signals (photoacoustic images) and fluorescence signals (autofluorescence images), and the ultrasonic signals and the fluorescence signals are respectively and simultaneously detected by the annular ultrasonic transducer and the photoelectric detector, so that the photoacoustic images and the fluorescence images of biological samples can be rapidly and high-contrast obtained, and the images of cell nuclei and cytoplasm are synchronously obtained (the prior single-mode photoacoustic and fluorescence technology can only obtain certain side information of the cell nuclei or cytoplasm is overcome), and the two images are automatically registered, so that the integrity of microscopic images of tissue samples is ensured. The method can directly observe the biopsy tissue sample of fresh materials, can simultaneously obtain the nucleus and cytoplasm images of the tissue sample on the premise of no slicing and no marking, and is expected to provide novel histological imaging technical support for pathological diagnosis in operation.
The application is proved by experiments, the developed label-free laser histological imaging system successfully realizes one-time scanning and two imaging on a mouse brain slice sample (about 7um thickness), and the registration of the sample nuclei and cytoplasm imaged by the system is accurate and the information is reliable through the comparison verification with a sample H & E staining image, as shown in figure 3. The image (a) in fig. 3 is a photoacoustic image obtained by imaging an ultrasonic signal of a mouse brain slice sample correspondingly, and shows that the imaging system specifically images the nucleus of the sample; fig. 3 (b) is a fluorescence diagram obtained by imaging the fluorescence signal of the mouse brain slice sample correspondingly, which shows the specific imaging of the imaging system to the cytoplasm of the sample; fig. 3 (c) is a fused diagram obtained by fusing a photoacoustic diagram and a fluorescence diagram of a mouse brain slice sample, and fig. 3 (d) is an H & E diagram obtained by H & E staining a mouse brain slice sample. The fusion map obtained by fusing the map (a) and the map (b) is in one-to-one correspondence with the H & E image of the sample, so that nondestructive acquisition of pathological section information is realized (in H & E staining, hematoxylin dye is combined onto DNA of cell nuclei in a nucleophilic dye form to enable the cell nuclei to be dark blue or purple, eosin dye is used for dyeing cytoplasm and extracellular matrix in an acid dye form to enable the cell nuclei to be pink), and the ratio of FIG. 3 is: 500um.
The application provides a marking-free laser histological imaging system and an imaging method. Compared with the prior art, the application generates laser through the laser and emits the laser into the dichroic mirror through the filter component; the dichroic mirror reflects the incident laser to the objective lens, and the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample; after the sample is irradiated by laser to generate fluorescence and penetrates through a dichroic mirror, the fluorescence is transmitted into a photoelectric detector through a transmission component, and a data acquisition card respectively acquires an ultrasonic signal obtained by detection of an annular ultrasonic transducer and a fluorescence signal obtained by detection of the photoelectric detector and outputs the ultrasonic signal and the fluorescence signal to an imaging processing terminal; the imaging processing terminal comprehensively processes the two received signals to generate a sample image, and combines fluorescence and photoacoustic detection, so that dyeing treatment is not required to be carried out on a tissue sample, and morphological structure images of two elements, namely a cell nucleus and a cell cytoplasm of a fresh tissue sample can be obtained simultaneously on the premise of no slicing and no marking, thereby greatly improving the imaging efficiency.
While the application has been described with reference to certain preferred embodiments, it will be understood by those skilled in the art that various changes and substitutions of equivalents may be made and equivalents will be apparent to those skilled in the art without departing from the scope of the application. Therefore, the protection scope of the application is subject to the protection scope of the claims.
Claims (10)
1. The system is characterized by comprising a laser, a filtering component, a dichroic mirror, an objective lens, a container, an annular ultrasonic transducer, a three-dimensional displacement platform, a transmission component, a photoelectric detector, a data acquisition card and an imaging processing terminal;
the laser is arranged at the upstream of the dichroic mirror, and the filter component is arranged between the laser and the dichroic mirror;
the annular ultrasonic transducer is arranged in the container, a coupling solution is placed in the container, and a sample is placed in the container; the three-dimensional displacement platform is fixedly connected with the container; the connecting line between the objective lens and the sample passes through a through hole arranged in the annular ultrasonic transducer;
the objective lens is arranged above the container, and the objective lens is positioned between the dichroic mirror and the container; the photoelectric detector is arranged on the other side of the dichroic mirror opposite to the objective lens, and the transmission component is arranged between the photoelectric detector and the dichroic mirror;
the laser generated by the laser is emitted into the dichroic mirror through the filtering component; the dichroic mirror reflects the incident laser to the objective lens, and the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample;
the sample is irradiated by laser to generate fluorescence and penetrates through the dichroic mirror, and then is injected into the photoelectric detector through the transmission component;
the data acquisition card is respectively in communication connection with the annular ultrasonic transducer, the photoelectric detector and the imaging processing terminal;
the data acquisition card respectively acquires ultrasonic signals obtained by detection of the annular ultrasonic transducer and fluorescent signals obtained by detection of the photoelectric detector and outputs the ultrasonic signals to the imaging processing terminal; the imaging processing terminal processes the ultrasonic signal and the fluorescence signal to generate a sample image.
2. The label-free laser histological imaging system of claim 1, wherein the sample is placed at the focal point of the objective lens exit beam.
3. The label-free laser histological imaging system of claim 1, wherein the laser is a short wave ultraviolet laser.
4. The marker-free laser histological imaging system of any of claims 1-3, wherein the filter assembly comprises a first lens, a second lens, and an incidence baffle;
the incident baffle is arranged between the first lens and the second lens, and an incident pinhole is arranged on the incident baffle;
the laser beam emitted by the first transmission mirror is focused at the incidence needle hole and emitted to the second lens through the incidence needle hole.
5. The marker-free laser histological imaging system of any of claims 1-3, wherein the transmissive assembly comprises a third lens, optical filter, and detection baffle;
the optical filter is arranged between the third lens and the detection baffle, and the third lens is positioned at the upstream of the fluorescent light beam emitted by the dichroic mirror; the detection baffle is provided with a detection pinhole;
the fluorescent light beam emitted by the optical filter is focused at the detection pinhole and is emitted to the photoelectric detector through the detection pinhole.
6. The label-free laser histological imaging system of claim 5, wherein the filter is a long pass filter.
7. A label-free laser histological imaging system according to any of claims 1-3, wherein the coupling solution is deionized water.
8. A method of label-free laser histological imaging, wherein the imaging method is applied in an imaging system according to any of claims 1 to 7, the imaging method comprising:
starting a laser to emit laser beams, filtering the laser beams by a filtering component, expanding the laser beams, and then injecting the laser beams into a dichroic mirror;
the dichroic mirror reflects the incident laser to the objective lens, the laser emitted by the objective lens passes through a through hole arranged in the annular ultrasonic transducer and irradiates a sample, and the sample and the coupling solution in the container are subjected to coupling action to generate an ultrasonic signal;
controlling the three-dimensional displacement platform to drive the container to move in three dimensions so as to enable laser emitted by the objective lens to scan the sample in three dimensions;
after the sample is irradiated by laser to generate fluorescence and penetrates through a dichroic mirror, the fluorescence is emitted into a photoelectric detector through a transmission component;
the data acquisition card acquires ultrasonic signals obtained by detection of the annular ultrasonic transducer, and fluorescence signals obtained by detection of the photoelectric detector and transmits the signals to the imaging processing terminal in real time;
the imaging processing terminal performs imaging processing on the ultrasonic signals and the fluorescent signals at the same time to generate a sample image.
9. The method of claim 8, wherein the laser emits laser light at a wavelength of 250-275nm.
10. The label-free laser histological imaging method of claim 8, wherein the fluorescence signal has a center wavelength of 290-310nm and 345-365nm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311174028.2A CN117030621A (en) | 2023-09-11 | 2023-09-11 | Marking-free laser histological imaging system and imaging method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311174028.2A CN117030621A (en) | 2023-09-11 | 2023-09-11 | Marking-free laser histological imaging system and imaging method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117030621A true CN117030621A (en) | 2023-11-10 |
Family
ID=88645006
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311174028.2A Pending CN117030621A (en) | 2023-09-11 | 2023-09-11 | Marking-free laser histological imaging system and imaging method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117030621A (en) |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102621115A (en) * | 2012-02-27 | 2012-08-01 | 华南师范大学 | Confocal simultaneous opto-acoustic imaging and fluorescence imaging method and device |
| WO2015003449A1 (en) * | 2013-07-12 | 2015-01-15 | 深圳先进技术研究院 | Optoacoustic-fluorescence dual-mode endoscope |
| CN105548102A (en) * | 2015-12-07 | 2016-05-04 | 华南师范大学 | Deep calcium ion concentration monitoring method using photoacoustic-fluorescent complementary principle and device using the same |
| US20160287211A1 (en) * | 2015-03-31 | 2016-10-06 | Ralph S. DaCosta | System and Method for Multi-Modal in Vivo Imaging |
| CN114264614A (en) * | 2022-01-25 | 2022-04-01 | 深圳大学 | Label-free photoacoustic pathological microscopic imaging system and imaging method |
| CN114324183A (en) * | 2022-01-26 | 2022-04-12 | 深圳大学 | Ultraviolet photoacoustic microscopic imaging system with large focal depth and imaging method |
| CN114468960A (en) * | 2021-12-21 | 2022-05-13 | 华南师范大学 | Endoscope device integrating optics, optoacoustic, OCT and fluorescence multi-mode forward imaging and imaging method thereof |
| CN115656120A (en) * | 2022-10-09 | 2023-01-31 | 中国科学院深圳先进技术研究院 | Dark field reflection ultraviolet optical microscopic imaging method and system |
-
2023
- 2023-09-11 CN CN202311174028.2A patent/CN117030621A/en active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102621115A (en) * | 2012-02-27 | 2012-08-01 | 华南师范大学 | Confocal simultaneous opto-acoustic imaging and fluorescence imaging method and device |
| WO2015003449A1 (en) * | 2013-07-12 | 2015-01-15 | 深圳先进技术研究院 | Optoacoustic-fluorescence dual-mode endoscope |
| US20160287211A1 (en) * | 2015-03-31 | 2016-10-06 | Ralph S. DaCosta | System and Method for Multi-Modal in Vivo Imaging |
| CN105548102A (en) * | 2015-12-07 | 2016-05-04 | 华南师范大学 | Deep calcium ion concentration monitoring method using photoacoustic-fluorescent complementary principle and device using the same |
| CN114468960A (en) * | 2021-12-21 | 2022-05-13 | 华南师范大学 | Endoscope device integrating optics, optoacoustic, OCT and fluorescence multi-mode forward imaging and imaging method thereof |
| CN114264614A (en) * | 2022-01-25 | 2022-04-01 | 深圳大学 | Label-free photoacoustic pathological microscopic imaging system and imaging method |
| CN114324183A (en) * | 2022-01-26 | 2022-04-12 | 深圳大学 | Ultraviolet photoacoustic microscopic imaging system with large focal depth and imaging method |
| CN115656120A (en) * | 2022-10-09 | 2023-01-31 | 中国科学院深圳先进技术研究院 | Dark field reflection ultraviolet optical microscopic imaging method and system |
Non-Patent Citations (1)
| Title |
|---|
| 闫宝运 等: "一体化的光声-荧光显微镜用于活体肿瘤成像", 激光生物学报, vol. 26, no. 03, 30 June 2017 (2017-06-30), pages 212 - 216 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP6775492B2 (en) | Systems and methods for controlling the imaging depth in tissues by fluorescence microscopy using UV excitation after staining with fluorescent agents | |
| US9964489B2 (en) | System and method for controlling depth of imaging in tissues using fluorescence microscopy under ultraviolet excitation following staining with fluorescing agents | |
| EP3164689B1 (en) | Novel methods of tissue processing for deep imaging | |
| JP5687201B2 (en) | Combined microscopy | |
| US20110224574A1 (en) | Methods and systems for tissue processing and imaging | |
| EP3414553A2 (en) | Method and apparatus for imaging unsectioned tissue speciments | |
| CN108348224A (en) | Biopsy device for coherent raman imaging | |
| CN110755042B (en) | Time pulse light sheet tomography method and system for realizing large-volume high-resolution | |
| WO2018175565A1 (en) | System and method for controlling depth of imaging in tissues using fluorescence microscopy under ultraviolet excitation following staining with fluorescing agents | |
| EP3550510A1 (en) | Method for producing image of biological sample and optical system using same | |
| CN117705773A (en) | Modularized multi-mode microscopic optical analysis system | |
| CN110726702A (en) | Optical layer cutting device adopting advanced optical interference microscopy | |
| JP7231345B2 (en) | Preparation of tissue sections using fluorescence-based detection | |
| CN117030621A (en) | Marking-free laser histological imaging system and imaging method | |
| Zhang et al. | Speckle illumination microscopy enables slide-free and non-destructive pathology of human lung adenocarcinoma | |
| WO2024082717A1 (en) | System and method for rapid and label-free imaging of biological tissues based on microscopy with ultraviolet single-plane illumination | |
| US20240192120A1 (en) | Systems and methods for preparing imaging and analyzing cytologic samples | |
| WO2023061068A1 (en) | Translational rapid ultraviolet-excited sectioning tomography assisted with deep learning | |
| Wu et al. | High-throughput, nondestructive, and low-cost histological imaging with deep-learning-assisted UV microscopy | |
| Fereidouni et al. | MUSE: a novel approach for slide-free microscopy | |
| Patel | Developing SCAPE Microscopy for Real-Time, Volumetric Imaging at the Point-of-Care | |
| Li | Label-Free Photoacoustic Microscopy for Biomedical and Point-of-Care Applications | |
| Hu | Dual-View Inverted Selective Plane Illumination Microscopy (diSPIM) Imaging for Accurate 3D Digital Pathology | |
| Lin | Volumetric stimulated Raman scattering microscopy | |
| Crosignani et al. | Ultra-deep imaging of turbid samples by enhanced photon harvesting |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |