A biological detection and optical imaging method based on a one-dimensional nanophotonic platform

Abstract

The present invention relates to the field of optical nano-imaging bioassays and discloses a bioassay and optical imaging method based on a one-dimensional nanophotonic platform. The method comprises the following steps: (1) a beam of white light is irradiated along a non-periodic direction on a one-dimensional nanostructure, and the generated scattering and diffraction signals are received using a microscope or mobile phone; (2) defects in the one-dimensional nanostructure disrupt the distribution of the local field, reduce the scattering intensity of the nanostructure, and change the color of the one-dimensional nanostructure; (3) in the visible light region, in addition to a scattering peak, another new diffraction peak can improve the sensitivity of the spectral response, allowing the measurement of size differences of 20 nanometers; (4) by modifying antibodies on the surface of the one-dimensional nanostructure, the detection of a single virus can be achieved, which is beneficial for the early diagnosis of in vitro diseases, especially in underdeveloped areas and areas with harsh biological experimental conditions. The technology is simple to operate, low-cost, and conducive to the rapid and high-throughput detection of nanoscale samples.

Description

Technical Field

[0001] The present invention relates to the technical field of optical nano-imaging bioassays, and in particular to a bioassay and optical imaging method based on a one-dimensional nano-photonic platform. Background Art

[0002] Nanoscale detection holds significant application value in areas such as colloidal self-assembly, environmental monitoring, and biomedical sensing and diagnostics. There is an urgent need to develop an efficient, low-cost biodetection platform to prevent the spread of viruses. Optical microscopy, as a nondestructive testing tool, has been widely used for large-scale, real-time characterization of the shape, size, and composition of samples. However, the imaging resolution of conventional optical microscopy is limited by the diffraction limit, making it difficult to resolve subwavelength details in the visible region. To overcome this limitation, fluorescence microscopy has been widely used for super-resolution imaging of single nanoparticles. While it facilitates observation of living biological samples, it requires complex fluorescent labeling and its detection efficiency is limited by photobleaching. Non-fluorescence characterization methods, including scattering-based dark-field microscopy and interferometric scattering microscopy, can rapidly detect and track isolated nanoparticles. The latter offers higher sensitivity for weakly scattering particles, but requires an additional coherent laser source to illuminate the sample and specialized data analysis to reconstruct the image. Furthermore, both techniques lack the specificity to distinguish nanoscale objects in multicomponent systems, posing considerable challenges in practical applications. Nanophotonic structures are easy to surface modify and can be prepared in large quantities. They have attracted widespread attention in the field of label-free and highly sensitive detection of nano-objects. The strong resonance effect of the subwavelength structure of metal / dielectric is usually used to amplify weak optical signals, thereby improving the sensitivity of spectrometer detection. For non-resonant enhanced detection, nanoscale objects can be directly observed near a semiconductor nanocavity or a pair of nanowires. Both methods require the use of a series of laser light sources in the optical system to adjust and optimize the performance of the photonic chip, which is a rather complex, expensive and time-consuming process. In addition, when the optical signals of the nanoparticles are very close, they have difficulty in identifying individual nanoparticles among multiple nanoparticles, especially viruses. Summary of the Invention

[0003] In order to improve the above technical problems, the present invention provides a detection chip comprising a one-dimensional nanostructure with biological probe molecules.

[0004] According to an embodiment of the present invention, the biological probe molecule can be selected from one, two or more of the following antibodies including but not limited to: IgM, IgG, Sig antibodies.

[0005] According to an embodiment of the present invention, the antibody is a fluorescently labeled antibody or a non-fluorescently labeled antibody.

[0006] Exemplarily, the biological probe molecule is a rabbit anti-goat Cy3 fluorescent antibody or a new coronavirus pseudovirus antibody.

[0007] According to an embodiment of the present invention, the biomolecule and the one-dimensional nanostructure may be connected by chemical bonding.

[0008] According to an embodiment of the present invention, the detection chip is capable of specifically identifying a target molecule corresponding to a biological probe molecule. Preferably, the target molecule may be a nanoscale biological particle, such as a nanoparticle, a viral particle, or a protein macromolecule. Preferably, the size of the nanoscale biological particle is no less than 50 nm, for example, 50-100 nm, such as 60 nm, 70 nm, 80 nm, or 90 nm.

[0009] According to an embodiment of the present invention, the one-dimensional nanostructure comprises nanoparticles, one-dimensional nanowires, one-dimensional nanorods, or one-dimensional nanocylinders. Preferably, the nanoparticles are of uniform size. Preferably, the one-dimensional nanostructure is self-assembled from nanoparticles, one-dimensional nanowires, one-dimensional nanorods, or one-dimensional nanocylinders of uniform size.

[0010] According to an embodiment of the present invention, the nanoparticles may be spherical, cubic or rod-shaped nanoparticles.

[0011] According to an embodiment of the present invention, the diameter of the nanoparticles is 100-600 nm, such as 150-500 nm, exemplified by 180 nm, 280 nm, 310 nm, and 360 nm.

[0012] According to an embodiment of the present invention, the material of the one-dimensional nanostructure can be a metal nanomaterial, a dielectric nanomaterial, or a polymer nanomaterial. For example, the metal nanomaterial can be selected from gold nanoparticles and / or silver nanoparticles; for example, the dielectric nanomaterial can be selected from one, two, or more of silicon nanoparticles, selenium nanoparticles, and silicon dioxide nanoparticles; for example, the polymer nanomaterial can be selected from polystyrene nanoparticles and / or polymethyl methacrylate nanoparticles. Exemplarily, the material of the one-dimensional nanostructure is a modified or unmodified polystyrene nanomaterial.

[0013] According to an exemplary embodiment of the present invention, the one-dimensional nanostructure may be a one-dimensional chain structure; preferably, it is a one-dimensional chain structure formed by self-assembly of modified or unmodified polystyrene nanoparticles.

[0014] According to an embodiment of the present invention, the one-dimensional nanostructure can generate two scattering peaks in the visible light region.

[0015] According to an embodiment of the present invention, the detection chip further comprises a substrate, and the one-dimensional nanostructure with the biological probe molecules is located on the surface of the substrate.

[0016] According to an embodiment of the present invention, the substrate may be selected from a silicon substrate, a glass substrate or a flexible polymer substrate.

[0017] According to an exemplary embodiment of the present invention, the detection chip comprises a substrate and a one-dimensional nanostructure with biological probe molecules located on the surface of the substrate;

[0018] The one-dimensional nanostructure is a one-dimensional chain structure formed by self-assembly of modified or unmodified polystyrene nanoparticles, and the biological probe molecule is a rabbit anti-goat Cy3 fluorescent antibody or a new coronavirus pseudovirus antibody.

[0019] The present invention also provides a method for preparing the above-mentioned detection chip, comprising the following steps: immersing a substrate containing a one-dimensional nanostructure modified with a carboxyl group in a mixed solution of NHS (N-hydroxysulfosuccinimide) and EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) to activate the carboxyl group; and then immersing the substrate in a solution containing a biological detection molecule for incubation to obtain the detection chip.

[0020] According to an embodiment of the present invention, the one-dimensional nanostructure, substrate and biological detection molecule all have the meanings as described above.

[0021] According to an embodiment of the present invention, the substrate containing the carboxyl-modified one-dimensional nanostructure can be obtained by self-assembly by dropping a suspension of carboxyl-modified nanoparticles, nanowires, nanorods or nanocylinders on the substrate.

[0022] According to an embodiment of the present invention, in the mixed solution of NHS and EDC, the molar concentration ratio of NHS to EDC is 1:(2-5), for example, 1:4.

[0023] According to an embodiment of the present invention, the time for activating the carboxyl group is 0.5-3 hours, for example, 1 hour.

[0024] According to an embodiment of the present invention, after the carboxyl groups are activated, the immersed substrate is cleaned, for example, by using PBS buffer to clean the substrate surface.

[0025] According to an embodiment of the present invention, the incubation time may be 3-10 hours, for example, 6 hours.

[0026] According to an embodiment of the present invention, the preparation method further comprises washing the substrate after incubation, for example, using PBS buffer to wash the substrate surface.

[0027] According to an exemplary embodiment of the present invention, the preparation method of the detection chip includes the following steps: immersing a substrate containing a one-dimensional chain structure formed by self-assembly of carboxyl-modified polystyrene nanoparticles in a mixed solution of NHS and EDC to activate the carboxyl groups; and then immersing the substrate in serum containing rabbit anti-goat Cy3 fluorescent antibody and incubating to obtain the detection chip.

[0028] According to an exemplary embodiment of the present invention, the method for preparing the detection chip includes the following steps: immersing a substrate containing a one-dimensional chain structure formed by self-assembly of carboxyl-modified polystyrene nanoparticles in a mixed solution of NHS and EDC to activate the carboxyl groups; and then immersing the substrate in serum containing antibodies to the new coronavirus pseudovirus and incubating to obtain the detection chip.

[0029] The present invention also provides application of the detection chip in detecting target molecules corresponding to the biological probe molecules.

[0030] According to an embodiment of the present invention, the target molecules and bioprobe molecules have the meanings as shown above.

[0031] According to an embodiment of the present invention, the detection chip can detect a single target molecule corresponding to the biological probe molecule.

[0032] The present invention also provides a biological detection and optical imaging method, comprising the following steps:

[0033] (1) immersing the detection chip in a system containing target molecules to obtain a detection chip that adsorbs the target molecules;

[0034] (2) using oblique incident light to illuminate the one-dimensional nanostructure in the detection chip that adsorbs the target molecule, and collecting an optical image;

[0035] (3) Analyzing the optical image to obtain an intensity distribution curve of the one-dimensional nanostructure, and obtaining the type, position, number and / or size of the adsorbed target molecules based on the intensity change.

[0036] According to an embodiment of the present invention, the detection chip has the meaning as described above.

[0037] According to an embodiment of the present invention, the system containing the target molecule may be serum containing the target molecule.

[0038] According to an embodiment of the present invention, the target molecule is adsorbed on the detection chip through specific recognition between the target molecule and the biological probe molecule contained in the detection chip. For example, the target molecule can be adsorbed on the top of the one-dimensional nanostructure, on the side of the one-dimensional nanostructure, and / or between the gaps.

[0039] According to an embodiment of the present invention, the inclined incident light means that the angle of the incident light is greater than or equal to 70 degrees.

[0040] According to an embodiment of the present invention, the wavelength of the incident light is in the range of 350 to 1100 nm.

[0041] According to an embodiment of the present invention, the incident light is white light. For example, the white light can be generated by a halogen lamp, a xenon lamp or an LED lamp.

[0042] According to an embodiment of the present invention, the incident light in step (2) is incident along a non-periodic direction of the one-dimensional nanostructure.

[0043] According to an embodiment of the present invention, step (2) includes using a mobile phone or an optical microscope to receive scattering and diffraction signals generated by the one-dimensional nanostructure in the detection chip that adsorbs the target molecules, and collecting an optical image.

[0044] According to one embodiment of the present invention, step (2) comprises: placing the detection chip having adsorbed target molecules on a stage of an optical microscope, irradiating the one-dimensional nanostructure in the detection chip having adsorbed target molecules along a non-periodic direction using inclined incident light, and collecting an optical image using a camera;

[0045] According to another embodiment of the present invention, in step (2), a mobile phone can be used to receive the scattering and diffraction signals generated by the one-dimensional nanostructure, and the target molecules adsorbed on the one-dimensional nanostructure can be observed using the mobile phone. Specifically, a microscope camera with an image magnification function is installed on the mobile phone, and the mobile phone camera is turned on to directly photograph the sample to obtain an optical image.

[0046] Preferably, the optical microscope is provided with an objective lens capable of receiving light and a camera capable of forming an image.

[0047] According to an embodiment of the present invention, the intensity distribution curve refers to the intensity value extracted from the grayscale value of each pixel point obtained by image processing software.

[0048] According to an embodiment of the present invention, the nano-defects in the one-dimensional nanostructure may disrupt the distribution of the local field, reduce the scattering intensity of the nanostructure, and change the color of the one-dimensional nanostructure.

[0049] Specifically, the size of the nano-defect is greater than or equal to 20 nm. The nano-defect can be caused by the unevenness of the structure itself, such as misalignment caused by uneven arrangement of nanoparticles, or it can be caused by the introduction of other substances from the outside, such as additional nanoparticles, viruses and / or protein molecules.

[0050] For example, when there is a defect in the one-dimensional nanostructure, the intensity of scattered light at the defect is smaller than the intensity of scattered light at a non-defective position.

[0051] For example, when defects are introduced into the one-dimensional nanostructure, its scattering spectrum will be changed, including changes in intensity and shifts in wavelength.

[0052] Preferably, the diameter of the nanoparticles constituting the one-dimensional nanostructure can be directly observed when it changes by at least 20 nm.

[0053] Preferably, the biological detection and optical imaging method comprises the following steps:

[0054] (a) A beam of white light is irradiated along a non-periodic direction onto a one-dimensional nanostructure within a detection chip that has adsorbed target molecules. The resulting scattering and diffraction signals are then received using an optical microscope or mobile phone to acquire an optical image.

[0055] (b) Defects in the one-dimensional nanostructure disrupt the distribution of the local field, reduce the scattering intensity of the nanostructure, and change the color of the one-dimensional nanostructure; in the visible light region, in addition to a scattering peak, another new diffraction peak improves the sensitivity of the spectral response, allowing the measurement of size differences of at least 20 nanometers; by analyzing the optical image, an intensity distribution curve of the one-dimensional nanostructure is obtained, and the type, position, number and / or size of the adsorbed target molecules are obtained based on the intensity change.

[0056] The present invention also provides a biological detection system, which includes the biological detection chip.

[0057] According to an embodiment of the present invention, the system further comprises an optical device and an imaging device, wherein the optical device is used to provide incident light, and the imaging device is used to collect optical images.

[0058] According to an embodiment of the present invention, the optical device and the imaging device may be provided simultaneously by an optical microscope.

[0059] According to an embodiment of the present invention, the imaging device may be a mobile phone with a microscope camera.

[0060] Beneficial effects of the present invention

[0061] The present invention provides a detection chip based on a one-dimensional nanostructure, a method for preparing the same, and a biological detection and imaging method and system utilizing the detection chip. The present invention utilizes a one-dimensional nanophotonic platform to achieve label-free, visualized detection of biological nanoparticles. During detection, a beam of white light is irradiated obliquely along a non-periodic direction onto the one-dimensional nanostructure of the detection chip, which specifically identifies and adsorbs target molecules. By adjusting the angle and direction of the incident light, defects in the one-dimensional nanostructure can be directly detected using a conventional optical microscope or mobile phone. These defects are caused by the one-dimensional nanostructure itself and by the adsorption of target molecules on the one-dimensional nanostructure. The type, location, number (even a single target molecule) and / or size of the target molecule can be identified by the scattering spectrum and color changes of the one-dimensional nanostructure.

[0062] Compared with traditional methods, the method of the present invention does not require fluorescent labeling and is non-destructive to biological samples, making detection simpler and faster. It is of great significance in immunology, physiology, and in vitro disease diagnosis, and is particularly suitable for underdeveloped areas and areas with harsh biological experimental conditions.

[0063] The method of the present invention is also simple to operate, low in cost, and portable, and can realize the detection of a single nanometer-scale object using a microscope or a mobile phone. BRIEF DESCRIPTION OF THE DRAWINGS

[0064] Figure 1 Schematic diagram of the principle of optical nanodetection of a single virus in Example 1 of the present invention (a); an optical photograph of a virus adsorbed on the surface of a one-dimensional nanostructure (left image in b) and a scanning electron microscope photograph (middle image in b); and a scattering intensity distribution curve of the one-dimensional nanostructure after virus adsorption (right image in b);

[0065] The scale of the optical photo is 1 micron, and the scale of the scanning electron microscope photo is 500 nanometers.

[0066] Figure 2 1 is a statistical diagram of the size distribution of different viruses in Example 1 of the present invention.

[0067] Figure 3 These are fluorescent photos before and after the antibody is modified on the surface of the one-dimensional nanostructure in Example 2 of the present invention.

[0068] Figure 4 These are optical photographs, electron microscope photographs, and intensity distribution curves of viruses adsorbed at different positions on the one-dimensional nanostructure in Example 3 of the present invention (the intensity distribution curves are from top to bottom: red channel, blue channel, green channel);

[0069] The scale of the optical photo is 1 micron, and the scale of the scanning electron microscope photo is 500 nanometers.

[0070] Figure 5 This is an electron microscope photograph of adenovirus and novel coronavirus pseudovirus adsorbed on the surface of a one-dimensional nanostructure in Example 4 of the present invention.

[0071] Figure 6 This is a photo of the one-dimensional nanostructure taken with a mobile phone in Example 5 of the present invention (by adjusting the camera magnification, optical signals of multiple viruses adsorbed on the one-dimensional nanostructure can be observed). DETAILED DESCRIPTION

[0072] The technical solutions of the present invention will be described in further detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanations of the present invention and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are encompassed within the scope of protection that the present invention is intended to protect.

[0073] Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0074] Example 1

[0075] like Figure 1 As shown, a suspension of carboxyl-modified polystyrene nanoparticles was dropped onto a clean, smooth silicon substrate. A one-dimensional chain-like nanostructure with a diameter of 310 nm and a refractive index of 1.59 was obtained through solution self-assembly. The resulting one-dimensional nanostructure was then immersed in a mixed solution of 0.05 mol N-hydroxysulfosuccinimide sodium salt (NHS) and 0.2 mol 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) to activate the surface carboxyl groups. After soaking for one hour, the residual solution on the surface was washed off with PBS buffer. The activated one-dimensional nanostructure was then incubated in a solution containing an antibody for six hours. The residual antibody on the substrate was then washed off with PBS buffer, resulting in a detection chip. The one-dimensional nanostructure chip modified with antibodies was placed in a human serum solution containing the corresponding virus (antigen) for incubation. After one hour of antibody-antigen specific recognition, unrecognized virus particles were washed with PBS.

[0076] An optical microscope was used to observe the virus-adsorbed chip. A xenon lamp was used as the white light source, with a wavelength of 400-760nm, an incident angle of 70°, and incident along a non-periodic direction. The objective lens had a magnification of 20, a working distance of 11mm, and a numerical aperture of 0.4. The camera imaged the light received by the objective lens with a resolution of 16 million pixels. This method can also be used to observe the adsorption of viruses on the surface of the structure using the camera built into the mobile phone. The collected photos were decomposed into three channels: red, green, and blue using the image processing software Image J, and the intensity change information of each single channel was extracted. When viruses are adsorbed on the surface of a one-dimensional nanostructure, the adsorbed viruses will change the color of the one-dimensional nanostructure and reduce the scattering intensity of the one-dimensional nanostructure. Therefore, one-dimensional nanostructures can be directly used to detect viruses of different sizes and numbers without labeling.

[0077] like Figure 2 As shown in the figure, the size distribution histogram shows that the average size of the four viruses is smaller than the optical diffraction limit. The specific operation steps are as follows: first, the four test solutions containing the new coronavirus pseudovirus (SARS-CoV-2), adenovirus (Adenovirus), lentivirus (Lentivirus), and herpes virus (EB virus) are diluted with PBS buffer to a concentration of 1x10 3 One microliter was dropped onto a copper grid, cooled with liquid nitrogen, and the size of the virus was observed using a scanning electron microscope.

[0078] Example 2

[0079] According to Example 1, a suspension of polystyrene nanoparticles modified with carboxyl groups was dropped onto a clean and smooth silicon substrate, and a one-dimensional chain structure was obtained by solution self-assembly. The diameter of the nanoparticles was 310 nm, and they were periodically and tightly arranged. The obtained one-dimensional structure was immersed in a mixed solution of 0.05 mol NHS and 0.2 mol EDC to activate the carboxyl groups on the surface of the nanoparticles. The one-dimensional structure after the carboxyl groups were activated was immersed in a fluorescently labeled rabbit anti-goat Cy3 fluorescent antibody solution, incubated at 4 ° C for 6 hours, and washed three times with PBS buffer solution. After activation, the carboxyl groups can more efficiently react with the amino groups in the antibody protein to form a chemical bond, so that the antibodies are evenly modified on the surface of the one-dimensional structure to obtain a detection chip. The chip with modified antibodies was placed on the stage of a confocal microscope, with a mercury lamp as the incident light source and an incident angle of 70 degrees.

[0080] like Figure 3As shown, the one-dimensional nanostructures before antibody modification exhibited no fluorescence signal under a confocal microscope. However, after modification with the fluorescent antibody, the one-dimensional nanostructures exhibited a yellow-green fluorescence signal under the confocal microscope, indicating that the antibodies were evenly distributed on the surface of the one-dimensional structures with uniform fluorescence intensity. Comparing scanning electron micrographs of the one-dimensional nanostructures before and after antibody modification revealed that the antibodies had little effect on the morphology of the one-dimensional nanostructures.

[0081] Example 3

[0082] According to the method in Example 1, an aqueous solution of polystyrene nanoparticles modified with carboxyl groups was dropped onto a clean and smooth silicon substrate, and a one-dimensional chain structure was obtained by solution self-assembly. The diameter of the nanoparticles was 310 nm, and they were periodically and closely arranged. The obtained one-dimensional structure was placed in a mixed solution of NHS and EDC in Example 1 to activate the carboxyl groups on the surface of the nanoparticles. The one-dimensional structure after the activated carboxyl group was placed in a new crown pseudovirus antibody (SARS-CoV-2 (2019-nCoV) Spike Antibody) solution and incubated at 4°C for 6 hours. After activation, the carboxyl group can more efficiently react with the amino group in the antibody protein to chemically bond, so that the antibody is evenly modified on the surface of the one-dimensional structure to obtain a detection chip. The structure chip after the modified antibody was cleaned with PBS buffer solution and placed in a new crown pseudovirus (SARS-CoV-2 (2019-nCoV) Spike Pseudovirus) solution and incubated at 37°C for 1 hour. After the incubation is complete, the one-dimensional nanostructure chip is rinsed three times with PBS buffer solution, blown dry, and placed on a microscope stage to characterize the structure after antibody virus (antigen) specific adsorption. The light source for optical detection is a xenon lamp with an incident angle of 70 degrees.

[0083] according to Figure 4 Scanning electron microscopy images show that virus adsorption on the sides, tops, and crevices of one-dimensional nanostructures produces distinct optical signals. Using the image processing software Image J, the optical images of adsorbed viruses are divided into three channels: red, green, and blue. It can be seen that the image intensity decreases at locations where the virus is adsorbed.

[0084] Example 4

[0085] According to the method in Example 1, an aqueous solution of polystyrene nanoparticles modified with carboxyl groups was dropped onto a clean and smooth silicon substrate, and a one-dimensional chain structure was obtained by solution self-assembly. The diameter of the nanoparticles was 310 nm and they were periodically and closely arranged. The obtained one-dimensional structure was placed in a mixed solution of NHS and EDC in Example 1 to activate the carboxyl groups on the surface of the nanoparticles. The one-dimensional structure after the carboxyl group activation was placed in a new crown pseudovirus antibody (SARS-CoV-2 (2019-nCoV) Spike Antibody) solution and incubated at 4°C for 6 hours. After activation, the carboxyl group can more efficiently undergo a chemical bonding reaction with the amino group in the antibody protein, so that the antibody is evenly modified on the surface of the one-dimensional structure to obtain a detection chip. After the antibody-modified structure chip was cleaned with a buffer solution, it was placed in a new crown pseudovirus (SARS-CoV-2 (2019-nCoV) Spike Pseudovirus) and an adenovirus solution, and the virus was incubated at 37°C for 1 hour. After the chip was incubated with the virus, it was rinsed with a buffer solution and placed on a microscope stage to characterize the optical signal of the structure after the antibody virus adsorption. The light source used was a xenon lamp with an incident angle of 70 degrees.

[0086] Figure 5 The results showed that the antibodies had a significant specific adsorption effect on the corresponding viruses, with more viruses being observed on the surface of the structure. However, the antibodies had no specific adsorption effect on non-corresponding viruses, with almost no viruses being observed on the surface of the structure.

[0087] Example 5

[0088] According to the method in Example 1, an aqueous solution of polystyrene nanoparticles with carboxyl modifications and particle sizes of 180nm, 280nm, 310nm, and 360nm were dropped onto a clean and smooth silicon substrate, and a large-area one-dimensional structure was obtained by solution self-assembly. According to the method in Example 1, the one-dimensional structure after the carboxyl group is activated is placed in a new coronavirus pseudovirus antibody (SARS-CoV-2 (2019-nCoV) Spike Antibody) solution and incubated at 4°C for 6 hours. After activation, the carboxyl group can more efficiently undergo chemical bonding reaction with the amino group in the antibody protein, so that the antibody is evenly modified on the surface of the one-dimensional structure to obtain a detection chip. After the antibody-modified structure chip is cleaned with a buffer solution, it is placed in the new coronavirus pseudovirus (SARS-CoV-2 (2019-nCoV) SpikePseudovirus) and incubated at 37°C for 1 hour. The unidentified viruses on the chip are rinsed with PBS buffer solution. A microscope camera is installed on the mobile phone, and the one-dimensional nanostructure can be directly observed by adjusting the magnification of the microscope camera. As Figure 6As shown in the image, at low magnification (15x), the color of the one-dimensional structure changes with size: a one-dimensional structure with a diameter of 180nm appears blue-green, a one-dimensional structure with a diameter of 280nm appears orange-red, a one-dimensional structure with a diameter of 310nm appears pink, and a one-dimensional structure with a diameter of 360nm appears purple. At high magnification (200x), the differences in optical signals generated by multiple viruses adsorbed on the surface of the one-dimensional nanostructure can be directly seen. By performing differential processing on optical images of the one-dimensional nanostructure before and after virus adsorption, the optical signals of the viruses can be directly obtained. The scale bar of the scanning electron microscopy image is 500.

[0089] The above describes the embodiments of the present invention. However, the present invention is not limited to the above embodiments. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included in the scope of protection of the present invention.

Claims

1. A detection chip, characterized in that: The detection chip comprises a substrate and a one-dimensional nanostructure with biological probe molecules located on the surface of the substrate; the one-dimensional nanostructure is a one-dimensional chain structure formed by self-assembly of modified or unmodified polystyrene nanoparticles; the biological probe molecules are connected to the one-dimensional nanostructure by chemical bonding; The biological probe molecule is selected from one, two or more of the following antibodies, including but not limited to: IgM, IgG, Sig antibodies; The detection chip can specifically identify target molecules corresponding to biological probe molecules.

2. The detection chip according to claim 1, characterized in that The antibody is a fluorescently labeled antibody or a non-fluorescently labeled antibody; The target molecule is a nanoscale biological particle selected from a virus particle or a protein macromolecule; the size of the nanoscale biological particle is not less than 50 nm; The substrate is selected from a silicon substrate, a glass substrate or a flexible polymer substrate; The one-dimensional nanostructure can generate two scattering peaks in the visible light region.

3. The detection chip according to claim 1, characterized in that: The biological probe molecule is rabbit anti-goat Cy3 fluorescent antibody or new coronavirus pseudovirus antibody.

4. The method for preparing the detection chip according to any one of claims 1 to 3, characterized in that: The preparation method comprises the following steps: immersing a substrate containing a one-dimensional chain structure formed by self-assembly of carboxyl-modified polystyrene nanoparticles in a mixed solution of N-hydroxysulfosuccinimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride to activate the carboxyl groups; and then immersing the substrate in a solution containing biological probe molecules for incubation to obtain the detection chip.

5. The preparation method according to claim 4, characterized in that The preparation method of the detection chip comprises the following steps: immersing a substrate containing a one-dimensional chain structure formed by self-assembly of carboxyl-modified polystyrene nanoparticles in a mixed solution of NHS and EDC to activate the carboxyl groups; then immersing the substrate in serum containing rabbit anti-goat Cy3 fluorescent antibody and incubating the solution to obtain the detection chip; Alternatively, the preparation method of the detection chip includes the following steps: immersing a substrate containing a one-dimensional chain structure formed by self-assembly of carboxyl-modified polystyrene nanoparticles in a mixed solution of NHS and EDC to activate the carboxyl groups; and then immersing the substrate in serum containing new coronavirus pseudovirus antibodies for incubation to obtain the detection chip.

6. Use of the detection chip according to any one of claims 1 to 3 in detecting target molecules corresponding to biological probe molecules.

7. The use according to claim 6, characterized in that The detection chip is used to detect a single target molecule corresponding to a biological probe molecule.

8. A biological detection and optical imaging method, characterized in that: The method comprises the following steps: (1) Immersing the detection chip according to any one of claims 1 to 3 in a system containing a target molecule to obtain a detection chip that adsorbs the target molecule; (2) using oblique incident light to illuminate the one-dimensional nanostructure in the detection chip that adsorbs the target molecule, and collecting an optical image; (3) Analyzing the optical image to obtain an intensity distribution curve of the one-dimensional nanostructure, and obtaining the type, position, number and / or size of the adsorbed target molecules based on the intensity change.

9. The method according to claim 8, characterized in that The system containing the target molecule is serum containing the target molecule; The adsorption of target molecules on the detection chip is achieved through specific recognition between the target molecules and the biological probe molecules contained in the detection chip, resulting in adsorption on the one-dimensional nanostructure surface; The inclined incident light refers to the incident light having an angle greater than or equal to 70 degrees; The wavelength range of the incident light is 350-1100 nm; The incident light is white light.

10. The method according to claim 9, characterized in that The target molecules are adsorbed on the top of the one-dimensional nanostructure, on the side of the one-dimensional nanostructure and / or between the gaps.

11. The method according to claim 8, characterized in that The incident light in step (2) is incident along the non-periodic direction of the one-dimensional nanostructure; Step (2) includes using a mobile phone or an optical microscope to receive scattering and diffraction signals generated by the one-dimensional nanostructure in the detection chip that adsorbs the target molecule, and collecting an optical image; Nano defects in the one-dimensional nanostructure can disrupt the distribution of the local field, reduce the scattering intensity of the nanostructure, and change the color of the one-dimensional nanostructure; The size of the nano-defect is greater than or equal to 20 nm.

12. A biological detection system, characterized in that: The system comprises the detection chip according to any one of claims 1 to 3.

13. The biological detection system according to claim 12, characterized in that: The system further includes an optical device and an imaging device, wherein the optical device is used to provide incident light, and the imaging device is used to collect optical images.

Images