Aptamer biosensor and method for CA125 detection

Abstract

The invention relates to the field of biomolecule detection, and provides an aptamer biosensor and method for CA125 detection. According to the imaging weak measurement system, two beam expanding lenses are introduced, so that the weak measurement system is improved, the quality of light beams is improved, and multi-channel detection is facilitated. In addition, the anti-CA125 aptamer is used as a biological probe to be modified on a dopamine film on the surface of the sensing chip, so that high-throughput detection of the tumor protein CA125 is realized. Wherein the prism is combined with the detection chip in advance, is modified by dopamine, and is used as a sensing surface of the sensing chip, so that functionalization of the surface of the sensing chip is facilitated. The aptamer biosensor based on imaging weak measurement provided by the invention has the characteristics of high throughput, rapidness, sensitivity, simplicity, convenience and low cost, shows a good application prospect in CA125 detection, and provides a new tool for early diagnosis and monitoring of ovarian cancer.

Description

Technical Field

[0001] The present invention relates to the field of biomolecule detection, and in particular to an aptamer biosensor and a method for CA125 detection. Background Art

[0002] Cancer Antigen 125 (CA125) is a tumor marker widely used in the diagnosis and monitoring of ovarian cancer. CA125 is a high molecular weight glycoprotein that is mainly secreted by ovarian cancer cells, but can also be secreted by other types of cancer cells and cells of certain benign diseases.

[0003] At present, the main methods used for CA125 detection are as follows:

[0004] (1) Enzyme-linked immunosorbent assay (ELISA): Through antigen-antibody reaction, detection is performed using enzyme-labeled secondary antibodies. This method has high sensitivity and specificity, but the operation is complicated and the detection time is long.

[0005] (2) Radioimmunoassay (RIA): It uses radiolabeled antibodies for detection and has high sensitivity, but requires radioisotopes and has certain safety risks.

[0006] (3) Chemiluminescent immunoassay (CLIA): It uses chemiluminescent reaction to detect antigen-antibody conjugates. It has high sensitivity, but the instrumentation is expensive.

[0007] Although the existing CA125 detection methods have been widely used in clinical practice, they still have the following shortcomings: limited detection sensitivity: some early cancers or low expression of CA125 levels are difficult to detect; long detection time: many methods require a long reaction time and cannot achieve rapid detection; complex operation: complex sample pretreatment and multi-step operation are required, which is not suitable for large-scale screening and on-site testing; high cost: some methods require expensive instruments and reagents, which increases the cost of detection.

[0008] Aptamer is a synthetic single-stranded oligonucleotide or peptide molecule composed of nucleic acid, which can specifically bind to the target molecule. Aptamer biosensors utilize the high specificity and high affinity of aptamers. Compared with traditional antibody sensors, they have the following advantages: High specificity and high sensitivity: Aptamers can be obtained through in vitro screening (SELEX) and have high specificity and affinity for target molecules; Good stability: Aptamers are more stable than antibodies to temperature, pH value and organic solvents, and are suitable for detection in a variety of complex environments; Low cost: The synthesis cost of aptamers is low, and the consistency between batches is good, which is suitable for large-scale production.

[0009] Since it was proposed in 1988, weak measurement technology has played an important role in the field of high-precision detection. Especially after the universally valuable frequency domain weak measurement was proposed in 2016, weak measurement biosensors have shown amazing potential in label-free biomolecule detection. However, with the continuous improvement of detection requirements, spectral point-based weak measurement aptamer biosensors can no longer meet current detection requirements and need to be further improved. Summary of the invention

[0010] To solve the above problems, the present invention proposes an aptamer biosensor and method for CA125 detection.

[0011] To achieve the above purpose, the present invention adopts the above technical solution:

[0012] An aptamer biosensor for CA125 detection, characterized in that it comprises an imaging weak measurement system and a sensor chip;

[0013] The imaging weak measurement system comprises a light source, a collimating lens, a first beam expander lens, a second beam expander lens, a front selective polarizer, a prism, a quarter wave plate, a rear selective polarizer, an imaging lens and a camera which are sequentially arranged in an optical path. The light emitted by the light source is collimated by the collimating lens, sequentially passes through the first beam expander lens and the second beam expander lens, and then passes through the front selective polarizer to enter the interior of the prism. The light beam is totally internally reflected inside the prism, and then sequentially enters the quarter wave plate, the rear selective polarizer to reach the imaging lens and the camera. The camera is used to record the image imaging of the total internal reflection interface.

[0014] The sensor chip includes the prism and the detection chip, and the detection chip is provided with a liquid flow channel. The prism and the detection chip are combined together. The detection chip is vertically placed on one side of the prism.

[0015] The detection chip is made of resin material, and the prism is directly bonded to the chip using AB glue. Use a syringe or peristaltic pump to slowly draw the solution into the chip channel.

[0016] The light source is collimated by a collimating lens to ensure that the emitted light is collimated light.

[0017] The first beam expanding lens and the second beam expanding lens are a group of lenses, which are used to expand the light source on the basis of ensuring collimation, so that the light intensity of the light beam is more uniform.

[0018] The angle between the front selective polarizer and the rear selective polarizer is 89° to 91°, so as to achieve polarization state adjustment of the light source.

[0019] The quarter wave plate is used to convert the phase difference into optical rotation.

[0020] The method for preparing the sensor chip comprises a substrate and a detection chip, wherein the substrate is a prism.

[0021] The detection chip is provided with a plurality of liquid flow channels; the prism and the detection chip are bonded together by AB glue to obtain a sensor chip.

[0022] The multiple liquid flow channels serve as experimental channels and reference channels in the experiment.

[0023] The CA125 detection method is based on the aptamer biosensor for CA125 detection, and the following steps are performed:

[0024] S1: Solution A and solution B are sequentially introduced into the liquid flow channel of the sensor chip to obtain real-time images when different solutions are passed through. Solution A and solution B are selected from pure water, PBS phosphate buffer solution, and sodium chloride solution to be used for statistical system sensitivity: ΔI is the statistical relative intensity change, ΔC is the concentration change, δT is the standard deviation of the system when stable, and δ is the system sensitivity.

[0025] S2: Start CA125 protein molecule detection experiment:

[0026] S21: depositing a binding layer in the liquid flow channel of the sensor chip to prepare a first sensing interface;

[0027] S22: coupling anti-CA125 aptamer molecules on the surface of the first sensing interface to prepare a second sensing interface;

[0028] S23: using a protein-free blocking solution to block the non-specific binding sites on the second sensor interface, and completing a phosphate buffer solution washing step to obtain a sensor chip that can be used for detection;

[0029] S3: Construct a prism sensor chip with anti-CA125 aptamer molecules as the recognition layer to achieve recognition and capture of CA125 protein molecules. By calculating the difference between the initial light intensity and the final light intensity when recognizing CA125 protein molecules, determine the different light intensity differences generated when capturing CA125 protein molecules of different concentrations. According to the real-time binding curve of the change of the output light intensity △I with time t and the different light intensity differences, the binding rate constant Ka and the standard binding curve formula of the binding kinetic curve are obtained to analyze the characteristics of the binding reaction. When detecting unknown concentrations of CA125 protein molecules, the concentration of CA125 protein molecules can be calculated according to the light intensity change and the standard binding curve formula to achieve CA125 detection.

[0030] The binding layer is a dopamine membrane.

[0031] The step of depositing a binding layer on the surface of the detection chip comprises:

[0032] The substrate surface is cleaned with a phosphate buffer solution; dopamine is prepared with a Tris solution of pH 8.0-9.0 to obtain a dopamine-Tris solution; the liquid flow channel of the sensor chip is modified with the dopamine-Tris solution for 10-60 minutes, and then the phosphate buffer solution is used to complete the cleaning.

[0033] Preferably, the imaging lens is used to achieve imaging of the liquid flow channel of the detection chip.

[0034] Preferably, the camera is used for image recording of the chip.

[0035] The present invention has the following beneficial effects:

[0036] In view of the shortcomings of the existing CA125 detection methods, the present invention proposes a weak measurement aptamer biosensor, a sensor chip and a method for CA125 detection. The imaging weak measurement system of the present invention improves the weak measurement system by introducing two beam expansion lenses, improves the light beam quality, and facilitates the realization of multi-channel detection. In addition, the anti-CA125 aptamer is used as a biological probe to modify the dopamine film on the interface of the sensor chip to achieve the detection of the tumor protein CA125. Among them, the prism is pre-combined with the detection chip and modified with dopamine, and used as the sensing surface of the sensor chip, which is conducive to the functionalization of the surface of the sensor chip. The aptamer biosensor based on imaging weak measurement provided by the present invention has the characteristics of high throughput, rapidity, sensitivity, simplicity and low cost, and shows good application prospects in CA125 detection, providing a new tool for early diagnosis and monitoring of ovarian cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 Modification and detection process for imaging weak measurement systems and sensor chips.

[0038] Numbers in the figure:

[0039] 1. Light source, 2 and 3. Collimating lens, 4. First beam expander lens, 5. Second beam expander lens, 6. Front polarizer, 7. Prism, 8. Detection chip, 9. Quarter wave plate, 10. Rear polarizer, 11.

[0040] Imaging lens, 12. Camera.

[0041] Figure 2a This is a picture of the multi-channel detection chip.

[0042] Figure 2b The sensor chip is obtained by bonding the prism and the detection chip.

[0043] Figure 3 To test the stability of the system using sodium chloride solutions of different concentrations.

[0044] Figure 4 The real-time reaction curve of the whole experimental process detected by the weak measurement system, including dopamine deposition, anti-CA125 aptamer fixation, blocking, PBS stabilization and CA125 protein molecule detection process.

[0045] Figure 5 These are pictures of the experimental flow channel and reference flow channel of the sensor chip taken by a camera during a certain experiment.

[0046] Figure 6a It is to detect different concentrations of CA125 protein molecules and record the real-time change curve of light intensity. By counting the difference between the initial light intensity and the final light intensity during this period, different light intensity differences can be obtained, which is the light intensity change caused by different concentrations of CA125 molecules.

[0047] Figure 6b The concentration of CA125 was used as the abscissa, and the different changes in light intensity caused by CA125 were used as the ordinate. Langmuir fitting was performed on these data to obtain the standard binding curve formula.

[0048] Figure 7 It is the result of specific experiment, which tests the feasibility of the sensor chip in detecting other proteins. DETAILED DESCRIPTION

[0049] The following is a detailed description of the embodiments of the present invention. It should be noted that the following description is only exemplary and is not intended to limit the scope and application of the present invention.

[0050] The present invention provides an aptamer biosensor and method for CA125 detection. To make the purpose, technical method and application of the present invention clearer and more specific, the present invention is further described in detail below. It should be understood that the embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0051] Example:

[0052] The reagents used in this experiment are listed below.

[0053] Alpha-fetoprotein (AFP) and CA125 were purchased from Beijing Zhipeng Company (Beijing, China). Human IgG and bovine serum albumin (BSA) were purchased from Bioss Biological Company (Beijing, China). Sodium chloride (NaCl), dopamine hydrochloride, and Tris buffer were purchased from Shanghai Aladdin Biological Co., Ltd. (Shanghai, China). Phosphate buffered saline (PBS) was purchased from Beijing Solarbio Technology Co., Ltd. (Beijing, China), and protein-free blocking solution (2%) was purchased from Shanghai Shenggong Biotechnology Co., Ltd. (Shanghai, China).

[0054] The CA125 aptamer sequence was synthesized by Shanghai Sangon Biotechnology Co., Ltd. (Shanghai, China).

[0055] The prism used in the present invention is a ZF6 prism, which is purchased from Fuzhou Alpha Optics Co., Ltd. (Fuzhou, Fujian, China).

[0056] The anti-CA125 aptamer sequence used in the present invention is artificially synthesized by Shanghai Shenggong Biotechnology Co., Ltd. The specific sequence used is: 5′-NH2CTCACTATAGGGAGACAAGAATAAACGCTCAA-3′.

[0057] In an embodiment of the present invention, an aptamer biosensor for CA125 detection includes building an imaging weak measurement system based on total internal reflection and preparing a sensor chip;

[0058] The imaging weak measurement system comprises a light source 1, collimating lenses 2 and 3, a first beam expander lens 4 and a second beam expander lens 5, a front selective polarizer 6, a prism 7, a quarter wave plate 9, a rear selective polarizer 10, an imaging lens 11 and a camera 12 which are sequentially arranged in an optical path. The light emitted from the light source 1 is collimated by the collimating lenses 2 and 3, sequentially passes through the first beam expander lens 4 and the second beam expander lens 5, and then enters the interior of the prism 7 through the front selective polarizer 6. After the light beam is totally internally reflected inside the prism 7, it sequentially enters the quarter wave plate 9, the rear selective polarizer 10 and reaches the imaging lens 11 and the camera 12. The camera 12 is used to record the image imaging of the total internal reflection interface; the angle between the front selective polarizer 6 and the rear selective polarizer 7 is 89° to 91°, so as to realize the polarization state adjustment of the light source.

[0059] The sensor chip includes a prism 7 and a detection chip 8. The detection chip 8 is provided with a plurality of liquid flow channels serving as experimental channels and reference channels in the experiment.

[0060] Weak measurement can amplify tiny signals by using the properties of biological signals, and can be used with aptamer sensor chips to achieve high-sensitivity detection of biological molecules. Figure 1 shown.

[0061] exist Figure 1In the figure, the light source 1 used is an LED, and the emitted light is collimated by collimating lenses 2 and 3. The light beam is expanded by the first and second beam expander lenses 4 and 5 in sequence to ensure collimation and minimize the influence of light intensity changes. The collimated light is selectively filtered by a polarizer. After passing through the front selective polarizer 6, the light passes through a ZF6 prism 7 with a refractive index of 1.73. During this process, the light beam undergoes total internal reflection on the inner surface of the prism 7, resulting in a significant phase difference between p-polarized light and s-polarized light. In order to convert the phase difference into a measurable optical rotation, an achromatic quarter-wave plate 9 is placed behind the prism. Subsequently, the collimated light passes through the rear selective polarizer 10. The entire internal reflection interface is projected onto the camera 12 through the imaging lens 11 for imaging.

[0062] When a protein molecule is captured by an aptamer immobilized on a detection chip, the effective refractive index of the detection chip surface increases because the refractive index of the protein molecule is greater than that of the aqueous solution. During total internal reflection at the chip surface, the phases of p-polarized light and s-polarized light respond differently to the change in the surface refractive index. By measuring the phase difference between p-polarized light and s-polarized light, molecules can be detected in real time and without labels. Weak measurement is a relatively non-traditional method characterized by its minimal invasiveness, which means that the measurement process does not significantly change the state of the system.

[0063] This technique uses the weak value amplification effect to amplify the original weak signal by several orders of magnitude. In this case, weak measurement can be used to analyze the phase difference between p-polarized light and s-polarized light. By analyzing the phase difference, the concentration of the corresponding protein molecules in the solution can be determined.

[0064] In the embodiment of the present invention, the aptamer biosensor chip for CA125 detection is prepared. The prism and the detection chip are glued together with AB glue to form a sensor chip, wherein the detection chip is respectively as follows: Figure 2a As shown, the sensor chip is Figure 2b shown.

[0065] 1. CA125 detection method

[0066] Follow these steps:

[0067] S1: Solution A and solution B are introduced into the liquid flow channel of the sensor chip in sequence to obtain real-time images when different solutions are passed through. Solution A and solution B are selected from pure water, PBS phosphate buffer solution, and sodium chloride solution to be used for statistical system sensitivity: ΔI is the statistical relative light intensity change, ΔC is the concentration change, δT is the standard deviation of the system when stable, and δ is the system sensitivity;

[0068] S2: Start CA125 protein molecule detection experiment:

[0069] S21: Depositing a binding layer dopamine film on the surface of the detection chip: cleaning the substrate surface with a phosphate buffer solution; preparing dopamine with a Tris tris hydroxymethylaminomethane solution at pH 8.0-9.0 to obtain a dopamine-Tris solution; modifying the substrate with the dopamine-Tris solution for 10-60 minutes, and then cleaning with a phosphate buffer solution to obtain a sensing interface 1;

[0070] S22: coupling anti-CA125 aptamer molecules on the surface of the first sensing interface to prepare a second sensing interface;

[0071] S23: blocking the non-specific binding sites on the second sensor interface, and completing a phosphate buffer solution washing step to obtain a sensor chip;

[0072] S3: Construct a prism sensor chip with anti-CA125 aptamer molecules as the recognition layer to achieve recognition and capture of CA125 protein molecules, obtain the binding kinetic curve of the output light intensity △I changing with time t, and obtain the binding rate constant Ka based on the binding kinetic curve.

[0073] 2. System stability evaluation:

[0074] The sensor chip was placed in the imaging weak measurement system, and pure water, 2 g / L, 4 g / L, 6 g / L, 8 g / L, and 10 g / L sodium chloride solutions were injected into the multiple liquid flow channels of the detection chip in turn to test the stability of the system. The results are as follows: Figure 3 shown.

[0075] 3. Experimental process of characterization of imaging weak measurement system:

[0076] In the modification process of the sensor chip, the flow channel is cleaned with 10mM PBS solution, and then 1 gram per liter of dopamine-tris solution (ph-8.5) is sent into the flow channel to react for 20 minutes, forming a sticky self-assembled layer on the surface of the flow channel, so that the prism surface has active groups, such as amino groups. Into different flow channels washed with PBS, an amino anti-CA125 aptamer solution with a concentration of 2uM is injected at a rate of 38uL / min and reacted for 60min. Subsequently, the flow channel is washed with PBS solution, and a protein-free blocking solution is injected into the flow channel to occupy non-specific binding sites and achieve the effect of blocking blank sites. After washing with PBS solution for 10min, different aptamer sensor chips are obtained.

[0077] The prepared aptamer sensor chip was used to capture CA125 protein molecules and react for 60 minutes. After the reaction, the unreacted CA125 protein molecules can be easily washed away in the chip flow channel by PBS washing buffer. The curve of the whole experimental process described above is as follows: Figure 4 shown.

[0078] 4. Gradient experiment:

[0079] Repeat the above-mentioned sensor chip preparation. After the chip preparation is completed, store it in a refrigerator at 2-8°C.

[0080] Using the sensor chip prepared above, PBS buffer was injected into the flow channel, reacted for 30 minutes, and the change of the average light intensity response value of each flow channel was recorded as the system background noise signal. After the system enters a stable state, the CA125 protein molecule solution is passed into the experimental channel, and the reference channel is passed into the PBS buffer. The system records the light intensity changes of the experimental channel and the reference channel in real time. The CA125 protein molecule solution is diluted with PBS buffer, and the concentrations are 5U / mL, 10U / mL, 20U / mL, 40U / mL, 60U / mL, and 100U / mL, respectively. The CA125 protein molecule is reacted in the flow channel for 60 minutes, then the PBS solution is passed, and washed to remove the CA125 protein molecules that do not participate in the reaction, and the entire experimental process is completed.

[0081] The entire experimental process uses the camera's built-in software to capture images of the flow channel, such as Figure 5 shown.

[0082] After the experiment was completed, Matlab was used to process the image and obtain the light intensity changes during the chip capturing CA125 protein molecules ( Figure 6a ).

[0083] Figure 6a These are the response curves for detecting CA125 protein molecules at different concentrations. The corresponding average relative light intensity changes are 37.24 au, 52 a.u., 85.9 au, 119 a.u., 151 a.u. and 193.7 au, respectively.

[0084] like Figure 6a As shown, the initial curve is a short section of PBS, which is used as the baseline to prove that the system has stabilized. Because PBS does not bind to the prism surface, PBS does not cause the curve to drift. However, when CA125 is captured by the anti-CA125 aptamer pre-immobilized on the prism surface, it causes the refractive index of the solution on the prism surface to change, which ultimately manifests as a change in light intensity, that is, a drift of the curve. Figure 6a The experimental results show that the concentration of CA125 protein molecules is positively correlated with the relative light intensity change. The light intensity changes caused by the specific reaction of CA125 protein with different concentrations and its aptamer are different.

[0085] For the Langmuir fit ( Figure 6b), the fitting equation of the response process of the experimental gradient relative to the change of light intensity is: I = (260.82*23.53*10 -3 *C) / (1+23.53*10 -3 *C), where I and C are the response relative light intensity (au) and the concentration of the analyte CA125 protein molecule (U / mL), respectively. Langmuir fitting is applicable to the reaction that the surface type changes due to the formation of a monolayer uniform adsorption after dopamine binds to the amino-aptamer, which is similar to our experimental binding process. The standard deviation of the system at stability is δT, which is 4.12au. When this value is exceeded, it can be assumed that the system can effectively distinguish the relative light intensity changes caused by changes in CA125 concentration and the fluctuations of the system itself. To ensure the accuracy of the experimental analysis, when the light intensity change exceeds 12.36au, which is 3 times the standard deviation, it can be considered that the light intensity change is caused by the specific reaction of CA125 and its aptamer. Our system and method can distinguish and detect, which is why when the concentration of CA125 protein molecules is low, the light intensity measured by the system can be used as an experimental phenomenon. The detection limit of the method proposed in the present invention for CA125 is 1.58U / mL, while the clinical detection limit of CA125 is 25U / mL, which proves that our proposed method can effectively meet clinical detection needs.

[0086] 5. Specificity verification experiment

[0087] See also Figure 7 . Three interfering proteins, human IgG, AFP and BSA, were used, with alpha-fetoprotein and human immunoglobulin G concentrations of 40 ng / mL and bovine serum albumin concentration of 100 ng / mL. Except for the reactive proteins, the other experimental conditions were the same as those of the gradient experiment. The data collection and processing methods were also consistent. The bar graph results clearly show that the anti-CA125 aptamer only binds specifically to CA125. When the CA125 concentration was 40 U / mL, the corresponding average relative light intensity was 121.21 au. The result of the interaction between the interfering proteins (BSA / AFP / hIgG) and the anti-CA125 aptamer was less than 35.00 au, indicating that the adsorption of the interfering proteins could be ignored during the experiment. Through these nonspecific results, it can be found that even at high concentrations, the nonspecific binding is very small compared to the specific results. Of course, in the ultra-high sensitivity analysis of CA125, the smaller the nonspecific binding results that may have an effect, the better. Specificity verification experiments showed that our proposed aptamer sensor specifically recognized CA125, and the effect of interfering proteins on the specific binding of the anti-CA125 aptamer to CA125 was negligible.

Claims

1. An aptamer biosensor for CA125 detection, characterized in that: Including imaging weak measurement system and sensor chip; The imaging weak measurement system comprises a light source, a collimating lens, a first beam expander lens, a second beam expander lens, a front selective polarizer, a prism, a quarter wave plate, a rear selective polarizer, an imaging lens and a camera which are sequentially arranged in an optical path. The light emitted by the light source is collimated by the collimating lens, sequentially passes through the first beam expander lens and the second beam expander lens, and then passes through the front selective polarizer to enter the interior of the prism. The light beam is totally internally reflected inside the prism, and then sequentially enters the quarter wave plate, the rear selective polarizer to reach the imaging lens and the camera. The camera is used to record the image imaging of the total internal reflection interface. The sensor chip includes the prism and the detection chip. The prism and the detection chip are combined together, and a liquid flow channel is provided on the detection chip.

2. The aptamer biosensor for CA125 detection according to claim 1, characterized in that: The light source is collimated by a collimating lens to ensure that the emitted light is collimated light.

3. The aptamer biosensor for CA125 detection according to claim 1, characterized in that: The first beam expanding lens and the second beam expanding lens are a group of lenses, which are used to expand the light source on the basis of ensuring collimation, so that the light intensity of the light beam is more uniform.

4. The aptamer biosensor for CA125 detection according to claim 1, characterized in that: The angle between the front selective polarizer and the rear selective polarizer is 89° to 91°, so as to achieve polarization state adjustment of the light source.

5. The aptamer biosensor for CA125 detection according to claim 1, characterized in that: The quarter wave plate is used to convert the phase difference into optical rotation.

6. The aptamer biosensor for CA125 detection according to claim 1, characterized in that The detection chip is provided with a plurality of liquid flow channels.

7. The aptamer biosensor for CA125 detection according to claim 6, characterized in that: The multiple liquid flow channels are respectively used as experimental channels and reference channels in the experiment.

8. CA125 detection method, characterized in that, Based on the aptamer biosensor for CA125 detection according to any one of claims 1 to 6, the following operating steps are performed: S1: Solution A and solution B are sequentially introduced into the liquid flow channel of the sensor chip to obtain real-time images when different solutions are passed through. Solution A and solution B are selected from pure water, PBS phosphate buffer solution, and sodium chloride solution to be used for statistical system sensitivity: is the statistical relative light intensity change, ΔC is the concentration change, δT is the standard deviation of the system when stable, and δ is the system sensitivity; S2: Start CA125 protein molecule detection experiment: S21: depositing a binding layer on the sensor chip to prepare a first sensing interface; S22: coupling anti-CA125 aptamer molecules on the surface of the first sensing interface to prepare a second sensing interface; S23: using a protein-free blocking solution to block the non-specific binding sites on the second sensor interface, and completing a phosphate buffer solution washing step to obtain a sensor chip that can be used for detection; S3: Construct a prism sensor chip with anti-CA125 aptamer molecules as the recognition layer to achieve recognition and capture of CA125 protein molecules. By calculating the difference between the initial light intensity and the final light intensity when recognizing CA125 protein molecules, the different light intensity differences generated when capturing CA125 protein molecules of different concentrations are determined. Based on the real-time binding curve of the output light intensity △I changing with time t and the different light intensity differences, the binding rate constant Ka of the binding kinetics curve and the standard binding curve formula are obtained to analyze the characteristics of the binding reaction. When detecting CA125 protein molecules of unknown concentration, the concentration of CA125 protein molecules can be calculated based on the light intensity change and the standard binding curve formula to achieve CA125 detection.

9. The CA125 detection method according to claim 8, characterized in that: The binding layer is a dopamine membrane.

10. The CA125 detection method according to claim 8, characterized in that: The step of depositing a binding layer on the surface of the detection chip comprises: The substrate surface is cleaned with a phosphate buffer solution; dopamine is prepared with a Tris solution of pH 8.0-9.0 to obtain a dopamine-Tris solution; the liquid flow channel of the sensor chip is modified with the dopamine-Tris solution for 10-60 minutes, and then the phosphate buffer solution is used to complete the cleaning.

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