A CHI3L1 protein detection method based on microfluidic fiber optic sensing

Abstract

This application discloses a method for detecting CHI3L1 protein based on microfluidic fiber optic sensing, belonging to the field of biomedical detection technology. The method includes: providing a microstructured fiber optic sensing unit with a through-flow microfluidic channel; immobilizing a CHI3L1 protein-specific antibody on the inner surface of the microfluidic channel; injecting the sample to be tested into the channel, causing the CHI3L1 protein to be specifically captured; and achieving quantitative detection of the CHI3L1 protein by detecting the drift of fiber optic transmission spectral characteristic parameters caused by antigen-antibody binding. This invention combines microfluidics with fiber optic sensing, utilizing antibody immobilization on the inner wall of the microfluidic channel to increase the interaction efficiency between the light field and biomolecules, significantly improving detection sensitivity. It has the advantages of rapid detection, low sample consumption, and label-free operation, making it suitable for early non-invasive screening and dynamic monitoring of clinical liver fibrosis.

Description

Technical Field

[0001] This application belongs to the field of biomedical detection technology, specifically relating to a method for detecting CHI3L1 protein based on microfluidic fiber optic sensing. Background Technology

[0002] Liver fibrosis, a common pathological process in the development of various chronic liver diseases, can progress to end-stage liver diseases such as cirrhosis and even liver failure if not intervened in time. Currently, liver biopsy remains the "gold standard" for the diagnosis and staging of liver fibrosis. However, this method is invasive and may cause complications such as bleeding, infection, and pain. Furthermore, sampling errors and inter-observer variability exist, limiting its widespread application in dynamic monitoring and early screening.

[0003] In recent years, non-invasive serum biomarker testing has gained clinical attention due to its high safety, ease of operation, and good reproducibility. Chitosan-3-like protein 1 (CHI3L1), a biomarker closely related to the development and progression of liver fibrosis, is highly expressed in liver tissue, and its serum level is positively correlated with the degree of fibrosis. Therefore, achieving sensitive and rapid detection of CHI3L1 protein is of great significance for the early diagnosis and disease assessment of liver fibrosis.

[0004] However, in the process of realizing this application, existing CHI3L1 detection technologies mostly rely on enzyme-linked immunosorbent assay (ELISA) and magnetic microparticle chemiluminescence assay. These traditional methods usually have the following limitations: (1) limited detection sensitivity, making it difficult to achieve accurate quantification below 10 pg / mL, which cannot meet the needs of early fibrosis identification; (2) complex detection process, long time consumption, and reliance on large instruments and professional operators, making it difficult to achieve real-time continuous and rapid dynamic monitoring; (3) low integration of existing technology platforms, making it difficult to build portable and automated detection devices, which limits its application prospects in clinical point-of-care testing.

[0005] Therefore, there is an urgent need to develop a new method for detecting CHI3L1 protein that is more sensitive, faster, and easier to operate. Summary of the Invention

[0006] The purpose of this application is to provide a CHI3L1 protein detection method based on microfluidic fiber optic sensing, which can solve the problems of insufficient sensitivity, cumbersome and time-consuming detection process and low system integration of existing CHI3L1 detection methods, and achieve high sensitivity, rapid and real-time detection of CHI3L1 protein, providing a new method with high sensitivity, simple operation and rapid detection for early diagnosis and monitoring of liver fibrosis.

[0007] To address the aforementioned technical problems, this application provides a method for detecting CHI3L1 protein based on microfluidic fiber optic sensing, comprising the following steps: S1. A microfluidic fiber optic sensing unit is provided, the sensing unit comprising a section of microstructured optical fiber with cladding air holes, wherein liquid inlet micropores and liquid outlet micropores connected to the cladding air holes are constructed on the sidewall of the optical fiber to form a through microfluidic channel. S2. The inner surface of the microfluidic channel is chemically modified to immobilize the CHI3L1 protein-specific antibody onto the inner surface; S3. The CHI3L1 antigen solution to be tested is injected into the microfluidic channel through the inlet micropore, so that it flows through the inner surface on which the antibody is fixed; S4. Input detection light into the microfluidic fiber optic sensing unit and collect the output optical signal; S5. Based on the change in spectral characteristic parameters of the output optical signal relative to the reference signal, quantitative detection of CHI3L1 protein in the test solution is achieved.

[0008] Furthermore, in step S1, the outer periphery of the cladding air holes is a ring-shaped optical fiber guiding structure.

[0009] Furthermore, in step S1, the diameter of the cladding air pores is 2 μm - 5 μm.

[0010] Furthermore, in step S2, the chemical modification includes sequential hydroxylation, silanization modification, and antibody crosslinking fixation.

[0011] Furthermore, the hydroxylation treatment employs an acid washing or immersion method; the silane coupling agent used for the silanization modification is 3-aminopropyltriethoxysilane (APTES) or 3-glycidyl etheroxypropyltrimethoxysilane (GPTMS); and the antibody crosslinking reaction fixation uses glutaraldehyde or an EDC / NHS system.

[0012] Furthermore, step S2 also includes: after antibody fixation, using a protein-based blocking agent to seal the microfluidic channels.

[0013] Furthermore, in step S3, the CHI3L1 antigen solution to be tested is a biological fluid sample.

[0014] Furthermore, the biofluid sample is one of serum, plasma, and standard solution. Further, in step S3, the flow rate of the CHI3L1 antigen solution in the microfluidic channel is controlled at 1 μL / min - 20 μL / min, and the residence time is 5 min - 30 min.

[0015] Further, in step S4, the light source is a broadband light source with a wavelength range of 600 nm - 1700 nm, and the detection light is coupled into the microfluidic fiber sensing unit through a single-mode fiber; the spectral characteristic parameters are at least one of resonant wavelength, light intensity, or full width at half maximum (FWHM).

[0016] The CHI3L1 protein detection method based on microfluidic fiber optic sensing described in this application increases the contact area between antigen and antibody and the efficiency of light field interaction by cross-linking and immobilizing antibodies on the inner wall of the microfluidic channel. By precisely controlling the flow rate and residence time of the sample solution and combining this with highly sensitive fiber optic spectroscopy, ultra-micro-level detection of CHI3L1 protein is achieved. The detection process is continuous within the microfluidic channel, and spectral changes can be monitored in real time after sample injection, enabling rapid acquisition of detection results. Furthermore, this method requires no sample labeling and is easy to operate. In addition, the system is easily integrated with portable optical devices, providing a powerful new tool for the early screening and dynamic monitoring of liver fibrosis. Attached Figure Description

[0017] To more clearly illustrate the technical solutions in the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0018] Figure 1 This is a schematic flowchart of the CHI3L1 protein detection method based on microfluidic fiber optic sensing provided in the embodiments of this application; Figure 2 This is a schematic diagram of the microfluidic fiber optic sensing unit used in the detection method of this application. Figure 3 This is a standard curve of resonant wavelength shift versus concentration obtained by detecting different concentrations of CHI3L1 standard in the embodiments of this application; Figure 4 This is an example of the output spectrum of a sensing unit detecting CHI3L1 antigen solutions of different concentrations in this application. Among them, Figure 4 (a) Figure 4 (b) Figure 4 (c) Figure 4 (d) in the figure corresponds to the spectral response diagrams for concentrations of 1000 pg / mL, 100 pg / mL, 10 pg / mL and 1 pg / mL, respectively. Detailed Implementation

[0019] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0020] The following description, in conjunction with the accompanying drawings, details a CHI3L1 protein detection method based on microfluidic fiber optic sensing provided in this application, through specific embodiments and application scenarios.

[0021] like Figure 1 The CHI3L1 protein detection method based on microfluidic fiber optic sensing, as shown in one embodiment, includes the following steps: S1. A microfluidic fiber optic sensing unit is provided, the sensing unit comprising a section of microstructured optical fiber with cladding air holes, wherein liquid inlet micropores and liquid outlet micropores communicating with the cladding air holes are constructed on the sidewalls of the optical fiber to form a through-flow microfluidic channel; such as Figure 2 As shown, in this embodiment, the core of the microfluidic fiber optic sensing unit is a 2 cm long microstructured fiber, preferably a photonic crystal fiber with a ring-shaped optical signal distribution around the cladding air hole, wherein the diameter of the cladding air hole is 2 μm-5 μm. Using the photonic crystal fiber as the optical transmission path in this embodiment, its light guiding mechanism allows some light to penetrate into the cladding air hole region in the form of an evanescent field. When the air hole is filled with liquid, the change in the refractive index of the liquid inside the hole causes a change in the effective refractive index or transmission loss spectrum of the fiber, thereby achieving optical detection of biomolecules. The selection of a 2 cm fiber segment ensures sufficient interaction between the light source and the CHI3L1 antigen solution to be tested. The 2 μm-5 μm diameter of the cladding air hole results in a very high specific surface area in the fiber. The refractive index change caused by antigen-antibody binding is accumulated and amplified within the fiber segment, converting the weak bio-binding signal into a significant and measurable spectral shift, achieving accurate detection of extremely low concentration antigen solutions at the pg / mL level. The inlet and outlet micropores are located on both sides of the microstructured optical fiber, allowing liquid to enter from below and exit from above. After liquid entry, the sample fills the air pores, enabling the antigen to fully bind to the antibodies distributed on the inner wall of the air pores. Furthermore, this facilitates the real-time and precise capture of spectral changes by the peripheral optical fiber's light-guiding structure, achieving accurate detection of sample concentration.

[0022] S2. The inner surface of the microfluidic channel is chemically modified to immobilize the CHI3L1 protein-specific antibody onto the inner surface.

[0023] Chemical modification includes sequential hydroxylation, silanization, and antibody crosslinking fixation. The hydroxylation is performed using acid washing or impregnation. The silane coupling agent used for silanization is 3-aminopropyltriethoxysilane (APTES) or 3-glycidyl etheroxypropyltrimethoxysilane (GPTMS). The antibody crosslinking fixation uses glutaraldehyde or an EDC / NHS system. Hydroxylation of the microfluidic channel inner surface provides active sites for the subsequent silanization reaction. In this application, acid washing or impregnation can thoroughly remove organic contaminants from the microfluidic channel inner surface and increase the surface silanol density. Furthermore, it has the advantages of simple operation, low cost, and suitability for in-situ treatment of the microfluidic channel inner surface in this embodiment. The silane coupling agent used is 3-aminopropyltriethoxysilane (APTES), whose terminal amino group (-NH2) can react with various crosslinking agents (such as glutaraldehyde), and can also directly couple with the carboxyl group on the antibody molecule after activation. 3-Glycidyl etheroxypropyltrimethoxysilane (GPTMS) achieves one-step immobilization because its terminal epoxy group can directly react with the amino, thiol, or hydroxyl groups on the antibody molecule through a ring-opening reaction, making the process simpler. Both reactions are carried out in aqueous or alcohol phases, compatible with microfluidic channel operations, and the resulting silane layer is stable, effectively preventing antibody detachment due to non-specific adsorption or physical erosion during subsequent detection. The cross-linking reaction firmly connects the CHI3L1 antibody molecule to the silanized channel surface via covalent bonds. Glutaraldehyde, as a bifunctional cross-linking agent, has two aldehyde groups that can form Schiff bases with the amino groups at the silane layer terminal and on the antibody molecule, respectively, thereby achieving cross-linking. This method is fast and simple to operate. EDC / NHS is suitable for the coupling of carboxyl and amino groups. If the surface is aminosilane, the antibody carboxyl group needs to be activated first; if the antibody is directly immobilized via epoxy groups, this step can be simplified. This system is carried out under mild conditions, which is beneficial for maintaining antibody activity. Both methods ensure that antibodies are fixed on the inner surface of the microfluidic channel in the appropriate direction and density, maximizing their antigen-binding activity.

[0024] In this embodiment, after antibody immobilization, the microfluidic channels are blocked using a protein-based blocking agent. This protein-based blocking agent includes, but is not limited to, bovine serum albumin, casein, and skim milk powder. Blocking the microfluidic channels aims to seal the remaining active sites not occupied by the antibody, minimizing non-specific adsorption of non-target proteins in subsequent detection, reducing background signal, and improving the signal-to-noise ratio and specificity.

[0025] S3. The CHI3L1 antigen solution to be tested is injected into the microfluidic channel through the inlet micropore, and the flow rate and residence time of the CHI3L1 antigen solution in the microfluidic channel are controlled.

[0026] The CHI3L1 antigen solution to be tested is a biofluid sample, including but not limited to serum, plasma, and standard solutions. The target of the detection method in this application is the CHI3L1 protein, which has been identified as an important biomarker for liver fibrosis-related diseases. Serum or plasma is the most commonly used clinical sample for non-invasive diagnosis of liver fibrosis-related diseases; therefore, serum or plasma is selected as the antigen solution in this application. The flow rate of the CHI3L1 antigen solution in the microfluidic channel is controlled at 1 μL / min - 20 μL / min. When the flow rate in the microfluidic channel is lower than 1 μL / min, the sample flow rate is too low, which can easily lead to uneven diffusion or stagnation. When the flow rate in the microfluidic channel is higher than 20 μL / min, the target antigen molecules, when flowing through the sensing area, are too fast and do not have time to diffuse to the antibody fixation sites on the inner wall of the channel before flowing away, reducing the antigen capture efficiency and detection sensitivity within the channel. The CHI3L1 antigen solution has a residence time of 5-30 minutes in the microfluidic channel. A residence time of 5 minutes is the shortest necessary reaction time, ensuring that most antigen-antibody specific bindings are achieved under conditions of sufficiently high antibody affinity and appropriate concentration, making it suitable for applications requiring high detection speed. A residence time of 30 minutes allows for the diffusion and binding of more target molecules, especially suitable for extremely low concentration samples, helping to improve the reliability and signal intensity of low-concentration detection.

[0027] S4. Input detection light into the microfluidic fiber optic sensing unit using a light source, collect the output light signal after sample absorption through the optical detection unit, record the spectral characteristic parameters, and plot the quantitative standard curve.

[0028] like Figure 3 The method obtains the resonant wavelength drift standard curve by detecting different concentrations of CHI3L1 standard; in step S4, the light source is a broadband light source with a wavelength range of 600 nm - 1700 nm, and the detection light is coupled into the microfluidic fiber sensing unit through a single-mode fiber; the spectral characteristic parameters are at least one of resonant wavelength, light intensity or half-maximum width.

[0029] In this embodiment, the biological signal of antigen-antibody specific binding completed in step 3 is converted into a precisely measurable change in spectral parameters through an optical sensing mechanism. This eliminates the need for fluorescent or chemical labeling of the target analyte, avoiding the complexity, increased cost, and potential impact on bioactivity that labeling processes may introduce. A broadband light source is used to excite or observe the broad-spectrum optical response of the sensing unit before and after target binding. The wavelength range is 600 nm - 1700 nm, within the absorption window of the protein detected in this embodiment, minimizing interference from the absorbance of the biological sample itself and ensuring that the signal originates solely from antigen-antibody binding. Single-mode fiber coupling of the detection light effectively ensures the purity, stability, and precise control of the spatial mode of the input light field, facilitating the acquisition of a clear and resolvable spectral response. Spectral characteristic parameters, such as resonance wavelength, intensity, and full width at half maximum (FWHM), quantitatively characterize the comprehensive optical effects caused by changes in refractive index or thickness of the sensing region due to antigen-antibody binding from different dimensions. The combination of multiple parameters enhances the reliability and information dimensionality of the method.

[0030] S5. Based on the change in spectral characteristic parameters, quantitative detection of CHI3L1 protein in the test solution is achieved.

[0031] This step is the final step in achieving quantitative concentration. For example... Figure 4 As shown, the binding of the test sample to the antibody causes a characteristic change in the output spectrum of the sensing unit. By automatically extracting characteristic parameters from the spectrum and then substituting these parameters into the input... Figure 3 Once a quantitative standard curve is established, the accurate concentration of CHI3L1 protein in the test sample can be calculated.

[0032] This application presents a microfluidic fiber optic sensing-based method for detecting CHI3L1 protein. By cross-linking and immobilizing antibodies on the inner wall of a microfluidic channel, the contact area between the antigen and antibody is increased. Precise control of the flow rate and residence time of the antigen solution enhances the antigen-antibody binding efficiency. Combined with highly sensitive optical detection, this method achieves ultra-micro-level detection of CHI3L1 protein (1 pg / mL). The detection process is continuous within the microfluidic channel, and spectral changes can be monitored in real time after sample injection, enabling rapid acquisition of results. Based on an integrated sensing unit, the operation is simple, and integration with a portable light source and spectrometer allows for portable detection, broadening its application scenarios.

[0033] The application and effects of the present invention will be further illustrated below through specific embodiments: Example 1: Fabrication and characterization of microfluidic fiber optic sensing units: 1.1 Selection and Fabrication of Microstructured Optical Fibers: An optical fiber guiding structure with a ring-shaped distribution of cladding air holes was selected. The fiber core diameter was 10 μm, the cladding air hole diameter was approximately 3.2 μm, and the spacing was 6.5 μm. A 2.0 cm length of optical fiber was cut, and single-mode optical fibers were welded to both ends as optical input and output interfaces. Micro-holes were fabricated on the side of the optical fiber using a femtosecond laser micromachining system: liquid inlet and liquid outlet micro-holes were symmetrically distributed at both ends of the fiber, with a center-to-center spacing of 1.0 cm; the hole diameter was 50±5 μm, and the depth extended to completely penetrate the cladding air holes; after fabrication, the fiber was ultrasonically cleaned with isopropanol for 10 min and dried with nitrogen for later use.

[0034] 1.2 Chemical modification of the inner surface of the microfluidic channel: (1) Hydroxylation treatment: The sensing unit was immersed in Piranha solution (concentrated sulfuric acid: 30% hydrogen peroxide = 9:1, v / v) and heated in a water bath at 80 ℃ for 1 h. After removal, it was rinsed with deionized water and ethanol in sequence, and dried with nitrogen to obtain an active surface rich in silanol groups. (2) Silanization modification: A 2% (v / v) ethanol solution of 3-aminopropyltriethoxysilane (APTES) was prepared and injected into the microfluidic channel and reacted at room temperature for 2 h. Then it was rinsed with ethanol 3 times and cured in an oven at 110 ℃ for 30 min. (3) Antibody fixation: The CHI3L1 monoclonal antibody was diluted with 10 mM phosphate buffer solution (pH=7.4) to a final concentration of 20 μg / mL, and 0.25% glutaraldehyde (v / v) was added as a crosslinking agent. The mixture was injected into the channel and incubated at room temperature for 2 h. Unbound antibodies were rinsed with phosphate buffer solution. (4) Blocking treatment: Inject blocking solution containing 1% bovine serum albumin in phosphate buffer solution, let stand at room temperature for 1 h, rinse with phosphate buffer solution and store at 4°C for later use.

[0035] 1.3 Structural Characterization and Detection: The morphology of the micropores was observed using a scanning electron microscope to confirm that the pore size was uniform and the channels were interconnected. Optical microscopy revealed that the inner walls of the channels after antibody modification were uniformly covered, with no blockage.

[0036] Example 2 Establishment of the CHI3L1 protein detection standard curve: 2.1 Standards and Reagents: Recombinant human CHI3L1 protein was prepared in phosphate buffer containing 0.1% bovine serum albumin to form the following concentration gradients: 0 pg / mL, 0.1 pg / mL, 0.25 pg / mL, 0.5 pg / mL, 0.75 pg / mL, 1.0 pg / mL, 1.5 pg / mL, 2.0 pg / mL, 5 pg / mL, 10 pg / mL, 50 pg / mL, 100 pg / mL, 500 pg / mL, 1 ng / mL, 5 ng / mL, and 10 ng / mL.

[0037] 2.2 Detection system setup: Light source: broadband light source preheated for 30 min, wavelength range: 600 nm - 1700 nm; spectrometer, integration time extended to 2 s, 10 spectra were continuously collected for each sample and the average value was taken; microfluidic injection pump, flow rate range 0.1 μL / min - 100 μL / min; real-time data acquisition software to record the spectrum, the entire detection system was set at 25.0 ± 0.5 ℃.

[0038] 2.3 Detection steps: (1) Connect the sensing unit to the optical path and microfluidic system; (2) Inject phosphate buffer baseline solution at a flow rate of 5 μL / min and record the initial resonance wavelength. l 0; (3) Inject standards of various concentrations sequentially at a flow rate of 5 μL / min and a residence time of 15 min; (4) Record the resonant wavelength after stabilization at each concentration point. l Calculate the drift amount Dl = l - l 0; (5) Rinse three times with phosphate buffer solution, restore baseline and then proceed with the next concentration test.

[0039] 2.4 Standard Curve: The average resonance wavelength shift corresponding to each concentration point was measured. Dl At CHI3L1 concentration C (pg / mL) is the x-axis. Dl The vertical axis represents the sum of the vertical and horizontal axes. Within the concentration range of 0.1 pg / mL to 10 ng / mL, the data exhibit a linear relationship. Dl =0.280 C +0.272 ( R 2 =0.997) 2.5 Validation of the detection limit Repeat assays were performed using a low concentration of CHI3L1 standard (1.0 pg / mL) close to the blank concentration (n=10). Simultaneously, parallel assays were performed using a blank sample containing only buffer (0 pg / mL) (n=10) to assess background signal fluctuations.

[0040] 2.6 Test results of samples: (1) Blank sample (0 pg / mL): Resonance wavelength shift ( Dl The mean value is 0.02 nm, and the standard deviation is 0.008 nm.

[0041] (2) 1.0 pg / mL CHI3L1 sample: resonant wavelength shift ( Dl The mean value is 0.25 nm, and the standard deviation is 0.03 nm.

[0042] 2.7 Evaluation of test results: According to international practice, a signal-to-noise ratio (S / N) ≥ 3 is used as the criterion for the detection limit. Here, signal (S) is the difference between the average signal of a 1.0 pg / mL sample and the average signal of a blank sample, and noise (N) is the standard deviation of the blank sample. The calculated S / N ≈ 28.75, which is much greater than 3.

[0043] The results showed that this method can generate a clear and distinguishable detection signal for 1.0 pg / mL CHI3L1 protein, verifying that its detection limit can reach 1.0 pg / mL.

[0044] Example 3 Human serum sample testing 3.1 Sample source: With the approval of the ethics committee, serum samples were collected from 24 patients with liver fibrosis (F1-F4 stage) and 24 healthy volunteers and stored at -80 ℃.

[0045] 3.2 Sample pretreatment: Serum samples were serially diluted with phosphate buffer solution according to the concentration ratio, centrifuged (10,000 rpm, 10 min), and the supernatant was directly detected.

[0046] 3.3 Comparative reagents (1) Detection method of this application: The microfluidic fiber optic sensing unit prepared in Example 1 is used, and the detection conditions are the same as in Example 2 (flow rate 5 μL / min, residence time 15min). (2) Commercial enzyme-linked immunosorbent assay (ELISA): The Human Chitinase 3-like 1 (CHI3L1) ELISA Kit (Cusabio, Cat No. CSB-E13111h) was used. The limit of detection of this kit is 15.6 pg / mL.

[0047] (3) Magnetic microparticle chemiluminescence method: Chitosan polysaccharide enzyme 3-like protein 1 detection kit (magnetic microparticle chemiluminescence method) (CHI-100Z) was used. The detection limit of this kit is 1.5 ng / mL (i.e. 1500 pg / mL).

[0048] 3.4 Parallel Testing and Results The above three methods were used to perform parallel tests on all 48 serum samples. The results are shown in Table 1. Table 1. Comparison of CHI3L1 detection results in clinical serum samples using three methods 3.4 Discussion and Conclusion This embodiment fully verifies the superiority of the technical solution of the present invention through parallel comparison with actual clinical samples: (1) High sensitivity and early detection capability: In the context of low concentration in healthy individuals, this detection method can achieve 100% detection and quantification, while the two control methods have a large number of "undetected" cases due to insufficient sensitivity. For patients in the early stage of liver fibrosis (F1 / F2), the concentration measured by this method is significantly higher than that of traditional detection methods (commercial enzyme-linked immunosorbent assay and magnetic particle chemiluminescence assay), indicating that it is more sensitive to small increases in concentration and is more conducive to the early diagnosis of the disease.

[0049] (2) High accuracy and reliability: The results of this detection method are highly correlated with those of commercial enzyme-linked immunosorbent assay, proving the accuracy of the quantitative results of this method, and not sacrificing accuracy for sensitivity.

[0050] (3) Small sample volume and fast detection speed: The sample volume and detection time required by this detection method are significantly smaller than those of traditional detection methods (commercial enzyme-linked immunosorbent assay and magnetic particle chemiluminescence assay). (4) More diverse application scenarios: This detection method can be an integrated portable system, which has a wider range of applications than traditional detection methods (commercial enzyme-linked immunosorbent assay and magnetic particle chemiluminescence assay), and is not limited to the laboratory.

[0051] The embodiments of this application have been described above with reference to the accompanying drawings. Specific examples have been used to illustrate the principles and implementation methods of this application. The description of the above embodiments is only for the purpose of helping to understand the core ideas of this application. However, this application is not limited to the specific embodiments described above. The specific embodiments described above are merely illustrative and not restrictive. Those skilled in the art can make many other forms under the guidance of this application without departing from the spirit and scope of the claims, and all of these forms are within the protection scope of this application.

Claims

1. A method for detecting CHI3L1 protein based on microfluidic fiber optic sensing, characterized in that, Includes the following steps: S1. A microfluidic fiber optic sensing unit is provided, the sensing unit comprising a section of microstructured optical fiber with cladding air holes, wherein liquid inlet micropores and liquid outlet micropores connected to the cladding air holes are constructed on the sidewall of the optical fiber to form a through microfluidic channel. S2. The inner surface of the microfluidic channel is chemically modified to immobilize the CHI3L1 protein-specific antibody onto the inner surface; S3. The CHI3L1 antigen solution to be tested is injected into the microfluidic channel through the inlet micropore, so that it flows through the inner surface on which the antibody is fixed; S4. Input detection light into the microfluidic fiber optic sensing unit and collect the output optical signal; S5. Based on the change in spectral characteristic parameters of the output optical signal relative to the reference signal, quantitative detection of CHI3L1 protein in the test solution is achieved.

2. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, In step S1, the outer periphery of the cladding air holes is a fiber optic light guide structure with a ring-shaped distribution.

3. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 2, characterized in that, In step S1, the diameter of the cladding air pores is 2 μm - 5 μm.

4. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, In step S2, the chemical modification includes sequential hydroxylation, silanization, and antibody crosslinking fixation.

5. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 4, characterized in that, The hydroxylation treatment is performed by acid washing or immersion; the silane coupling agent used for the silanization modification is 3-aminopropyltriethoxysilane (APTES) or 3-glycidyl etheroxypropyltrimethoxysilane (GPTMS); the antibody crosslinking reaction is fixed using glutaraldehyde or an EDC / NHS crosslinking system.

6. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, Step S2 further includes: after antibody fixation, using a protein-based blocking agent to seal the microfluidic channels.

7. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, In step S3, the CHI3L1 antigen solution to be tested is a biological fluid sample.

8. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 7, characterized in that, The biofluid sample is one of serum, plasma, and standard solution.

9. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, In step S3, the flow rate of the CHI3L1 antigen solution in the microfluidic channel is controlled at 1 μL / min - 20 μL / min, and the residence time is 5 min - 30 min.

10. The CHI3L1 protein detection method based on microfluidic fiber optic sensing according to claim 1, characterized in that, In step S4, the light source is a broadband light source with a wavelength range of 600 nm to 1700 nm, and the detection light is coupled into the microfluidic fiber sensing unit through a single-mode fiber; the spectral characteristic parameters are at least one of resonant wavelength, light intensity, or full width at half maximum (FWHM).

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