CN114910642A - Method for rapidly detecting characteristics of microscopic substances or structures - Google Patents
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Abstract
The invention relates to a method for rapidly detecting the characteristics of microscopic substances or structures, which comprises the following steps: (1) reacting and connecting the microscopic substance to be detected and analyzed with the magnetic microspheres to form a magnetic microsphere combination, and dispersing the magnetic microsphere combination in a sample solution to be analyzed; (2) and simultaneously, strictly limiting and conforming the spatial orientation of the anisotropic magnetic microsphere combination in the sample liquid by applying fluid limitation and magnetic field control, singly and sequentially flowing through a detection analysis position along a specific track, and measuring and counting the characteristics of the microsphere combination one by one in a uniform spatial orientation to finish the method. Compared with the prior art, the method can rapidly analyze the characteristics of a large number of single microscopic substances or structures in a short time, and further obtain effective information through analysis.
Description
Technical Field
The invention belongs to the technical field of microscopic substance detection and analysis, and relates to a method for rapidly detecting characteristics of microscopic substances or structures.
Background
In scientific research and engineering technology, with the progress of science and technology, a more intensive understanding and more precise application are required to analyze and study the composition and interaction of microscopic substances such as various molecules, proteins, nucleic acids, cell bodies, organic macromolecules and the like. Direct imaging methods such as electron microscopy are not only expensive, but also have low analysis efficiency and long detection time, and are not conducive to rapid analysis of a large number of samples. On the other hand, with the help of special microspheres, such as nanometer or micrometer scale, microscopic substances such as molecules to be analyzed are connected with the microspheres to form a special structure or microsphere combination, and by means of optical means and the like, characteristic signals of the microsphere combination can be simply and rapidly detected and analyzed, so that characteristics of the microscopic substances, such as composition and interaction, reaction kinetics, classification, concentration determination and the like, can be further analyzed.
On one hand, the single microsphere combination body dispersed in the sample liquid flows through the analysis light beam in a specific track in a very small limited space to measure the characteristic optical signal, so that the characteristics of a large number of single microsphere combination bodies can be quickly analyzed in a short time, and effective information can be further obtained through analysis.
For example, capillary techniques may be used to confine the fluid, allowing the individual microsphere arrays to flow through the analysis beam in a specific trajectory. The capillary has an inner diameter very fine, e.g. 10 microns, and the sample flow containing the particles to be analyzed is a single, non-stratified fluid. Flow Cytometry techniques may also be used for Flow confinement by allowing individual microsphere conjugates to Flow through the analytical beam in a specific trajectory using layered Flow, using sheath fluid confinement and modulation of sample Flow.
Specifically, the flow analysis technique is a powerful advanced technique for rapidly analyzing single particles, and the object to be detected by the flow analyzer is cells, particles or microsphere combinations suspended in a liquid. In the flow analysis technique, as shown in fig. 1, using hydrodynamic focusing principle (hydrodynamic focusing), in an optical flow cell 6, for example, a quartz glass material with 200 × 200 μm square inner hole, a sheath fluid 2(sheath) wraps a sample fluid 3(sample fluid) and flows forward, and the sheath fluid 2 and the sample fluid 3 are layered and do not mix; the sheath liquid limits the sample liquid to make the diameter of the sample liquid flow small, such as several micrometers, so that the analyzed particles, cells or microsphere combinations are arranged in a line and pass through the analysis light beams one by one; meanwhile, the diameter of the sample liquid flow can be adjusted by adjusting the pressure and flow rate of the sheath liquid and the sample liquid, respectively.
As shown in fig. 1, in the optical flow cell 6, the sheath liquid 2 lines up the microparticles, cells, or microsphere combinations to be analyzed in the sample liquid 3, and rapidly flows through the analyzing light beam 1 such as a detecting laser beam one by one. A high-precision optical system collects and detects characteristic optical signals, such as multi-angle scattered light scattered by particles and fluorescence of multiple wavelengths excited; through complex electronic signal processing and computer data analysis, information such as a plurality of physical structural characteristics and biological expression characteristics of tens of thousands of cells or particles can be obtained in a short time. The flow cytometer may be equipped with a multi-color laser, and multiple scattered light and fluorescence detectors to achieve powerful cell and particle analysis functions. In the characteristic optical signal detection, the scattered light of different angles reflects the physical structure characteristics of the particles, cells or microsphere combination, for example, the forward scattered light and the side scattered light can reflect the size, the internal structure and other characteristics of the particles or microsphere combination. Fluorescence detection at different wavelengths can reflect the characteristics of molecular expression. For example, in order to detect the expression of a specific protein molecule, a proper fluorescent marker is used for marking, for example, PE fluorescent dye is used for detecting emitted fluorescence at the wavelength of 585nm under the excitation of 532nm laser, so that quantitative information of the expression of the specific protein molecule can be obtained, and important basis is provided for scientific research and clinical diagnosis.
However, in the capillary or flow analysis techniques described above, the sample fluid stream is typically on the order of microns, e.g., 5-20 microns in diameter, due to hydrodynamic properties, and the size of the microsphere combination being analyzed is approximately between 50-500 nm. Even if the sample stream is as small as possible, the sample stream is much larger in diameter than the nanoscale microsphere combination being analyzed. Therefore, the spatial orientation of the microsphere-bound product being analyzed in the sample fluid stream is random, as shown in FIG. 1, and thus the spatial and optical characteristics of the microsphere-bound product cannot be accurately analyzed by optical means.
Therefore, how to realize the rapid and effective detection of the characteristics of the microscopic substances or structures is very valuable.
Disclosure of Invention
The invention aims to provide a method for rapidly detecting the characteristics of microscopic substances or structures. In order to analyze and study the composition and interaction of microscopic substances, such as various molecules, proteins, nucleic acids, cell bodies, organic polymers, etc., the microscopic substances to be analyzed, such as molecules, can be reacted and connected with the microspheres by means of special microspheres, such as nano or micron scale, to form a special structure or microsphere combination, and the characteristic signals of the microsphere combination can be simply and rapidly detected by means of optical means, etc., so as to analyze the characteristics of the microscopic substances, such as composition and interaction, reaction kinetics, classification and concentration determination, etc. In particular, in bioassays, quantitative analysis of target antigen or antibody molecules is widely used for diagnosis of diseases.
The purpose of the invention can be realized by the following technical scheme:
a method for rapidly detecting a characteristic of a microscopic substance or structure, comprising the steps of:
(1) reacting and connecting the microscopic substances to be detected and analyzed with the magnetic microspheres to form a magnetic microsphere combination, and dispersing the magnetic microsphere combination in sample liquid to be analyzed;
(2) and simultaneously, strictly limiting and conforming the spatial orientation of the anisotropic magnetic microsphere combination in the sample liquid by applying fluid limitation and magnetic field control, singly and sequentially flowing through a detection analysis position along a specific track, and measuring and counting the characteristics of the microsphere combination one by one in a uniform spatial orientation to finish the method.
Furthermore, the microscopic substance is protein molecules, nucleic acid molecules, organic macromolecules or cell bodies which can be connected with the magnetic microspheres in a reaction way.
Further, the protein molecule is an antigen or an antibody.
Further, the magnetic field control specifically comprises: and applying an external magnetic field to enable the anisotropic magnetic microsphere combination in the sample liquid to reach a consistent spatial orientation under the action of the magnetic field.
Furthermore, the external magnetic field is generated by a permanent magnet or an electromagnet with controllable direction and strength.
Furthermore, the direction of the external magnetic field is consistent with the flow direction of the magnetic microsphere combination through the detection analysis position.
Further, the fluid limitation is specifically: the bead-bound bodies are caused to flow individually and sequentially through the detection assay site in a specific trajectory using a stratified fluid control technique, or a capillary technique using a single sample flow.
Furthermore, the layered fluid control technology is specifically as follows: the method comprises the steps of regulating the pressure and the flow velocity of sheath liquid and sample liquid by utilizing a hydrodynamic focusing principle and adopting a technology of limiting the sample liquid flow by the sheath liquid, so that the diameter of the sample liquid flow is reduced.
Furthermore, the characteristics of the microsphere combination body are analyzed by adopting an optical means, and the detection analysis position is the position where the analysis light beam and the sample flow formed by the sample liquid are intersected.
Further, the characteristic optical signal detected is scattered light, fluorescence or excited luminescence.
Compared with the prior art, the invention simultaneously applies fluid restriction and magnetic field control to enable a single microstructure (such as a magnetic microsphere combination combined with a microscopic substance) to pass through a detection analysis position (such as an analysis light beam) in a specific track and strictly consistent spatial orientation, and then measures a corresponding characteristic signal, so that a large amount of characteristics of the single microscopic substance or structure can be rapidly analyzed in a short time, and effective information can be further obtained through analysis.
Drawings
FIG. 1 is a schematic diagram of a prior art detection process;
FIG. 2 is a schematic view of the binding of a microscopic substance to magnetic microspheres;
FIG. 3 is a schematic view of the detection process of the present invention;
FIG. 4 is a schematic view of another detection process of the present invention;
FIG. 5 is a schematic view of the detection process in example 1;
FIG. 6 is a two-dimensional scattergram measured in example 1;
the notation in the figure is:
1-analyzing light beam, 2-sheath liquid, 3-sample liquid, 4-external magnetic field, 5-capillary, 6-optical flow chamber, 7-magnetic microsphere combination, 8-first photoelectric detector and 9-second photoelectric detector.
Detailed Description
The invention is described in detail below with reference to the figures and specific embodiments. The present embodiment is implemented on the premise of the technical solution of the present invention, and a detailed implementation manner and a specific operation process are given, but the scope of the present invention is not limited to the following embodiments.
The detection method of the present invention is explained in detail below.
In order to analyze and study the composition and interaction of microscopic substances, such as various molecules, proteins, nucleic acids, cell bodies, organic polymers, etc., the microscopic substances to be analyzed, such as molecules, can be reacted and connected with the microspheres by means of special microspheres, such as nano or micron scale, to form a special structure or microsphere combination, and the characteristic signals of the microsphere combination can be simply and rapidly detected by means of optical means, etc., so as to analyze the characteristics of the microscopic substances, such as composition and interaction, reaction kinetics, classification and concentration determination, etc. In particular, in bioassays, quantitative analysis of target antigen or antibody molecules is widely used for diagnosis of diseases.
Microscopic substances to be analyzed, such as molecules, are reacted with microspheres to be linked, and in the reaction solution, the microsphere conjugates have various forms, such as a Dimer structure (Dimer) formed by two microsphere conjugates, and also forms of monomeric microspheres, trimers, tetramers, etc., which are generally in the nanometer scale.
Analyzing microsphere conjugates in a sample fluid, such as dimers, monomer microspheres, trimers, tetramers, etc., to accurately distinguish the types and amounts thereof, such as total number or amount of each class, can provide a lot of useful information, such as the concentration of target antigens to be detected, etc.
In particular, if the characteristics of the dimer in the sample liquid, such as the spatial structure characteristics, the distance between two microspheres, the optical characteristic signal, etc., can be accurately analyzed, more useful information can be provided, and a powerful means is provided for researching the composition and interaction of molecules, developing a new detection instrument.
Using capillary or flow analysis techniques, as shown in fig. 1, a characteristic optical signal can be measured by flowing a single microsphere conjugate dispersed in a sample fluid through an analysis beam in a specific trajectory in a very small confined space. Thus, it is possible to rapidly analyze the characteristics of a large number of individual microsphere combinations in a short time, and further to obtain effective information through analysis.
However, due to the hydrodynamic properties, the sample fluid stream typically has a diameter on the micrometer scale, e.g., 5-20 micrometers, while the microspheres or microsphere combinations being analyzed are typically on the nanometer scale. Even if the diameter of the sample stream is minimized, the sample stream has a diameter that is much larger than the associated nanoscale microspheres, such as dimers, trimers, tetramers, etc., that are being analyzed. Therefore, the spatial orientation of the microsphere conjugates being analyzed in the sample fluid stream is random, as shown in FIG. 1, and thus the spatial and optical characteristics of anisotropic microsphere conjugates cannot be accurately analyzed by optical means, and thus the types of dimers, trimers and tetramers cannot be accurately resolved, resulting in confusion. It is impossible to precisely analyze intrinsic characteristics of the dimer in the reaction solution, such as a spatial substructure difference, a distance difference between two microspheres, and the like.
Effective information can only be obtained by analyzing when all dimer structures flow in the vertical direction through the analyzing laser beam, for example, by measuring the characteristic optical signal by sequentially passing the analyzing beam in uniform spatial orientation with the spatial orientation of the anisotropic microsphere assembly being analyzed in the sample stream strictly defined.
According to the electromagnetic principle, a specially controlled magnetic field can be used for controlling the magnetic microspheres or the combination body to complete the operations of separation, movement, rotation and the like.
The magnetic microsphere, such as a microsphere containing a superparamagnetic characteristic substance, is used for reacting and connecting the microscopic substance to be analyzed, such as molecules, with the magnetic microsphere to form a special structure or a magnetic microsphere combination, so that the magnetic microsphere or the combination can be controlled by an external magnetic field to move, rotate and the like. In particular, a dimer structure formed by connecting two magnetic microspheres flowing in a sample fluid flow is magnetized in an external magnetic field, the spatial orientation of the dimer structure of the magnetic microspheres finally coincides with the direction of the external magnetic field under the action of a magnetic field force, and analysis light beams such as focused laser beams sequentially flow one by one in a uniform spatial orientation to detect various characteristic optical signals such as scattered light at different angles, fluorescence at different wavelengths or excited luminescence, so that the types of dimers, trimers and tetramers in the sample fluid can be accurately distinguished, and the spatial substructure characteristics of the dimers in the sample fluid can be accurately analyzed.
Accordingly, the present invention provides a method for rapidly detecting characteristics of microscopic substances or structures, comprising the steps of:
(1) reacting and connecting the microscopic substances to be detected and analyzed with the magnetic microspheres to form a magnetic microsphere combination, and dispersing the magnetic microsphere combination in a sample solution to be analyzed;
(2) and simultaneously, strictly limiting and conforming the spatial orientation of the anisotropic magnetic microsphere combination in the sample liquid by applying fluid limitation and magnetic field control, singly and sequentially flowing through a detection analysis position along a specific track, and measuring and counting the characteristics of the microsphere combination one by one in a uniform spatial orientation to finish the method.
In some embodiments, the microscopic material is a protein molecule, a nucleic acid molecule (e.g., a DNA molecule, etc.), an organic polymer, or a cell body that can be linked to the magnetic microsphere by reaction. Furthermore, the protein molecule is an antigen or an antibody. More specifically, the antigenic molecule may be e.g. procalcitonin PCT or the like.
In some embodiments, the magnetic field control is in particular: and applying an external magnetic field to enable the anisotropic magnetic microsphere combination in the sample liquid to reach a consistent spatial orientation under the action of the magnetic field.
Furthermore, the external magnetic field is generated by a permanent magnet or an electromagnet with controllable direction and intensity.
Furthermore, the direction of the external magnetic field is consistent with the flow direction of the magnetic microsphere combination through the detection analysis position.
In some embodiments, the fluid restrictions are in particular: the microsphere-bound bodies are individually flowed sequentially through the detection and analysis site in a specific trajectory using a laminar fluid control technique, or a capillary technique using a single sample flow.
Furthermore, the layered fluid control technology is specifically as follows: the method utilizes the hydrodynamic focusing principle, adopts the technology of limiting the sample liquid flow by the sheath liquid, and adjusts the pressure and the flow velocity of the sheath liquid and the sample liquid so as to reduce the diameter of the sample liquid flow as much as possible.
In some embodiments, the characteristics of the microsphere conjugates are analyzed by optical means, and the detection analysis position is the position where the analysis beam intersects the sample fluid formed by the sample fluid to be analyzed.
In some embodiments, the characteristic optical signal detected is scattered light, fluorescence, or excited luminescence.
In some embodiments, the magnetic microspheres used have a diameter of 50-1000 nm, preferably, the magnetic microspheres have a diameter of 100-500 nm.
Specifically, after the microscopic substances to be analyzed are combined with the magnetic microspheres, the present invention can simultaneously apply fluid confinement and magnetic field control to strictly confine the spatial orientation of the anisotropic microscopic substances or structures to be analyzed to be consistent, and allow the individual microscopic substances or structures to sequentially flow through the analysis position by the analysis light beam or the like in a specific trajectory.
More specifically, taking optical analysis as an example, as shown in fig. 3, in the detection analysis, a sample liquid containing the magnetic microsphere bonded bodies 7 is sent to a flow optical detection system to be analyzed. The diameter of the sample liquid 3 is reduced as much as possible by respectively adjusting the pressure and the flow of the sheath liquid 2 and the sample liquid 3, the flow track of the analyzed magnetic microsphere combination 7 is limited, the sheath liquid arranges the analyzed magnetic microsphere combination 7 into a line, and the magnetic microsphere combination 7 flows through the analysis light beams one by one quickly; meanwhile, a specially controlled external magnetic field 4 is applied, the direction and the strength of the magnetic field can be precisely controlled, for example, the direction of the external magnetic field 4 is consistent with the flowing direction of the analyzed magnetic microsphere combination 7 flowing through the analyzing light beam, so that the anisotropic magnetic microsphere combination 7 with the superparamagnetic characteristic can gradually reach a consistent spatial orientation under the action of the magnetic field in the flowing process. As shown in FIG. 3, the spatial orientation of the dimers of the magnetic microspheres is initially random and does not coincide with the direction of flow; by accurately controlling the direction and the strength of external magnetic fields such as an electromagnetic field and the like, in the flowing process, the magnetic microsphere dimer gradually reaches a consistent spatial orientation under the action of the magnetic field, and finally all the magnetic microsphere dimer structures sequentially flow through the analysis light beams one by one according to the uniform spatial orientation (such as along the vertical direction) to measure characteristic optical signals. The same effect is also obtained with the trimer, tetramer, etc. magnetic microsphere combination 7.
Detection of various characteristic optical signals, including scattered light at different angles, fluorescence at different wavelengths, or excited luminescence. Specifically, the magnetic microspheres are not linked to a label, and physical characteristic optical signals, such as scattered light at different angles, are detected. The magnetic microspheres may also be attached to different labels, producing different characteristic optical signals. The magnetic microspheres are attached to a fluorescent label and excited to produce a fluorescent signal when flowing through an analytical beam. The magnetic microspheres are linked to a luminescent label and are excited to luminesce when flowing through an analytical beam or by a substrate.
The applied magnetic field can be realized in various forms, such as a magnetic field generated by a specially-shaped permanent magnet or an electromagnet of a coil winding; the direction and the strength of the magnetic field generated by the permanent magnet are fixed, the direction and the strength of the magnetic field generated by the electromagnet can be adjusted and controlled, and particularly, a plurality of groups of paired electromagnets can generate alternating rotating electromagnetic fields, so that the use is convenient.
Various characteristic optical signals such as scattered light at different angles can be detected by analyzing the characteristic optical signals, so that the types of dimers, trimers and tetramers in the sample liquid can be accurately distinguished, the number of each category is counted, and a lot of useful information such as the concentration of the target antigen to be detected and the like can be provided.
In particular, if the characteristics of the dimer in the sample liquid, such as the spatial structure characteristics, the distance between two microspheres, the optical characteristic signal, etc., can be accurately analyzed, more useful information can be provided, and a powerful means for studying the structure and interaction of molecules can be provided.
In the same principle, as shown in fig. 4, a capillary 5 may also be used to restrict the flow trajectory of the microspheres or combinations to be analyzed; meanwhile, a specially controlled magnetic field is applied, the direction and the strength of the external magnetic field 4 such as the electromagnetic field are accurately controlled, in the flowing process, the magnetic microsphere dimer gradually reaches a consistent spatial orientation under the action of the magnetic field, and finally all the magnetic microsphere dimer structures flow through the analysis light beam according to the uniform spatial orientation (such as the vertical direction) to measure characteristic optical signals.
The above embodiments may be implemented individually, or in any combination of two or more.
The method in the above embodiment is described in more detail with reference to specific examples.
In each of the following embodiments or examples,
unless otherwise indicated, all materials or processing techniques are conventional and commercially available materials or conventional processing techniques in the art.
Example 1:
among the bio-protein molecular detection assays, immunoassays based on antigen-antibody reactions are widely used for diagnosis of diseases. Procalcitonin PCT is a commonly used marker of inflammation, and its concentration in the plasma of normal humans is very low, typically less than 0.5 ng/mL. If the body has serious bacterial infection and sepsis, the index is rapidly increased, and the degree of the increase is positively correlated with the severity of inflammation. For analyzing the concentration of the target antigen molecule PCT, as shown in fig. 2, a magnetic microsphere is prepared by using a carboxylated superparamagnetic microsphere (Ademtech corporation) with a particle size of 500 nm, and the surface of the magnetic microsphere is coated with a first mouse monoclonal antibody (Mab-PCT, kyada biotechnology corporation) and a second mouse monoclonal antibody (Mab-PCT, kyada biotechnology corporation) against the PCT antigen, respectively, and the first antibody and the second antibody respectively correspond to different epitopes of the target antigen molecule PCT. Two kinds of magnetic microspheres with different antibodies react with a sample to be detected, such as a human serum sample, at 37 ℃ for 15 minutes, the first antibody and the second antibody are specifically combined with different epitopes of a target antigen molecule PCT respectively to form immune complexes, and the two magnetic microspheres are connected into a microsphere combination to form a Dimer structure (Dimer). According to different antigen concentrations, in the reaction solution, besides the dimer formed by two magnetic microspheres, there are the microsphere combination forms such as unbound monomer microspheres and polymers.
As shown in fig. 5, the sample liquid containing the magnetic microsphere conjugate 7 is sent to a flow optical detection system and analyzed. The flow path of the magnetic microspheres or the combination to be analyzed is limited by respectively adjusting the pressure and the flow rate of the sheath liquid 2 and the sample liquid 3, reducing the diameter of the sample liquid flow as small as 5 microns, and the sheath liquid arranges the magnetic microsphere combination 7 to be analyzed into a line and rapidly flows through an analysis light beam such as a detection laser beam one by one; meanwhile, a pair of electromagnets arranged above and below is used for applying a specially controlled external magnetic field 4, the direction and the strength of the magnetic field can be accurately controlled, for example, the direction of the external magnetic field is consistent with the flowing direction of the analyzed magnetic microsphere combination flowing through the analysis light beam, so that the magnetic microsphere combination with the superparamagnetic characteristic gradually reaches a uniform spatial orientation under the action of the magnetic field in the flowing process, as shown in fig. 5, the spatial orientation of the magnetic microsphere dimer is random at the beginning and is not consistent with the flowing direction; by accurately controlling the direction and the strength of the electromagnetic field, in the flowing process, the magnetic microsphere dimer gradually reaches a consistent spatial orientation under the action of the magnetic field, and finally, characteristic optical signals are measured by analyzing light beams in all the magnetic microsphere dimer structures in the vertical direction.
Specific analysis the beam was focused on the center of the sample stream using a 660nm red laser (HL6545, Hitachi corporation) with beam shaping. The particle or microsphere combination flows through a focused laser beam, characteristic optical signals detect scattered light at different angles, and a first photoelectric detector 8 is used for detecting the intensity of the scattered light at an angle which is 10 degrees away from the same direction of an analysis light beam; and detecting the scattered light intensity II by using a second photodetector 9 at an angle of 90 degrees to the vertical direction of the analysis light beam. The optical signal is converted into an electrical signal by a photodetector, the electrical signal is subjected to signal processing and digital sampling, the signal is received and processed by a computer, and a two-dimensional scattergram of data points obtained by measuring one sample is displayed as shown in fig. 6, wherein each point in the two-dimensional scattergram corresponds to characteristic optical signal measurement of one particle or microsphere combination, and the vertical axis of the two-dimensional scattergram is 10-degree scattered light intensity I, and the horizontal axis of the two-dimensional scattergram is 90-degree scattered light intensity II. .
Because various magnetic microsphere combinations pass through the analysis laser beam one by one in a uniform spatial direction, the characteristics of different microsphere combinations are optically analyzed, different groups of data points in a two-dimensional scatter diagram correspond to different microsphere combinations, as shown in figure 6, wherein, the first is a monomer microsphere, the second is a dimer, and the third is a polymer. Thus, the species and the number of the dimer, monomer microsphere, polymer, etc., in the sample solution, such as the total number or the number of each class, can be accurately distinguished. For a sample measurement and analysis, the number of the dimers in classification statistics is in direct proportion to the concentration of the antigen PCT in a certain target substance antigen concentration range, so that the concentration of the target substance antigen PCT to be detected can be obtained through analysis.
Example 2:
compared to example 1, most of the same is done, except that the fluid restriction in this example is achieved using capillary technology:
in particular, the capillary 5 can also be used to limit the flow trajectory of the microspheres or combinations to be analyzed; meanwhile, a specially controlled external magnetic field 4 is applied, as shown in fig. 4, by precisely controlling the direction and the strength of the external magnetic field 4, in the flowing process, the magnetic microsphere dimer gradually reaches a uniform spatial orientation under the action of the magnetic field, and finally all the magnetic microsphere dimer structures flow through the analysis laser beam one by one according to the uniform spatial orientation, such as the vertical direction, so as to measure characteristic optical signals.
The embodiments described above are described to facilitate an understanding and use of the invention by those skilled in the art. It will be readily apparent to those skilled in the art that various modifications to these embodiments may be made, and the generic principles described herein may be applied to other embodiments without the use of the inventive faculty. Therefore, the present invention is not limited to the above embodiments, and those skilled in the art should make modifications and alterations without departing from the scope of the present invention.
Claims (10)
1. A method for rapidly detecting a characteristic of a microscopic substance or structure, comprising the steps of:
(1) reacting and connecting the microscopic substances to be detected and analyzed with the magnetic microspheres to form a magnetic microsphere combination, and dispersing the magnetic microsphere combination in a sample solution to be analyzed;
(2) and simultaneously, strictly limiting and conforming the spatial orientation of the anisotropic magnetic microsphere combination in the sample liquid by applying fluid limitation and magnetic field control, singly and sequentially flowing through a detection analysis position along a specific track, and measuring and counting the characteristics of the microsphere combination one by one in a uniform spatial orientation to finish the method.
2. The method of claim 1, wherein the microscopic material is a protein molecule, a nucleic acid molecule, an organic polymer or a cell body that can be reacted with the magnetic microsphere.
3. The method of claim 2, wherein the protein molecules are antigens or antibodies.
4. The method according to claim 1, wherein the magnetic field control is specifically: and applying an external magnetic field to enable the anisotropic magnetic microsphere combination in the sample liquid to reach a consistent spatial orientation under the action of the magnetic field.
5. A method for rapid characterization of microscopic materials or structures according to claim 4 wherein the applied magnetic field is generated by a permanent magnet or an electromagnet of controllable direction and intensity.
6. The method of claim 4, wherein the direction of the applied magnetic field is aligned with the direction of the flow of the bound magnetic microspheres through the assay site.
7. The method for rapid characterization of microscopic materials or structures according to claim 1, wherein said fluid confinement is specifically: the microsphere-bound bodies are individually flowed sequentially through the detection and analysis site in a specific trajectory using a laminar fluid control technique, or a capillary technique using a single sample flow.
8. The method of claim 7, wherein the layered fluid control technique is specifically: the method is characterized in that the pressure and the flow velocity of sheath liquid and sample liquid are adjusted by utilizing the hydrodynamic focusing principle and adopting the technology of limiting the sample liquid flow by the sheath liquid, so that the diameter of the sample liquid flow is reduced.
9. The method of claim 1, wherein the characteristics of the microsphere conjugates are measured and analyzed by optical means, and the detection and analysis position is a position where the analyzing beam intersects with a sample fluid.
10. The method of claim 1, wherein the characteristic optical signal is scattered light, fluorescence or excited luminescence.
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