Micrometer-resolution magnetic imaging of tumor tissue

Abstract

The present invention provides a method for magnetic imaging of tumor tissue with micron resolution, comprising: labeling tissue samples with superparamagnetic nanoparticles, wherein the superparamagnetic nanoparticles are labeled on tumor marker proteins of the tissue samples; attaching diamonds to the tissue samples to form a diamond-tumor tissue slice-cover glass structure; using the NV color center two-dimensional magnetic sensor in the diamond to measure the magnetic field of the tissue sample to obtain a magnetic field image of the tissue sample; and processing the magnetic field image with a deep learning model to obtain a micron resolution magnetic imaging map of the tissue sample. The method for magnetic imaging of tumor tissue provided by the present invention has high signal stability, low background, and can achieve absolute quantification. It can realize magnetic and optical multimodal correlation imaging of biological tissues. Both magnetic and optical imaging can achieve micron-level spatial resolution. It can also be compatible with commercial optical microscopes and is easy to promote and use.

Description

Technical Field

[0001] The invention relates to the field of imaging, and in particular to a method for magnetic imaging of tumor tissue with micron resolution. Background Art

[0002] Cancer is one of the major diseases that affect human health. The study of the molecular mechanism and accurate diagnosis of cancer are the basis for the effective prevention and treatment of tumor diseases. Traditional tissue imaging methods mainly include HE staining, immunohistochemistry, immunofluorescence, etc. In recent years, multispectral imaging methods based on tyramide signal amplification, multichannel imaging based on mass spectrometry detection, and some other label-free spectral imaging methods have also emerged.

[0003] Traditional optical imaging methods are often plagued by background optical signals and signal instability, and can only provide relative quantitative information of tumor markers, while various optical methods cannot be used simultaneously on the same tissue slice, for example:

[0004] Hematoxylin-eosin staining (HE staining) is mainly a chemical staining based on the differential binding of different cell components to dyes. The diagnostic criteria vary from person to person and are more dependent on experience. HE staining lacks tumor marker information because it has no specific immune markers. HE staining forms obvious color signals while producing a large amount of autofluorescence, so it cannot be used simultaneously with optical methods such as immunohistochemistry and immunofluorescence.

[0005] Immunohistochemistry (IHC) can be divided into immunohistochemistry and immunofluorescence. Immunohistochemistry combines the specificity of immune response and the visibility of histochemistry. With the help of optical microscope, various antigens can be imaged at the cellular and subcellular levels. Immunohistochemistry has the advantages of simple operation, stable signal, long-term storage of samples, and low cost. However, it cannot perform accurate quantitative analysis of tumor markers, and can only perform relative quantitative grading. It is more dependent on the experience of clinical pathologists, and it is difficult to achieve multi-target co-labeling. In tissues such as glands and liver, due to the influence of endogenous enzyme activity, non-specific binding or high background staining is often prone to occur, interfering with judgment. In addition, immunohistochemistry is difficult to use together with other optical methods on the same tissue section.

[0006] Immunofluorescence technology uses fluorescent molecules to label antigens. Through spectral differentiation, it can achieve simultaneous labeling and multi-channel imaging of multiple tumor markers. However, immunofluorescence technology also has obvious disadvantages, such as the instability of fluorescence signals, background fluorescence in tissues, the dimensionless nature of fluorescence signal intensity that makes absolute quantification impossible, and the inability to be used together with HE staining and immunohistochemistry.

[0007] Conventional magnetic resonance imaging is widely used in clinical medicine, but its application at the tissue level is limited due to its low spatial resolution. The magnetic resonance technology based on nitrogen vacancy color centers (NV color centers) in diamond developed in recent years provides a very attractive microscopic magnetism method. NV color centers are used as a quantum magnetic sensor that can detect magnetic fields in nearby samples with high sensitivity, high resolution and high stability, and due to the good biocompatibility of diamond materials, they are very suitable for the detection of biological samples. The use of NV color centers has achieved magnetic imaging of biological samples from nanometer resolution to micrometer resolution, but due to some technical obstacles, magnetic imaging (or magnetic resonance imaging) with micrometer resolution or subcellular resolution at the tissue level has not yet been achieved. Summary of the invention

[0008] In view of this, the main purpose of the present invention is to provide a method to partially solve at least one of the above-mentioned technical problems.

[0009] In order to achieve the above object, as one aspect of the present invention, a method for magnetic imaging of tumor tissue with micrometer resolution is provided, comprising:

[0010] Labeling a tissue sample by superparamagnetic nanoparticles, wherein the superparamagnetic nanoparticles are labeled on a tumor marker protein in the tissue sample;

[0011] attaching diamond to the tissue sample to form a diamond-tumor tissue slice-cover glass structure, wherein the tumor tissue slice belongs to the tissue sample;

[0012] Using the NV color center two-dimensional magnetic sensor in the diamond to measure the magnetic field of the tissue sample to obtain a magnetic field image of the tissue sample; and

[0013] The magnetic field image is processed by a deep learning model to obtain a micron-level resolution magnetic imaging image of the tissue sample.

[0014] According to one embodiment of the present invention, the step of labeling the tissue sample with superparamagnetic nanoparticles further comprises:

[0015] The tissue sample is subjected to immunomagnetic labeling treatment, wherein the immunomagnetic labeling includes a superparamagnetic nanoparticle incubation process so that the tumor marker protein in the tissue sample is combined with the superparamagnetic nanoparticle specific label.

[0016] According to an embodiment of the present invention, it also includes:

[0017] The diamond is processed to form a high-density NV color center with a depth of 10 nm to 110 nm in a first surface of the diamond, and the first surface of the diamond is in contact with the tissue slice.

[0018] According to an embodiment of the present invention, attaching the diamond to the tissue sample further comprises:

[0019] In the diamond-tumor tissue slice-cover glass structure, ultraviolet glue is dripped into the tumor tissue slice test area to make the upper surface of the diamond contact the tumor tissue slice test area; and

[0020] A metal clamp is used to uniformly apply force to the diamond-tumor tissue slice-cover glass structure, and the force direction is perpendicular to the second surface of the diamond and the cover glass surface in contact with the metal clamp, so that the pretreated tissue slice and the diamond are closely attached, and the second surface of the diamond is the surface opposite to the first surface of the diamond.

[0021] According to an embodiment of the present invention, it also includes:

[0022] Attaching the diamond-tumor tissue slice-cover glass structure as a whole to a dove prism, wherein the lower surface of the diamond is placed on the dove prism;

[0023] The diamond-tumor tissue slice-cover glass structure is glued to the Dove prism using ultraviolet glue; wherein the diamond-tumor tissue slice-cover glass structure is placed at the center of the Dove prism; and

[0024] The diamond-tumor tissue slice-cover glass structure is properly pressed to make the diamond-tumor tissue slice-cover glass structure parallel to the Dove prism.

[0025] According to one embodiment of the present invention, the superparamagnetic nanoparticle labeling of tissue samples further comprises:

[0026] Before the superparamagnetic nanoparticles are used to label tissue samples, the coverslip is pretreated with poly-lysine, and the tumor tissue slices are placed on the coverslip to fix the tumor tissue slices on the coverslip.

[0027] According to an embodiment of the present invention, it also includes:

[0028] The laser light is refracted onto the diamond through the Dove prism so as to excite the NV color center in the diamond.

[0029] According to an embodiment of the present invention, it also includes:

[0030] The superparamagnetic nanoparticles are magnetized by an external magnetic field.

[0031] According to an embodiment of the present invention, it also includes:

[0032] Using microwaves, the spin state of the NV color center in the diamond is controlled.

[0033] According to an embodiment of the present invention, it also includes:

[0034] A CMOS camera is used to collect fluorescence data of the NV color center in the diamond to obtain a magnetic field image.

[0035] According to an embodiment of the present invention, it also includes:

[0036] The tissue sample to be tested may be other types of biological tissue samples except tumor tissue samples.

[0037] The tumor tissue magnetic imaging method provided by the present invention has high signal stability, low background, and can achieve absolute quantification. It can realize magnetic and optical multimodal correlation imaging of biological tissues. Both magnetic and optical imaging can achieve micron-level spatial resolution. It is also compatible with commercial optical microscopes and can be easily promoted and used. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] The above and other objects, features and advantages of the present invention will become more apparent through the following description of the embodiments of the present invention with reference to the accompanying drawings, in which:

[0039] Figure 1 A flow chart of an embodiment of the present invention is schematically shown;

[0040] Figure 2 A schematic diagram showing the labeling of tumor marker proteins by superparamagnetic particles in the tumor tissue magnetic imaging method of the present invention is shown schematically;

[0041] Figure 3 A schematic diagram of a two-dimensional magnetic sensor of NV color centers in diamond used in the present invention is schematically shown;

[0042] Figure 4 The schematic diagram shows the structure of the diamond-tumor tissue slice-cover glass structure of the present invention being bonded to the Dove prism;

[0043] Figure 5 The schematic diagram schematically shows that the superparamagnetic nanoparticles used in the present invention generate a weak magnetic field after being magnetized;

[0044] Figure 6 The figure schematically shows a device for imaging using a tumor tissue magnetic imaging method according to the present invention. DETAILED DESCRIPTION

[0045] In order to make the objectives, technical solutions and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with specific embodiments and with reference to the accompanying drawings.

[0046] The terms used herein are only for describing specific embodiments and are not intended to limit the present invention. The terms "include", "comprises", etc. used herein indicate the existence of the features, steps, operations and / or components, but do not exclude the existence or addition of one or more other features, steps, operations or components.

[0047] All terms (including technical and scientific terms) used herein have the meanings commonly understood by those skilled in the art, unless otherwise defined. It should be noted that the terms used herein should be interpreted as having a meaning consistent with the context of this specification, and should not be interpreted in an idealized or overly rigid manner.

[0048] When using expressions such as "at least one of A, B, and C, etc.", they should generally be interpreted according to the meaning of the expression commonly understood by those skilled in the art (for example, "a system having at least one of A, B, and C" should include but is not limited to a system having A alone, B alone, C alone, A and B, A and C, B and C, and / or A, B, C, etc.).

[0049] The present invention provides a method for magnetic imaging of tumor tissue. Figure 1 The flowchart of the embodiment of the present invention is schematically shown.

[0050] like Figure 1 As shown, the method includes operations S101-104.

[0051] In operation S101 , a tissue sample is labeled with superparamagnetic nanoparticles, wherein the superparamagnetic nanoparticles are labeled on tumor marker proteins in the tissue sample.

[0052] In operation S102, diamond is attached to the tissue slice to form a diamond-tumor tissue slice-cover glass structure; wherein, since conventional glass slides are too thick, which will affect the microwave application and light transmission effects, and the cover glass is thinner than the conventional glass slide, the tumor tissue slice is attached to the cover glass to facilitate subsequent imaging.

[0053] In operation S103 , a magnetic field of the tissue sample is measured using a two-dimensional magnetic sensor of an NV color center in diamond to obtain a magnetic field image of the tissue sample.

[0054] In operation S104, the magnetic field image is processed by a deep learning model to obtain a magnetic imaging image of the tissue sample.

[0055] According to an embodiment of the present invention, before performing operation S101, the cover glass is pretreated with poly-lysine, and the tumor tissue slice is placed on the cover glass, so that the tumor tissue slice is fixed on the cover glass.

[0056] More specifically, tissue samples can be prepared through the following process, and the preparation process can be frozen sections or paraffin-embedded sections; for frozen sections, conventional OCT-embedded tumor tissue is used for frozen sections, and a section thickness of 5 to 10 μm is selected; for paraffin-embedded sections, tissue samples need to undergo conventional fixation, dehydration and embedding before sectioning, and a section thickness of 5 μm is selected. Paraffin-embedded sections also need to undergo antigen repair before tumor tissue magnetic imaging operations can be performed; tumor tissue sections are attached to coverslips, and the thickness of the coverslips can be between 0.16 and 0.19 mm. The coverslips need to be pretreated with poly-L-lysine and soaked in 0.1 mg / ml poly-L-lysine for half an hour before the tumor tissue sections can be fixed on the coverslips.

[0057] According to an embodiment of the present invention, the tissue sample to be tested may be other types of biological tissue samples except tumor tissue samples.

[0058] More specifically, other types of biological tissue samples need to contain corresponding biomarkers, which may be characteristic proteins. The corresponding biomarkers may be immunomagnetically labeled, and other types of biological tissue samples may then utilize the micron-resolution tumor tissue magnetic imaging method of the present invention to obtain magnetic imaging images.

[0059] Figure 2 The schematic diagram of the tumor marker protein being labeled by superparamagnetic particles in the tumor tissue magnetic imaging method according to one embodiment of the present invention is schematically shown. The process of preparing the immunomagnetic labeled tissue sample is as follows:

[0060] (1) Place the prepared tissue sample in a culture dish or a moist box and wash it three times with phosphate buffered saline (PBS).

[0061] (2) Fixation: Fix in 4% paraformaldehyde (PFA) at room temperature for 15 minutes, or fix in -20°C pre-frozen 100% methanol for 5 minutes.

[0062] (3) Wash with phosphate-buffered saline (PBS) three times, 5 minutes each time, for a total of 15 minutes.

[0063] (4) Blocking: Block at room temperature for 30 minutes. The blocking solution formula is: 2% bovine serum albumin (BSA), 0.1% Triton X-100 (Triton X-100), 1× PBS. The pH value of the blocking solution is maintained at 7.4.

[0064] (5) Primary antibody incubation: Incubate the blocked tissue samples in the primary antibody solution at 4°C overnight. The antibody incubation solution formula is: 1% BSA, 0.1% Triton X-100, 1× PBS, pH 7.4.

[0065] (6) The next day, the cells were washed five times with phosphate-buffered saline (PBS), each time for 5 minutes, for a total of 25 minutes.

[0066] (7) Biotin-secondary antibody incubation: Incubate the tissue sample in a biotin-modified secondary antibody solution at room temperature for 1 hour. The biotin secondary antibody can be goat anti-mouse IgG H&L (Biotin) (Abcam, ab6788). The concentration of the antibody working solution is 2 μg / ml, i.e., a 1:1000 dilution. The formula of the antibody incubation solution is: 1% BSA, 0.1% Triton X-100, 1× PBS, pH 7.4.

[0067] (8) Wash with phosphate-buffered saline (PBS) five times, 5 minutes each time, for a total of 25 minutes.

[0068] After the above operations (1)-(8) are completed, a labeled intermediate product is obtained in which a primary antibody factor is bound to the tumor marker protein in the tissue sample, a secondary antibody factor is bound to the primary antibody factor, and the secondary antibody factor is modified with biotin.

[0069] (9) Magnetic particle incubation: Incubate the tissue sample in a solution of streptavidin-coated superparamagnetic nanoparticles at room temperature for 30 minutes. The concentration of the magnetic particle working solution is 50 μg Fe / ml, i.e., a 1:20 dilution. The incubation solution formula is: 0.1% Triton X-100, 1× PBS, pH 7.4.

[0070] (10) Wash with phosphate buffered saline (PBS) five times, 5 minutes each time, for a total of 25 minutes; during this period, stain the cell nucleus with 4',6-diamidino-2-phenylindole (DAPI) for 5 minutes. Optionally, stain the cell nucleus with 4',6-diamidino-2-phenylindole (DAPI) for 5 minutes during the third or fourth wash with phosphate buffered saline (PBS) to ensure adequate staining of the cell nucleus.

[0071] After the above operations (1)-(10) are completed, we get Figure 2 The schematic diagram of tumor marker proteins labeled with superparamagnetic particles is shown, wherein the superparamagnetic nanoparticles are coated in streptavidin, and streptavidin and biotin modified on the secondary antibody factor undergo antibody-antigen reaction and specifically bind to each other, so that the superparamagnetic nanoparticles are labeled on the tumor marker proteins in the tissue sample.

[0072] According to an embodiment of the present invention, the diamond used in operation S102 may be a diamond having a high-density NV color center with a depth of 10 nm to 110 nm formed in the first surface.

[0073] More specifically, Figure 3 The schematic diagram of the NV color center two-dimensional magnetic sensor in diamond used in the present invention is shown schematically, in which the first plane in the figure is set as the xoy plane, and the z direction is set perpendicular to the xoy plane; 14N is implanted into the bulk diamond of electronic grade purity and

[100] crystal orientation + Ions are injected into diamond at different energies and concentrations to form five interconnected ion layers. The ion concentration of each layer is 10^13 / cm 2 After annealing at 1000°C for 4 hours, a high-density NV color center with a depth of 10nm to 110nm is formed in the first surface of the diamond, and the concentration of the NV color center is about 1×10^12 / cm 2 Such NV color center two-dimensional magnetic sensors in diamond have better magnetic sensitivity and measurement efficiency, and the z-direction thickness of 100nm does not affect the realization of micron resolution.

[0074] According to an embodiment of the present invention, in operation S102, forming the diamond-tumor tissue slice-cover glass structure comprises the following steps:

[0075] (1) Rapidly washing the tissue sample labeled with superparamagnetic nanoparticles with deionized water and blowing the tissue sample dry;

[0076] (2) dripping ultraviolet glue on the tissue sample to be tested area, buckling the first surface of the diamond on the tissue sample to be tested area, and using a metal clamp to uniformly apply force to the diamond-tumor tissue slice-cover glass structure, the force direction is perpendicular to the second surface of the diamond in contact with the metal clamp and the cover glass surface, so that the pretreated tissue slice and the diamond are closely attached, and the second surface of the diamond is the surface opposite to the first surface of the diamond;

[0077] (3) Place the bonded diamond and tissue slice under a UV lamp and irradiate with UV light for 5 minutes to cure the UV glue;

[0078] (4) Apply UV glue to the center of the upper surface of the dove prism, buckle the second surface of the diamond on the UV glue, adjust the position of the diamond and appropriately press the diamond-tumor tissue slice-cover glass structure so that the diamond-tumor tissue slice-cover glass structure is parallel to the dove prism and is located in the center of the dove prism.

[0079] (5) Place the Dove prism with the diamond-tumor tissue slice-cover glass structure under a UV lamp and irradiate with UV light for 5 minutes to solidify the UV glue.

[0080] (6) Use a glass cutter to remove the cover glass outside the diamond and clean the glass fragments with a brush and an ear suction bulb.

[0081] Figure 4 The schematic diagram of the structure of the diamond-tumor tissue slice-cover glass structure of the present invention bonded to the Dove prism is shown schematically. The superiority of the two-dimensional magnetic sensor based on the diamond NV color center mainly comes from the sub-nanometer size, high sensitivity, high resolution, high stability and good biocompatibility of the NV color center quantum magnetic sensor. In order to give full play to these advantages, the tissue sample needs to be close to the magnetic sensor. After the formation of the above-mentioned diamond-tumor tissue slice-cover glass structure, the tissue slice is tightly bonded to the first surface of the diamond, and at least the bonding state does not change during the entire measurement process, so that the superparamagnetic nanoparticles on the tumor marker protein can be easily detected by the NV color center two-dimensional magnetic sensor in the diamond.

[0082] According to an embodiment of the present invention, in operation S103, the method further includes:

[0083] The laser is refracted onto diamond through a Dove prism to excite the NV color center in the diamond.

[0084] More specifically, a laser with a wavelength of 532nm is used to excite the NV color center in the diamond. Since a larger imaging area can provide richer and more global tumor tissue information, a larger area of ​​the tissue sample to be tested requires a larger power of the laser used to excite the NV color center. Therefore, the laser cannot be transmitted through the objective lens, and it is necessary to refract the laser onto the diamond through a Dove prism to excite the NV color center in the diamond for magnetic measurement.

[0085] According to an embodiment of the present invention, in operation S103, the method further includes:

[0086] The superparamagnetic nanoparticles are magnetized by an external magnetic field.

[0087] More specifically, an external magnetic field is applied to magnetize the superparamagnetic nanoparticles marked on the tissue sample, and the magnetic field can be generated by a permanent magnet. The external magnetic field of the magnet and the magnetic field generated by the magnetized superparamagnetic nanoparticles can increase the Zeeman effect of the NV color center in the diamond. The weak magnetic field of the tissue sample can be obtained by subtracting the external magnetic field from the total static magnetic field measured.

[0088] Figure 5The schematic diagram shows that the superparamagnetic nanoparticles used in the present invention generate a weak magnetic field after being magnetized. The NV color center two-dimensional magnetic sensor in the diamond can measure the magnetic field of tumor tissue attached to the surface of the diamond. The magnetic field can cause the NV color center in the diamond to undergo Zeeman splitting, and the fluorescence count decreases at the resonance frequency of the continuous wave spectrum (CW spectrum). Therefore, the total static magnetic field information can be inferred by measuring the Zeeman splitting resonance frequency. The magnetic field image of the tissue sample can be obtained by fitting and reconstructing the fluorescence signals of each pixel in the imaging area.

[0089] According to an embodiment of the present invention, in operation S103, the method further includes:

[0090] Using microwaves, the spin state of the NV color center in the diamond is controlled.

[0091] More specifically, by placing microwave lines on the coverslip surface above the diamond-tumor tissue slice-cover glass structure, controllable microwaves can be generated; by changing the microwave frequency, the spin state of the NV color center can be controlled, the fluorescence of the NV color center in different spin states can be read and a continuous wave spectrum (CW spectrum) can be obtained, thereby achieving the measurement of the magnetic field felt by the NV color center.

[0092] An embodiment of the present invention can use a CMOS camera to collect data, ensuring high sampling efficiency, and the pixel scale of 100-300nm ensures the realization of micron-level imaging resolution. The core optical path of the magnetic imaging system uses the optical path of an optical microscope, which is equipped with a multi-channel fluorescence imaging and bright field imaging system, which can meet the multi-modal correlation imaging of magnetism and light, and can be used in combination with most conventional optical imaging methods. The imaging area can be sub-millimeter, and the imaging resolution can be micron or sub-micron.

[0093] An embodiment of the present invention can refer to the single-cell segmentation and statistical software in optical imaging to perform inverse reconstruction of magnetic images, carry out magnetic quantification of tumor markers at the single-cell level, and use the cell nucleus labeled with 4',6-diamidino-2-phenylindole (DAPI) as important auxiliary information for single-cell analysis.

[0094] Figure 6 The figure schematically shows a device for imaging using a tumor tissue magnetic imaging method according to the present invention.

[0095] like Figure 6 As shown, in operation S103, it also includes:

[0096] (1) Using an optical microscope, find the tissue sample area to be tested in the diamond-tumor tissue slice-cover glass structure, focus, and collect a fluorescent image of 4',6-diamidino-2-phenylindole (DAPI). The collected fluorescent image of 4',6-diamidino-2-phenylindole (DAPI) represents the fluorescent image collected of the cell nucleus;

[0097] (2) Switch the optical microscope detection channel to the NV fluorescence channel and adjust the laser size and the angle of incidence on the Dove prism to make the fluorescence intensity in the field of view large and evenly distributed;

[0098] (3) By setting parameters such as microwave frequency range, camera exposure time, and data accumulation times, the LabVIEW program is used to control the synchronization of the CMOS camera and microwave source to complete the NV color center fluorescence data acquisition;

[0099] (4) Processing the collected NV color center fluorescence data through the MATLAB program, accumulating the data at each pixel, and obtaining the continuous wave spectrum (CW spectrum) of each pixel after accumulation;

[0100] (5) By Lorentz fitting the continuous wave spectrum (CW spectrum), the corresponding NV color center frequency diagram can be obtained;

[0101] (6) fitting the frequency graph by a polynomial to obtain a fitting result of the total static magnetic field, and deducting the frequency graph of the external magnetic field to obtain a frequency graph of the tissue sample;

[0102] (7) converting the frequency map of the tissue sample into a magnetic field image to obtain the magnetic field image of the tissue sample;

[0103] (8) A deep learning model called a conditional generative adversarial network is used to train the model and establish a mapping relationship between the magnetic field image and the corresponding magnetic particle marker protein, so as to realize the recognition and inverse analysis of the magnetic field signal and obtain the magnetic imaging map of the tissue sample. The magnetic imaging map can obtain intuitive information on the distribution and expression level of tumor marker proteins in the tissue sample.

[0104] The tumor tissue magnetic imaging method according to the above embodiment of the present invention uses superparamagnetic nanoparticles as magnetic signal labels to replace the fluorescent labels and enzyme labels in the traditional optical method, and indirectly marks the superparamagnetic nanoparticles on the tumor marker protein of the tissue sample through the antibody-antigen reaction; the superparamagnetic nanoparticles are detected by the NV color center two-dimensional magnetic sensor in the diamond, and the magnetic imaging of the tumor tissue with micron-level resolution can be achieved. The tumor tissue magnetic imaging method of the above embodiment of the present invention has high signal stability, low background, and can achieve absolute quantification. Because magnetism is another physical quantity different from light, electricity, etc., the tumor tissue magnetic imaging method of the above embodiment of the present invention can realize the multimodal correlation imaging of magnetism and light of biological tissues. The magnetic imaging device of the present invention is compatible with optical microscopes, and the present invention proposes a complete set of solutions from sample preparation to magnetic detection, and then to image processing and analysis, which is easy to promote and use.

[0105] The flow chart and block diagram in the accompanying drawings illustrate the possible architecture, function and operation of the system, method and computer program product according to various embodiments of the present invention. In this regard, each box in the flow chart or block diagram can represent a module, a program segment, or a part of a code, and the above-mentioned module, program segment, or a part of a code contains one or more executable instructions for realizing the specified logical function. It should also be noted that in some alternative implementations, the functions marked in the box can also occur in a different order from the order marked in the accompanying drawings. For example, two boxes represented in succession can actually be executed substantially in parallel, and they can sometimes be executed in the opposite order, depending on the functions involved. It should also be noted that each box in the block diagram or flow chart, and the combination of the boxes in the block diagram or flow chart can be implemented with a dedicated hardware-based system that performs a specified function or operation, or can be implemented with a combination of dedicated hardware and computer instructions.

[0106] It will be appreciated by those skilled in the art that the features described in the various embodiments and / or claims of the present invention may be combined and / or combined in various ways, even if such combinations and / or combinations are not explicitly described in the present invention. In particular, the features described in the various embodiments and / or claims of the present invention may be combined and / or combined in various ways without departing from the spirit and teachings of the present invention. All of these combinations and / or combinations fall within the scope of the present invention.

[0107] The embodiments of the present invention are described above. However, these embodiments are only for the purpose of illustration, and are not intended to limit the scope of the present invention. Although each embodiment is described above, it does not mean that the measures in each embodiment cannot be used in combination. The scope of the present invention is defined by the attached claims and their equivalents. Without departing from the scope of the present invention, those skilled in the art may make various substitutions and modifications, which should all fall within the scope of the present invention.

Claims

1. A method for magnetic imaging of tumor tissues with micron resolution, comprising: labeling a tissue sample with superparamagnetic nanoparticles, wherein the superparamagnetic nanoparticles are labeled on tumor marker proteins in the tissue sample; attaching diamond to the tissue sample to form a diamond - tumor tissue section - coverslip structure, and the tumor tissue section belongs to the tissue sample; using a nitrogen - vacancy color center (NV color center) two - dimensional magnetic sensor in the diamond to measure the magnetic field of the tissue sample, and obtaining a magnetic field image of the tissue sample; and processing the magnetic field image through a deep - learning model to obtain a magnetic imaging map of the tissue sample with micron resolution.

2. The method according to claim 1, wherein, the step of labeling the tissue sample with superparamagnetic nanoparticles further comprises: performing immunomagnetic labeling on the tissue sample, and the immunomagnetic labeling includes a superparamagnetic nanoparticle incubation process to enable specific labeling and binding of tumor marker proteins in the tissue sample with the superparamagnetic nanoparticles.

3. The method according to claim 1, further comprising: processing the diamond to form high - density NV color centers with a depth of 10 nm to 110 nm in the first surface of the diamond, and the first surface of the diamond is in contact with the tissue section.

4. The method according to claim 3, further comprising: in the diamond - tumor tissue section - coverslip structure, dropping ultraviolet glue on the area to be measured of the tumor tissue section, so that the first surface of the diamond contacts the area to be measured of the tumor tissue section; and using a metal fixture to apply a uniform force to the diamond - tumor tissue section - coverslip structure, and the force application direction is perpendicular to the second surface of the diamond and the coverslip surface contacted by the metal fixture, so that the tissue section and the diamond are closely attached, and the second surface of the diamond is the surface opposite to the first surface of the diamond.

5. The method according to claim 1, further comprising: attaching the whole diamond - tumor tissue section - coverslip structure to a Dove prism, wherein the lower surface of the diamond is placed on the Dove prism; using ultraviolet glue for pasting between the diamond - tumor tissue section - coverslip structure and the Dove prism; wherein the diamond - tumor tissue section - coverslip structure is placed at the center of the Dove prism; and appropriately pressing the diamond - tumor tissue section - coverslip structure to make the diamond - tumor tissue section - coverslip structure parallel and closely attached to the Dove prism.

6. The method according to claim 1, further comprising: pretreating the coverslip with polylysine before the operation of labeling the tissue sample with superparamagnetic nanoparticles, and placing the tumor tissue section on the coverslip to fix the tumor tissue section on the coverslip.

7. The method according to claim 5, further comprising: refracting laser to the diamond through the Dove prism to excite the NV color centers in the diamond.

8. The method according to claim 1, further comprising: using an external magnetic field to magnetize the superparamagnetic nanoparticles through the external magnetic field. Using microwaves, the spin state of the NV color center in the diamond is controlled.

9. The method according to claim 1, further comprising: include: A complementary metal oxide semiconductor (CMOS) camera is used to collect fluorescence data of the NV color center in the diamond to obtain a magnetic field image.

10. The method according to any one of claims 1 to 9, further comprising: include: The tissue sample to be tested may be other types of biological tissue samples except tumor tissue samples.

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