Prism-type laser-excited fluorescence detection system

Abstract

This invention discloses a prism-type laser-excited fluorescence detection system, relating to the field of bioimaging technology. The system includes: a thin-film window, a fluorescent membrane, a prism, a laser, and a camera. The fluorescent membrane is positioned below the thin-film window and adheres to the top surface of the prism. The thin-film window holds the biological sample to be detected. The laser emitted by the laser, with the side of the prism as its incident surface, refracts upon entering the incident surface and strikes the fluorescent membrane, generating a fluorescence signal. This allows the camera to detect the fluorescence signal illuminating the biological sample, thus creating a bioimaging effect. The laser is configured with a first incident angle such that when the laser strikes the fluorescent membrane, total internal reflection occurs within the fluorescent membrane. The system provided by this invention can avoid laser damage or interference with the biological sample.

Description

Technical Field

[0001] This invention relates to the field of bioimaging technology, and in particular to a prism-type laser-excited fluorescence detection system. Background Technology

[0002] QDM imaging (Quantum Dot Magnetic) is a bio-imaging technique that uses magnetic nanoparticles (MNPs) labeled and attached to NV color centers. By leveraging the fact that the spin quantum states of the luminescent defects of the NV color centers in diamond are easily affected by the surrounding microwave and static magnetic fields, and can be read out using lasers, the distribution of the magnetic or microwave field around the biological sample can be measured. This enables quantitative and non-destructive biomagnetic imaging, featuring high spatial resolution, a large field of view, a wide dynamic range of detectable magnetic fields, and fast imaging speed.

[0003] In practical applications, the lens is aligned with the sample wells on the multi-well plate, and pure gas is pressurized by introducing it into the lens-sample well cavity. This causes the bottom of the sample multi-well cavity to adhere to the fluorescent membrane, enabling the detection of batches of biological samples. The fluorescent membrane contains material with NV centers, or other types of membranes can be used.

[0004] NV centers (Nitrogen-Vacancy Centers) are nitrogen-vacancy defect centers, a common type of solid-state qubit widely used in quantum information science and nano-optics. To excite NV centers to produce light, a common method is optical excitation, where a laser of appropriate wavelength is irradiated onto the NV center. A common excitation wavelength is 532 nm. After the laser passes through the center, the photon energy is transferred to the electronic levels of the NV center. This leads to transitions in the electronic levels of the NV center. When a transition occurs between the ground-state and excited-state electronic levels of the NV center, photons are emitted. The wavelengths of these emitted photons are typically in the infrared range, approximately 640-800 nm.

[0005] Traditional laser optical paths use optical fibers or various mirrors to guide the laser beam onto the NV color center. In this case, a technical solution is needed to avoid laser damage or impact on biological samples. Summary of the Invention

[0006] This invention aims to at least partially address one of the technical problems in related technologies. Therefore, one objective of this invention is to provide a prism-type laser-excited fluorescence detection system to avoid laser damage or interference with biological samples.

[0007] According to a first aspect of the present invention, a prism-type laser-excited fluorescence detection system is provided, the system comprising: a thin-film window, a fluorescent film, a prism, a laser, and a camera; wherein,

[0008] The fluorescent film is placed below the thin film window and is attached to the top surface of the prism;

[0009] The thin film window is used to hold the biological sample to be tested;

[0010] The laser emitted by the laser has the side of the prism as the incident surface. After entering the incident surface, it is refracted and incident on the fluorescent film to generate a fluorescent signal, so that the camera can detect the fluorescent signal illuminating the biological sample for bioimaging.

[0011] The laser is configured to have a first incident angle, which satisfies the condition of being greater than or equal to a second incident angle. The second incident angle is determined based on the angle between the incident surface and the top surface, the refractive index of the laser in air, and the refractive index of the laser in the prism, such that when the laser is incident on the fluorescent film, total internal reflection occurs in the fluorescent film.

[0012] Optionally, the second incident angle is determined as follows:

[0013]

[0014] Where, θ i θ represents the second incident angle; θ0 represents the angle between the incident surface and the top surface; n1 represents the refractive index of the laser in air; n2 represents the refractive index of the laser in the prism.

[0015] Optionally, when the biological sample is placed on a porous plate and the fluorescent membrane is attached to the transparent membrane of the porous plate, the second incident angle is determined as follows:

[0016]

[0017] Where, θ i θ represents the second incident angle; θ0 represents the angle between the incident surface and the top surface; n1 represents the refractive index of the laser in air; n2 represents the refractive index of the laser in the prism; n3 represents the refractive index of the laser in the transparent film, and n3 < n2.

[0018] Optionally, the system may also include an objective lens, a lens, and a filter;

[0019] The objective lens is located above the thin-film window and is used to collect the fluorescence signal, which is then focused by the lens and transmitted to the camera.

[0020] The filter is disposed between the lens and the camera.

[0021] Optionally, the angle between the incident surface and the top surface is an acute angle.

[0022] Optionally, the first incident angle is the sum of the angle between the incident surface and the top surface and the refraction angle of the laser entering the prism.

[0023] Optionally, the target angle of the first incident angle is determined in the following manner:

[0024] Using the second incident angle as the initial angle of the first incident angle, the angle of the first incident angle is adjusted according to a preset step starting from the initial angle;

[0025] The monitoring power of the laser at the exit surface of the prism after each adjustment is determined, and the laser power incident on the fluorescent film is determined based on the determined monitoring power and the corresponding incident power.

[0026] The target angle of the first incident angle is determined based on the maximum power among all laser powers.

[0027] Optionally, when the incident surface and the exit surface are symmetrically arranged with respect to the vertical plane containing the center of the fluorescent film, the laser power incident on the fluorescent film is determined as follows:

[0028] The first transmittance of the laser at the incident surface is calculated using the first incident angle, the refractive index of the laser in air and the refractive index of the laser in the prism, and the incident laser polarization component at the incident surface, based on Fresnel's formula.

[0029] The second transmittance of the laser incident on the fluorescent film is calculated using the first incident angle, the angle between the top surface of the prism and the incident surface, the refractive index of the laser in the fluorescent film and the refractive index of the laser in the prism, and the incident laser polarization component on the surface of the laser incident on the fluorescent film, according to the Fresnel formula.

[0030] The third transmittance of the laser at the exit surface of the prism is calculated using the exit angle of the laser at the exit surface of the prism, the refractive index of the laser in air and the refractive index of the laser in the prism, and the polarization component of the incident laser at the exit surface, based on Fresnel's formula.

[0031] The laser power incident on the fluorescent film is determined as follows:

[0032]

[0033] Wherein, P represents laser power; a represents the first transmittance; b represents the second transmittance; c represents the third transmittance; P1 represents the determined monitoring power; and P0 represents the incident power corresponding to the determined monitoring power.

[0034] Optionally, the system also includes a protective frame and a radiating structure;

[0035] The fluorescent film is installed in the middle of the protective frame, and the height of the protective frame is higher than the height of the fluorescent film;

[0036] The radiating structure is installed on the outside of the protective frame.

[0037] Optionally, the system also includes a prism holder and a coating;

[0038] The prism is mounted on the prism bracket, and the coating is connected to the prism bracket and the system grounding electrode.

[0039] In the solution provided by the embodiments of the present invention, a prism is set at the bottom of the fluorescent membrane so that the laser can enter the fluorescent membrane through the refraction of the prism. Furthermore, by setting the first incident angle to satisfy the angle condition determined by the second incident angle, the laser undergoes internal total internal reflection in the fluorescent membrane. On the one hand, this can improve the fluorescence excitation effect, and on the other hand, it can avoid the laser irradiating the biological sample and damaging or affecting the biological sample.

[0040] Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. Attached Figure Description

[0041] Figure 1 This is a schematic diagram of the structure of the first prism-type laser-excited fluorescence detection system provided in the embodiments of the present invention;

[0042] Figure 2a This is a schematic diagram of the structure of the second prism-type laser-excited fluorescence detection system provided in the embodiments of the present invention;

[0043] Figure 2b This is a schematic diagram of the structure of the third prism-type laser-excited fluorescence detection system provided in the embodiments of the present invention;

[0044] Figure 3a This is a schematic diagram of the structure of the first type of prism provided in the embodiment of the present invention;

[0045] Figure 3b This is a schematic diagram of the structure of the second type of prism provided in an embodiment of the present invention;

[0046] Figure 3c This is a schematic diagram of the structure of the third type of prism provided in the embodiment of the present invention;

[0047] Figure 4a This is a schematic diagram of a scenario for the first laser power detection method provided in an embodiment of the present invention;

[0048] Figure 4b This is a schematic diagram of a scenario for the second laser power detection method provided in this embodiment of the invention. Detailed Implementation

[0049] Embodiments of the present invention are described in detail below, examples of which are illustrated in the accompanying drawings, wherein the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are exemplary and intended to explain the present invention, and should not be construed as limiting the present invention.

[0050] The prism-type laser-excited fluorescence detection system of the present invention is described below with reference to the accompanying drawings.

[0051] In one embodiment of the present invention, see Figure 1 A prism-type laser-excited fluorescence detection system is provided, comprising: a thin-film window, a fluorescent film, a prism, a laser, and a camera; wherein,

[0052] The fluorescent film is placed below the thin film window and is attached to the top surface of the prism;

[0053] A thin-film window is used to hold the biological sample to be tested;

[0054] The laser emitted by the laser has the side of the prism as the incident surface. After entering the incident surface, it is refracted and incident on the fluorescent film to generate a fluorescent signal, so that the camera can detect the fluorescent signal to illuminate the biological sample for bioimaging.

[0055] The laser is configured to have a first incident angle, which satisfies the condition of being greater than or equal to a second incident angle. The second incident angle is determined based on the angle between the incident surface and the top surface, the refractive index of the laser in air, and the refractive index of the laser in the prism, so that when the laser is incident on the fluorescent film, internal total internal reflection occurs in the fluorescent film.

[0056] exist Figure 1 In the embodiments shown, the camera can specifically be a CMOS (Complementary Metal Oxide Semiconductor) camera.

[0057] In one embodiment of the present invention, a diamond sheet containing NV color centers can be used as a fluorescent film.

[0058] like Figure 1As shown, the thin-film window can be a multi-well plate, with several thin-film windows containing several biological samples to be tested. By moving the multi-well plate, the thin-film windows move sequentially above the fluorescent membrane, and the biological samples are measured and analyzed based on the fluorescence signal generated by the fluorescent membrane. A prism is placed at the bottom of the fluorescent membrane, so that the top surface of the fluorescent membrane is in contact with the top surface of the prism, and the laser light enters the fluorescent membrane through the prism. Figure 1 The incident laser is refracted and reaches the top surface of the prism, illuminating the fluorescent film. The side of the prism on the side of the incident laser is the incident surface, and the side of the prism on the side of the reflected laser after reflection by the fluorescent film is the exit surface. The detector receives the exit laser and obtains the power of the exit laser, i.e., the monitoring power.

[0059] For example Figure 1 As shown, the system also includes an objective lens, a lens, and a filter; the objective lens, located above the thin-film window, is used to collect fluorescence signals, which are then focused by the lens and transmitted to the camera. In this way, bio-imaging of the fluorescence signal can be observed in the camera.

[0060] The filter is positioned between the lens and the camera. On one hand, it filters out light other than the fluorescence signal; on the other hand, it blocks the camera lens, preventing noise light signals from the surrounding area from entering the lens and interfering with the detection.

[0061] Bioimaging can include imaging of microwave fields and imaging of static magnetic fields. For example, microwave field imaging can be performed using a radiating structure composed of microstrip lines. Figure 2a As shown, the system also includes a protective frame and a radiating structure; a fluorescent film is installed in the middle of the protective frame, the NV color center is inside the fluorescent film, and the height of the protective frame is higher than the height of the fluorescent film; the radiating structure is installed on the outside of the protective frame, thereby providing a microwave field for the measurement of the NV color center. By applying an excitation laser to the NV color center, the magnetic field distribution in the biological sample within the porous plate film window above the diamond sheet can be detected based on the fluorescence signal of the NV color center, or whether the biological sample within the film window contains magnetic beads. By providing a microwave field, the magnitude of the magnetic field at each location can be obtained more specifically, or noise light information can be eliminated by modulating the microwave field, thereby improving the detection sensitivity of the fluorescence signal.

[0062] With a protective frame in place, on the one hand, it facilitates better adhesion between the silicon nitride film in the thin film window and the diamond sheet, and when the thin film window is filled with gas, it can limit and guide the deformation direction of the silicon nitride film; on the other hand, it can protect the diamond sheet from damage.

[0063] Based on the above principles, it is known that no magnetic or electric fields that could interfere with the detection should exist around the NV color center. Since the porous plate needs to be moved to detect samples in the thin film windows at different locations, the movement of the porous plate is prone to generating static electricity. To overcome the interference of static electricity, the system is also equipped with a prism support and a coating. Figure 2b As shown, the prism is mounted on a prism holder, exposing the incident and exit surfaces on both sides of the prism holder. A coating is applied to the top and bottom of the substrate of the base prism holder, and this coating is connected to the prism holder and the system ground electrode to discharge static electricity. In this mounting configuration, the microstrip line, NV color center, radiating structure, and etched area all face upwards towards the prism holder, facilitating camera observation.

[0064] Specifically, for the etched area, the middle of the coating on one side of the substrate with coatings on both sides is etched to remove the coating, leaving the microstrip line exposed, while preventing the microstrip line from connecting with other ground coatings. Alternatively, the microstrip line can be fabricated and attached to the location where the coating has been removed by the substrate etching.

[0065] As described above, in the solution provided by the embodiments of the present invention, by setting a prism at the bottom of the fluorescent membrane, the laser can enter the fluorescent membrane through the refraction of the prism. Furthermore, by setting a first incident angle to satisfy the angle condition determined by the second incident angle, the laser undergoes total internal reflection in the fluorescent membrane. On the one hand, this can improve the fluorescence excitation effect, and on the other hand, it can avoid the laser irradiating the biological sample and damaging or affecting the biological sample.

[0066] In one embodiment, the second incident angle is determined as follows:

[0067]

[0068] Where, θ i θ represents the second incident angle; θ0 represents the angle between the incident surface and the top surface; n1 represents the refractive index of the laser in air; n2 represents the refractive index of the laser in the prism.

[0069] In one embodiment, when the biological sample is placed on a multi-well plate and the fluorescent membrane is attached to the transparent membrane of the multi-well plate, the second incident angle is determined as follows:

[0070]

[0071] Where, θ i θ represents the second incident angle; θ0 represents the angle between the incident surface and the top surface; n1 represents the refractive index of the laser in air; n2 represents the refractive index of the laser in the prism; n3 represents the refractive index of the laser in the transparent film, and n3 < n2.

[0072] In one embodiment, the angle between the incident surface and the top surface is an acute angle. Specifically, as shown in the example... Figure 3aThe prism can have a trapezoidal cross-section, with the upper base being longer than the lower base. In this case, the angle θ0 between the incident surface and the top surface is acute. In other embodiments, the prism can also have an inverted triangular cross-section.

[0073] like Figure 3a , 3b As shown in 3c, the first incident angle is θ. i θ t Let θ be the angle of refraction, and θ0 be the angle between the incident surface and the top surface. Figure 3a The middle case is where the length of the upper base is greater than the length of the lower base. Figure 3b The middle case is where the length of the upper base is equal to the length of the lower base. Figure 3c This refers to the case where the length of the upper base is less than the length of the lower base. Conversely, in... Figure 3a In this design, the angle of incidence is more adjustable; while satisfying the condition of being greater than or equal to the second angle of incidence, the angle of incidence can be made either very large or very small. Other structures, such as... Figure 3b and above Figure 3c As shown, the incident angle of the laser at the incident surface has very little room for adjustment, and the incident angle is generally too large. This leads to excessive reflectivity of the laser power, resulting in insufficient laser power entering the fluorescent film. Consequently, the power of the laser needs to be increased, causing energy waste and affecting the lifespan of various optical components in the optical path.

[0074] by Figure 3a Taking the optimal prism structure as an example, since the refractive index of air is small and close to that of a vacuum, when the laser enters the fluorescent film from the prism, regardless of which has a larger refractive index, as long as the incident angle of the laser entering the fluorescent film from the prism is greater than the critical incident angle for total internal reflection when entering the air from the prism, the laser entering the fluorescent film can undergo total internal reflection.

[0075] Furthermore, in Figure 3a In the prism structure described above, the first incident angle is the sum of the angle between the incident surface and the top surface, and the refraction angle of the laser entering the prism. Therefore, θ in the above embodiment can be obtained. i Two calculation formulas.

[0076] To reduce the laser's reflectivity at the prism's incident surface, in practice, the incident angle of the laser at the prism's incident surface can be set slightly larger than θ. i This is to ensure that the laser entering the fluorescent film can undergo total internal reflection.

[0077] In one embodiment, the target angle of the first incident angle is determined as follows:

[0078] Using the second incident angle as the initial angle of the first incident angle, the angle of the first incident angle is adjusted according to a preset step starting from the initial angle;

[0079] The monitoring power of the laser at the exit surface of the prism after each adjustment is determined, and the laser power incident on the fluorescent film is determined based on the determined monitoring power and the corresponding incident power.

[0080] The target angle of the first incident angle is determined based on the maximum power among all laser powers.

[0081] A step refers to a fixed angle. The adjustment process involves rotating the laser, increasing the initial incident angle by a fixed angle each time, for example, by one degree each time.

[0082] The laser power at the exit surface is obtained by using a thermoelectric laser power meter, which is the monitored power.

[0083] After each adjustment, the incident power corresponding to the monitored power is the laser power at the incident surface when the laser enters the prism from the laser after this adjustment.

[0084] By optimizing through the above adjustment process, the first incident angle corresponding to the optimal laser power can be found for detection, thereby reducing power loss and saving energy.

[0085] In addition, when the incident angle of the laser on the prism incident surface is fixed, the incident position of the laser on the incident surface is also fixed for a prism with a defined structure. In addition to the incident angle on the prism incident surface affecting the laser power entering the fluorescent film, the incident angle of the incident laser at the interface between the fluorescent film and the prism, and the laser's travel distance in the prism, also affect the laser power entering the fluorescent film.

[0086] like Figure 4a and Figure 4b As shown, the relative positions of the thin-film window, the fluorescent membrane, and the prism remain unchanged. Figure 4a and Figure 4b The incident surface is the same face of the prism, and the exit surface is also the same face on the other side of the prism. The difference lies in the fact that the angle at which the laser emits the laser rotates, causing changes in the path of the incident and reflected lasers, and correspondingly, the position of the detector receiving the laser also changes.

[0087] Therefore, by obtaining the laser power entering the fluorescent film, the first incident angle of the laser at the prism incident surface can be further optimized.

[0088] To obtain the laser power entering the fluorescent film, the incident and exit surfaces of the prism can first be symmetrically arranged with respect to the vertical plane containing the center of the fluorescent film, such as... Figure 4aThe dashed lines shown are designed so that the path of the incident laser in the prism is equal to the path of the reflected laser. If the incident and exit surfaces of the prism are asymmetrical relative to the vertical plane containing the center of the fluorescent film, when the angles of the incident and reflected lasers change simultaneously, the path of the reflected laser in the prism and its angle at the exit surface will differ from those of the incident laser. This makes it difficult to calculate the laser power entering the fluorescent film based on the laser power monitoring value obtained from the detector.

[0089] When the incident and exit surfaces are symmetrically arranged with respect to the vertical plane containing the center of the fluorescent film, the laser power incident on the fluorescent film is determined as follows:

[0090] The first transmittance of the laser at the incident surface is calculated using the first incident angle, the refractive index of the laser in air and the refractive index of the laser in the prism, and the incident laser polarization component at the incident surface, based on Fresnel's formula.

[0091] The second transmittance of the laser incident on the fluorescent film is calculated using the first incident angle, the angle between the top surface of the prism and the incident surface, the refractive index of the laser in the fluorescent film and the refractive index of the laser in the prism, and the incident laser polarization component on the surface of the laser incident on the fluorescent film, based on the Fresnel formula.

[0092] The third transmittance of the laser at the exit surface of the prism is calculated using the exit angle of the laser at the exit surface of the prism, the refractive index of the laser in air and the refractive index of the laser in the prism, and the polarization component of the incident laser at the exit surface, based on Fresnel's formula.

[0093] The laser power incident on the fluorescent film is determined as follows:

[0094]

[0095] Where P represents laser power; a represents first transmittance; b represents second transmittance; c represents third transmittance; P1 represents the determined monitoring power; and P0 represents the incident power corresponding to the determined monitoring power.

[0096] In the above embodiment, the top surface of the prism is the interface between the prism and the fluorescent film; since the symmetry relationship with respect to the vertical plane where the center of the fluorescent film is located is preset, the exit angle is equal to the first incident angle, which is convenient for measurement and calculation.

[0097] In the above process, the calculation of the first, second, and third transmittance has no specific order; they are listed step by step for ease of explanation.

[0098] To ensure the accuracy of the above measurements and calculations, the preconditions can be controlled in advance to ensure that the fluorescent film is uniformly thick and flatly attached to the top surface of the prism, and that the plane containing the incident, refracted, and reflected laser beams is perpendicular to the side edges of the prism. The prism can be considered as a right prism, and the edge between the two trapezoidal cross sections of the prism is the side edge.

[0099] According to the above method, assuming that all other parameters remain unchanged, the incident angle of the laser on the prism incident surface is adjusted in preset steps, gradually increasing from θ0. Each adjustment calculates the laser power incident on the fluorescent film based on the monitored laser power P1 at the exit surface, thereby selecting the maximum laser power entering the fluorescent film, and determining the incident angle at which the maximum laser power is achieved as the target angle.

[0100] It should be noted that the logic and / or steps represented in the flowchart or otherwise described herein, for example, can be considered as a sequenced list of executable instructions for implementing logical functions, and can be embodied in any computer-readable medium for use by, or in conjunction with, an instruction execution system, apparatus, or device (such as a computer-based system, a processor-included system, or other system that can fetch and execute instructions from, an instruction execution system, apparatus, or device). For the purposes of this specification, "computer-readable medium" can be any means that can contain, store, communicate, propagate, or transmit programs for use by, or in conjunction with, an instruction execution system, apparatus, or device. More specific examples (a non-exhaustive list) of computer-readable media include: an electrical connection having one or more wires (electronic device), a portable computer disk drive (magnetic device), random access memory (RAM), read-only memory (ROM), erasable and editable read-only memory (EPROM or flash memory), fiber optic devices, and portable optical disc read-only memory (CDROM). Alternatively, the computer-readable medium may be paper or other suitable media on which the program can be printed, since the program can be obtained electronically, for example, by optically scanning the paper or other medium, followed by editing, interpreting, or otherwise processing as necessary, and then stored in a computer memory.

[0101] It should be understood that various parts of the present invention can be implemented in hardware, software, firmware, or a combination thereof. In the above embodiments, multiple steps or methods can be implemented in software or firmware stored in memory and executed by a suitable instruction execution system. For example, if implemented in hardware, as in another embodiment, it can be implemented using any one or a combination of the following techniques known in the art: discrete logic circuits having logic gates for implementing logical functions on data signals, application-specific integrated circuits (ASICs) having suitable combinational logic gates, programmable gate arrays (PGAs), field-programmable gate arrays (FPGAs), etc.

[0102] In the description of this specification, references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0103] In the description of this invention, it should be understood that the terms "center," "longitudinal," "lateral," "length," "width," "thickness," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," "outer," "clockwise," "counterclockwise," "axial," "radial," and "circumferential" indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are used only for the convenience of describing this invention and simplifying the description, and are not intended to indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this invention.

[0104] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this invention, "a plurality of" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0105] In this invention, unless otherwise explicitly specified and limited, the terms "installation," "connection," "linking," and "fixing," etc., should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral part; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; they can refer to the internal communication of two components or the interaction between two components, unless otherwise explicitly limited. Those skilled in the art can understand the specific meaning of the above terms in this invention according to the specific circumstances.

[0106] In this invention, unless otherwise explicitly specified and limited, "above" or "below" the second feature can mean that the first feature is in direct contact with the second feature, or that the first feature is in indirect contact with the second feature through an intermediate medium. Furthermore, "above," "over," and "on top" of the second feature can mean that the first feature is directly above or diagonally above the second feature, or simply that the first feature is at a higher horizontal level than the second feature. "Below," "below," and "under" the second feature can mean that the first feature is directly below or diagonally below the second feature, or simply that the first feature is at a lower horizontal level than the second feature.

[0107] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present invention.

Claims

1. A prism laser induced fluorescence detection system, characterized by, The system comprises a thin film window, a fluorescent film, a prism, a laser, a camera, wherein The fluorescent film is placed below the thin film window and adheres to the top surface of the prism; The thin film window is used to hold the biological sample to be detected; The laser emitted by the laser is incident on the side surface of the prism as the incident surface, and after refraction, it is incident on the fluorescent film to generate a fluorescent signal, so that the camera detects the biological imaging of the biological sample irradiated by the fluorescent signal; The laser is arranged to have a first incident angle, which satisfies the condition of being greater than or equal to a second incident angle, the second incident angle is determined based on the included angle between the incident surface and the top surface, the refractive index of laser in air and the refractive index of laser in the prism, so that when the laser is incident on the fluorescent film, internal total reflection occurs in the fluorescent film; The fluorescent film is a diamond sheet containing NV color centers, and the biological imaging is biological magnetic imaging; The target angle of the first incident angle is determined in the following manner: The angle of the second incident angle is used as the initial angle of the first incident angle, and the angle of the first incident angle is adjusted by a preset step from the initial angle; Determine the monitoring power of the laser at the exit surface of the prism after each adjustment, and determine the laser power incident in the fluorescent film based on the determined monitoring power and the corresponding incident power; The target angle of the first incident angle is determined based on the maximum power in each laser power; In the case where the incident surface and the exit surface are symmetrically arranged with respect to the vertical plane where the center of the fluorescent film is located, the laser power incident in the fluorescent film is determined in the following manner: The first transmittance of the laser at the incident surface is calculated according to the Fresnel formula by using the first incident angle, the refractive index of laser in air and the refractive index of laser in the prism, and the incident laser polarization component of the incident surface; The second transmittance of the laser incident on the fluorescent film is calculated according to the Fresnel formula by using the first incident angle, the included angle between the top surface of the prism and the incident surface, the refractive index of laser in the fluorescent film and the refractive index of laser in the prism, and the incident laser polarization component of the surface of the fluorescent film; The third transmittance of the laser at the exit surface of the prism is calculated according to the Fresnel formula by using the exit angle of the laser at the exit surface of the prism, the refractive index of laser in air and the refractive index of laser in the prism, and the incident laser polarization component of the exit surface; The laser power incident in the fluorescent film is determined in the following manner: wherein, represents the laser power; represents the first transmittance; represents the second transmittance; represents the third transmittance; represents the determined monitoring power; represents the incident power corresponding to the determined monitoring power.

2. The prism laser induced fluorescence detection system of claim 1, wherein, The second incident angle is determined in the following manner: wherein denotes the second angle of incidence; denotes the angle between the incidence plane and the top surface; denotes the refractive index of laser light in air; denotes the refractive index of laser light in the prism.

3. The prism laser induced fluorescence detection system of claim 1, wherein, When the fluorescent film adheres to the transparent film of the multi-well plate, the second incident angle is determined in the following manner: wherein denotes the second angle of incidence; denotes the angle between the top surface and the incidence plane; denotes the refractive index of laser light in air; denotes the refractive index of laser light in the prism; denotes the refractive index of laser light in the transparent film, and .

4. The prism laser induced fluorescence detection system of claim 1, wherein, The system further comprises an objective lens, a lens and a filter; The objective lens is located above the thin film window and is used to collect the fluorescent signal, which is focused by the lens and then transmitted to the camera; The filter is arranged between the lens and the camera.

5. The prism laser induced fluorescence detection system of claim 1, wherein, The included angle between the incident surface and the top surface is an acute angle.

6. The prism laser induced fluorescence detection system of claim 5, wherein, The first incident angle is the sum of the included angle between the incident surface and the top surface and the refraction angle of the laser entering the prism.

7. The prism laser induced fluorescence detection system of claim 1, wherein, The system further comprises a protective frame and a radiation structure; The fluorescent film is installed in the middle of the protective frame, and the height of the protective frame is higher than that of the fluorescent film. The radiation structure is installed outside the protective frame.

8. The prism laser induced fluorescence detection system of claim 1, wherein, The system further comprises a prism support and a plating layer. The prism is installed on the prism support, and the plating layer is connected with the prism support and the system ground.

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