CN111351848B - Preparation method of sensor, sensor and detection method of sensor - Google Patents

Abstract

The invention belongs to the technical field of point-of-care testing, and in particular relates to a preparation method of a sensor for detecting early markers of cardiac injury, a sensor prepared by the preparation method, and a detection method of the sensor. Among them, the sensor prepared by the preparation method (due to the small size of the piezoelectric thin film resonator, only in the order of millimeters) is small in size, which is conducive to large-scale and low-cost manufacturing using semiconductor technology, and can be integrated in wearable electronic devices or In other small electronic devices; in addition, the sensor prepared by the preparation method can detect early markers of cardiac injury based on the principle of mass sensitivity, which reduces the interference of the liquid sample itself and is beneficial to improve the detection accuracy of early markers of cardiac injury; The invention also proposes a detection method based on the sensor prepared by the above preparation method, which is beneficial to realize real-time measurement of early markers of cardiac injury.

Description

A method for preparing a sensor, a sensor, and a method for detecting the sensor

technical field

The invention belongs to the technical field of point-of-care testing, and in particular relates to a preparation method of a sensor for detecting early markers of cardiac injury, a sensor prepared by the preparation method, and a detection method of the sensor.

Background technique

Cardiovascular disease is one of the most serious diseases that endanger human health and life. Among them, acute myocardial infarction is the most common and most dangerous. Early diagnosis and treatment of acute myocardial infarction is the key to reducing its mortality and improving patient outcomes. Early markers of cardiac injury are important indicators for clinical diagnosis of myocardial infarction, myocardial ischemia, heart failure and other cardiac diseases.

Early markers of cardiac injury mainly include creatine kinase MB isoenzyme (CK-MB), cardiac troponin (cTn), heart-type fatty acid-binding protein (h-FABP), and B-type natriuretic peptide (BNP). Currently, most routine laboratory tests for early markers of cardiac injury are based on chemiluminescence, enzyme-linked immunosorbent assay (ELISA), and immunoturbidimetry. While these methods provide precise, reliable, and quality-controlled results, they require complex equipment, milliliter volumes of samples, and specialized manipulation.

In recent years, the rise of point-of-care testing (POCT) technology has revolutionized the behavior of medicine. POCT equipment has short analysis time and low sample consumption, which is of great significance for the diagnosis of critical and major diseases such as acute myocardial infarction. At present, a variety of promising POCT cardiac injury early marker sensors have been developed for the detection of cardiac biomarkers, including electrochemical, magnetic, fluorescence and other principles. E.g:

Patent Document 1 discloses a biochip and a detection method for the detection of five myocardial markers, including a biosensor, a wafer platform and a spotting antibody coated on the wafer platform, and the spotting antibody can couple with magnetic beads. The combined antibody and the marker protein form an immune complex, and the concentration of the marker protein is determined by measuring the strength of the magnetoresistive signal on the complex on the biosensor.

Patent Document 2 discloses a magnetic particle microfluidic biochip and detection method for myocardial infarction and heart failure, including a PCB board with a microfluidic channel, a biosensor is arranged in the microfluidic channel, and the spotting antibody can be coupled with magnetic beads. The marker protein forms an immune complex, and the concentration of the marker protein is determined by measuring the strength of the complex magnetoresistance signal on the wafer platform.

Patent Document 3 discloses a method for preparing a label-free electrochemical sensor for cardiac troponin I and a method for detecting cTnI. A new label-free type of sensor is constructed by combining on-site preparation of electrochemically active substances with biological immune responses. The sensor specifically reacts the sensor with the target molecule (cardiac marker cTnI) to obtain the cardiac marker antigen-antibody binding layer, thereby causing electrochemical activity disturbance and regular changes in the output electrochemical signal.

Patent Document 4 discloses a preparation method of a ferrocene-based covalent organic framework modified electrode and a method for electrochemically detecting troponin, and the invention patent application provides a ferrocene with high selectivity and high detection sensitivity Preparation method of modified electrode based on covalent organic framework and its application in electrochemical sensor.

However, the above technical solutions based on the principles of electrochemistry, magnetism or fluorescence have the following defects:

(1) The size of the device is large, and it is difficult to form a miniature integrated test system; (2) The test of magnetoresistance or electrochemical detection principle is easily affected by the dielectric and magnetic properties of the liquid test sample itself, resulting in the final The test result is inaccurate.

prior art literature

Patent Literature

Patent Document 1: Publication number: CN 108845146A, publication date: November 20, 2018;

Patent Document 2: Publication number: CN 108663525A, publication date: October 16, 2018;

Patent Document 3: Publication number: CN 110161100A, publication date: August 23, 2019;

Patent Document 4: Publication number: CN 110044987A, publication date: July 23, 2019.

SUMMARY OF THE INVENTION

One of the objectives of the present invention is to provide a method for preparing a sensor for detecting early markers of cardiac injury, so as to prepare a sensor capable of detecting early markers of cardiac injury.

In order to achieve the above object, the present invention adopts the following technical solutions:

A method for preparing a sensor for detecting early markers of cardiac injury, comprising the following steps:

I. Preparation of piezoelectric thin film sensor;

II. Deposit a silicon oxide layer on the surface of the upper electrode of the piezoelectric thin film sensor;

III. Making microchannel sidewalls on the surface of the silicon oxide layer;

IV. A glass cover plate is arranged on the upper surface of the side wall of the microchannel to form a microchannel;

V. Assemble an antibody for early marker of cardiac injury on the surface of the silicon oxide layer in the microfluidic channel.

Preferably, in step I, the piezoelectric thin film sensor includes a diaphragm type piezoelectric thin film sensor, a solid mounted piezoelectric thin film sensor with an acoustic reflection layer, or an air gap structure piezoelectric thin film sensor.

Preferably, in step II, the silicon oxide layer is obtained by a method of magnetron sputtering deposition.

Preferably, in step III, the sidewall of the microchannel is made of SU8 negative glue, polydimethylsiloxane or polyimide material, and is made by ordinary photolithography, soft photolithography or nano-imprinting method; The height of the runner side wall is 1-5 mm.

Preferably, in step IV, prior to disposing the glass cover plate, the surface of the glass cover plate and the sidewall of the microchannel is treated with particles such as oxygen.

Preferably, in step V, the assembly process of the early cardiac injury marker antibody is as follows:

Firstly, deionized water and ethanol were passed into the microfluidic channel to clean the surface;

Then, the ethanol solution of aminopropyltriethoxysilane is introduced to interact with the hydroxyl group of the silicon oxide layer to form an amino surface;

Further feed into glutaraldehyde aqueous solution to carry out aldehyde group modification;

Re-pass the phosphate buffered saline solution of the early marker antibody of cardiac injury for covalent binding of the antibody;

Unbound aldehyde groups are blocked by a final passage of bovine serum albumin in phosphate buffered saline.

Preferably, in step V, the early marker antibody of cardiac injury assembled on the surface of the silicon oxide layer in the microfluidic channel comprises creatine kinase MB isoenzyme, cardiac troponin, cardiac fatty acid binding protein or B-type natriuretic peptide antibody .

The second purpose of the present invention is to provide a sensor for detecting early markers of cardiac injury, the sensor is small in size, and the sensor can effectively improve the detection accuracy of the concentration of early markers of cardiac injury.

In order to achieve the above object, the present invention adopts the following technical solutions:

A sensor for detecting early markers of cardiac injury, which is prepared by the above-mentioned preparation method of the sensor.

The third purpose of the present invention is to provide a sensor detection method for detecting early cardiac injury markers, so as to realize real-time measurement of the concentration of early cardiac injury markers.

In order to achieve the above object, the present invention adopts the following technical solutions:

A detection method of a sensor for early marker detection of cardiac injury, based on the sensor mentioned above;

The detection method of the sensor includes the following steps:

s1. Pass pure serum sample into the microfluidic channel to measure the resonance frequency of the piezoelectric thin film resonator;

Take the resonant frequency of the resonator when the resonator is in a steady state as the frequency baseline for the measurement;

s2. Passing serum standard solutions containing different concentrations of early cardiac injury markers into the microchannel in turn, and continuously measuring the resonance frequency of the piezoelectric thin film resonator under the serum standard solutions of different concentrations of early cardiac injury markers;

Through the above process, a plurality of resonant frequency variation curves with time are sequentially obtained;

s3. Take the stable value of the resonance frequency in each change curve, and then compare it with the frequency baseline respectively, and use the frequency shift value as the response of the sensor to obtain the concentration correction curve of the sensor for early markers of cardiac injury;

Among them, the frequency shift value = the frequency baseline value - the stable value of the resonance frequency;

s4. Pass the serum sample to be tested in the microfluidic channel;

s5. Continuously measure the resonant frequency of the piezoelectric thin film resonator, obtain the change curve of the resonant frequency with time, take the frequency stability value on the change curve, compare it with the frequency baseline, and use the frequency shift value as the sensor response;

s6. Comparing with the concentration calibration curve in step s3, obtain the concentration of the early cardiac injury marker corresponding to the sensor response.

Preferably, in step s2, after each time the serum standard solution of the early marker of cardiac injury with different concentrations is injected, sodium dodecyl sulfate needs to be used to restore the sensor after the antibody of the early cardiac injury marker is adsorbed.

The present invention has the following advantages:

As described above, the present invention proposes a method for preparing a sensor for detecting early markers of cardiac injury. The sensor prepared by this method (due to the small size of the piezoelectric thin film resonator, only in the order of millimeters) has a relatively small size. It is small, which is conducive to large-scale and low-cost manufacturing using semiconductor technology, and can be integrated into wearable electronic devices or other small electronic devices. The sensor prepared by the above method can detect the early markers of cardiac injury based on the principle of mass sensitivity, which reduces the interference of the liquid sample itself, and is beneficial to improve the accuracy of the detection of the early markers of cardiac injury; in addition, the present invention also proposes a method based on the above The detection method of the sensor prepared by the method is beneficial to realize the real-time measurement of the early markers of cardiac injury.

Description of drawings

FIG. 1 is a flowchart of a method for preparing a sensor for detecting early markers of cardiac injury in Example 1 of the present invention;

2 is a schematic structural diagram of a piezoelectric thin-film sensor in Embodiment 1 of the present invention;

3 is a schematic diagram of the assembly of a sensor for detecting early markers of cardiac injury in Example 1 of the present invention;

4 is a schematic structural diagram of a sensor for detecting early markers of cardiac injury in Example 2 of the present invention;

5 is a flowchart of a method for detecting a sensor for detecting early markers of cardiac injury in Embodiment 3 of the present invention;

Figure 6 is a frequency-time curve diagram of the sensor used for the detection of early cardiac injury markers in Example 3 of the present invention to pure serum and a serum standard solution of cardiac troponin (cTnI), an early cardiac injury marker;

FIG. 7 is a graph showing the concentration calibration curve of cardiac troponin (cTnI) in Example 3 of the present invention.

FIG. 8 is a schematic diagram showing the results of measurement and comparison using the detection method in Example 3 of the present invention and the conventional chemiluminescence method.

Wherein, 101-piezoelectric layer, 102-upper electrode, 103-lower electrode, 104-support layer, 105-silicon substrate, 106-silicon oxide layer, 107-microchannel sidewall, 108-glass cover plate;

109-early marker antibody of cardiac injury, 110-acoustic reflection layer, 111-air gap, 112-microfluidic channel.

Detailed ways

The present invention is described in further detail below in conjunction with the accompanying drawings and specific embodiments:

Example 1

This Example 1 describes a method for preparing a sensor for detecting early markers of cardiac injury.

As shown in Figure 1, the preparation method of the sensor includes the following steps:

I. Preparation of piezoelectric thin film sensor.

The piezoelectric thin-film sensor prepared in Example 1 is, for example, a diaphragm-type piezoelectric thin-film sensor, as shown in FIG. 2( a ).

The sensor uses an aluminum nitride film as the piezoelectric layer 101, the c-axis has an inclination angle of 24 degrees with the vertical direction, and the thickness is 1 micrometer. The upper electrode 102 is made of gold material, and the lower electrode 103 is made of tungsten material, and the thicknesses are both 100 nanometers.

The support layer 104 is a silicon nitride film with a thickness of 800 nanometers.

The silicon substrate 105 below the sonication region is completely etched to form a diaphragm structure.

Of course, the piezoelectric thin film sensor made in Example 1 can also be a solid assembled piezoelectric thin film sensor with an acoustic reflection layer, as shown in Figure 2(b), or an air gap type piezoelectric thin film sensor, as shown in Figure 2(b) 2(c).

Here, in FIG. 2( b ), reference numeral 110 denotes an acoustic reflection layer, and in FIG. 2( c ), reference numeral 111 denotes an air gap.

The following takes a diaphragm type piezoelectric thin film sensor as an example to specifically describe the preparation process of the sensor in the first embodiment.

II. Deposit a silicon oxide layer 106 on the surface of the upper electrode of the fabricated piezoelectric thin film sensor (a diaphragm type piezoelectric thin film sensor is shown in FIG. 3( a ), as shown in FIG. 3( b ) .

The silicon oxide layer 106 is obtained by the method of magnetron sputtering deposition, and the deposition thickness is 200 nanometers.

The function of the silicon oxide layer 106 is to provide a hydroxyl surface for assembling the sensitive antibody, and at the same time isolate the test liquid from the electrode.

III. Fabricating the sidewalls 107 of the microchannels on the surface of the silicon oxide layer 106, as shown in FIG. 3(c). It can be seen from FIG. 3( c ) that there are two sidewalls 107 of the microchannels prepared in Example 1, and they are both located on the surface of the silicon oxide layer 106 .

The sidewall 107 of the micro-channel is preferably made of polydimethylsiloxane (PDMS), and is fabricated by soft photolithography using common photoresist as a template. Of course, the sidewall 107 of the microchannel can also be made of SU8 negative adhesive or polyimide (PI) material.

For example, the fabrication process of the sidewall 107 of the microfluidic channel also includes ordinary photolithography or nanoimprinting methods.

The height of the side wall 107 of the microchannel is 1-5 mm, for example, it can be 2 mm.

IV. A glass cover plate 108 is arranged on the upper surface of the side wall 107 of the micro-channel to form a micro-channel, as shown in FIG. 3(d).

Before setting the glass cover plate 108, the surface of the glass cover plate and the sidewall of the microchannel needs to be treated with particles such as oxygen.

The role of the surface treatment of particles such as oxygen is to ensure the connection effect between the sidewall 107 of the microchannel and the glass cover plate 108 , to ensure the reliability of the microchannel, and to avoid leakage of the solution in the microchannel.

The effect of designing a micro-channel on the piezoelectric film sensor in Example 1 is to enable the sensor prepared by this method to perform real-time continuous measurement on the sample solution to be tested introduced into the micro-channel based on the principle of mass sensitivity, It is beneficial to ensure the accuracy of the measurement results, and at the same time, it is beneficial to change the solution, and realize the rapid measurement of the sample (no need to wait for drying).

V. Assemble the early marker antibody 109 of cardiac injury on the surface of the silicon oxide layer 106 in the microfluidic channel, as shown in FIG. 3(e).

Taking cardiac troponin as an example to illustrate the assembly process of the early marker antibody of cardiac injury, the details are as follows:

First, deionized water and ethanol are poured into the microfluidic channel 112 to clean the surface.

Then a 2% ethanol solution of aminopropyltriethoxysilane (APTES) was introduced and soaked for 60 minutes. The amino surface is formed due to the interaction of the silane groups of APTES with the hydroxyl groups of the silicon oxide layer.

A 5% aqueous solution of glutaraldehyde was further passed through for 30 minutes to modify the surface of the amino group with aldehyde groups.

Covalent binding of the antibody was carried out by infusing a 10 μg/ml solution of cardiac troponin (cTnI) in phosphate buffered saline (PBS) for 2 hours.

Unbound aldehyde groups were blocked by a final passage of 0.1% bovine serum albumin (BSA) in PBS for 30 minutes to minimize non-specific binding effects. Through the above process, the assembly process of cardiac troponin antibody is realized.

The method for assembling other early markers of cardiac injury such as creatine kinase MB isoenzyme (CK-MB), heart-type fatty acid-binding protein (h-FABP), and B-type natriuretic peptide (BNP) is the same as the above method. It is not repeated here.

A sensor for detecting early markers of cardiac injury can be prepared by the preparation method in Example 1.

Due to the small size of the sensor prepared in this embodiment 1, it is advantageous to use semiconductor technology for large-scale and low-cost manufacturing, and can be integrated into wearable electronic devices or other small electronic devices.

In addition, the sensor prepared in Example 1 can detect early markers of cardiac injury based on the principle of mass sensitivity, which is beneficial to reduce the interference of the liquid sample itself, thereby improving the detection accuracy of early markers of cardiac injury.

Example 2

This embodiment 2 describes a sensor for detecting early markers of cardiac injury, which is prepared based on the method for preparing a sensor for detecting early markers of cardiac injury in the above-mentioned embodiment 1.

As shown in FIG. 4 , the sensor includes a piezoelectric thin film sensor, a silicon oxide layer 106 , a sidewall of a microfluidic channel 107 , a glass cover plate 108 and an antibody 109 , an early marker of cardiac injury.

The silicon oxide layer 106 is located on the surface of the upper electrode 102 of the piezoelectric thin film sensor.

The sidewalls 107 of the microchannels are disposed on the surface of the silicon oxide layer 106 .

The glass cover plate 108 covers the upper surface of the microchannel sidewall 107 and is connected to the microchannel sidewall 107 to form the microchannel 112 .

The antibody 109, an early marker of cardiac injury, is assembled on the surface of the silicon oxide layer 106 in the microfluidic channel.

The cardiac injury early marker antibody 109 in this example 2 includes creatine kinase MB isoenzyme (CK-MB), cardiac troponin (cTnI), heart-type fatty acid-binding protein (h-FABP) or B-type urinary sodium Peptide (BNP) and other antibodies.

The advantages of the sensor in this embodiment 2 have been described in detail in Embodiment 1, and will not be repeated here.

In Example 2, different sensitive (ie, early cardiac injury markers) antibodies can be assembled for different sensors to form a sensor array, which can simultaneously perform joint detection of multiple early cardiac injury markers.

Example 3

This embodiment 3 describes a detection method for a sensor for detecting early markers of cardiac injury, and the detection method is implemented based on the sensor for detecting early markers for cardiac injury described in Embodiment 2 above.

The following takes cardiac troponin as an example to illustrate the detection process of early markers of cardiac injury.

As shown in Figure 5, the detection method of the sensor used for the detection of early markers of cardiac injury includes the following steps:

s1. Pass the pure serum sample into the microchannel 112, and measure the resonance frequency of the piezoelectric thin film resonator.

The inlet and outlet of the microchannel 112 use a syringe needle for input and output of liquid samples, respectively, and inject liquid samples through a syringe pump and a flow pump, and use a network analyzer or frequency measurement circuit to measure the resonator frequency.

Due to the mass and damping load, the resonator frequency of the thin film sensor drops significantly after the liquid (here refers to the pure serum sample mentioned above) is introduced, and the resonant frequency in the steady state should be taken as the frequency baseline, as shown in Figure 6 .

s2. Serum standard solutions containing different concentrations of cardiac troponin (cTnI) are sequentially introduced into the microchannel 112. Due to the reaction between antibodies and antigens, the resonant frequency of the resonator will gradually decrease and stabilize, as shown in Figure 6. Show.

The resonant frequencies of the resonators under different concentrations of serum standard solutions of early markers of cardiac injury were continuously measured.

Since each time a certain concentration of serum standard solution of cardiac troponin (cTnI) is injected, a set of frequency variation curves with time will be obtained. Therefore, multiple resonance frequency variation curves with time can be sequentially obtained through the above process.

The serum standard solution concentration of cardiac troponin (cTnI) shown in Figure 6 was 0.1 ng/ml.

After each injection of serum standard solutions of cardiac troponin (cTnI) with different concentrations, 1% sodium dodecyl sulfate (SDS) should be used to restore the sensor after antibody adsorption of early marker of cardiac injury, so that the sensor can be reuse.

Of course, this Example 3 is not limited to the use of SDS, and other liquids with the same or similar functions as SDS can also be used to restore the sensor after the adsorption of the antibody of the early marker of cardiac injury, so that the sensor can be reused.

s3. Take the stable value of the resonant frequency in each change curve, and then compare it with the frequency baseline respectively, and use the frequency shift value as the response of the sensor to obtain the concentration correction curve of the sensor for early markers of cardiac injury, as shown in Figure 7.

Among them, the squares represent the data points obtained by the actual test (the lines above and below the squares represent the standard deviation of 5 measurements), and the dashed lines represent the linear fitting of the test results. It can be seen from Fig. 7 that in the logarithmic coordinate system, the concentration of cardiac troponin (cTnI) has an approximate linear relationship with its frequency shift value. Wherein, frequency shift value=frequency baseline value-stable value of resonance frequency.

s4. Pass the serum sample to be tested in the microfluidic channel.

The serum sample to be tested in this Example 3 is a human blood sample obtained through a standard blood collection process.

s5. Continuously measure the resonant frequency of the piezoelectric thin-film resonator to obtain a change curve of the resonant frequency with time, take the frequency stability value on the change curve, compare it with the frequency baseline, and use the frequency shift value as the sensor response.

s6. Compare the concentration calibration curve in step s3 to obtain the cardiac troponin (cTnI) concentration corresponding to the sensor response.

The above process is the detection method of cardiac troponin (cTnI) concentration.

The detection process for other early markers of cardiac injury such as creatine kinase MB isoenzyme (CK-MB), heart-type fatty acid-binding protein (h-FABP), and B-type natriuretic peptide (BNP) is the same as the above method. It is not repeated here.

This example 3 is conducive to realizing real-time dynamic detection of early markers of cardiac injury, and the detection results are accurate and reliable. The specific principle is analyzed as follows: the liquid flowing in the test can wash away the substances that are not bound to the antibody on the sensor surface, thereby reducing the sensor surface. non-specific adsorption, thereby improving accuracy. In addition, the microfluidic channel has a fixed channel and cavity volume, so that the sample volume entering the sensitive area of the sensor can be precisely controlled, which improves the reliability and repeatability of the test.

Since the piezoelectric thin film resonator is fabricated by silicon semiconductor process, it can be integrated in a micro integrated test system. The dynamic measurement of flowing liquid can realize continuous and automated measurement with other components (such as blood separation, centrifugation, etc.), and after one measurement is completed , after a new liquid (such as SDS) can be introduced to clear the surface, multiple automated repeated measurements can be realized.

In addition, this Example 3 also compared the results of the detection by the method of the present invention with the results of the conventional chemiluminescence method, and the comparison results are shown in FIG. 8 . Among them, the square represents the detection result of the data point obtained by the actual test, and the dotted line represents the result of linear fitting to the test result. By comparing the results detected by the method of the present invention with the results obtained by the conventional chemiluminescence method, the results are consistent, indicating that the detection method in Example 3 of the present invention has better detection accuracy.

Of course, the above descriptions are only the preferred embodiments of the present invention, and the present invention is not limited to the above-mentioned embodiments. , and obvious deformation forms, all fall within the essential scope of this specification, and should be protected by the present invention.

Claims (8)

1. A method for detecting a sensor for early marker detection of cardiac injury, based on a sensor for early marker detection of cardiac injury, the method for preparing the sensor comprising the steps of:

I. preparing a piezoelectric film sensor;

depositing a silicon oxide layer on the surface of an upper electrode of the piezoelectric film sensor;

III, manufacturing a micro-channel side wall on the surface of the silicon oxide layer;

IV, arranging a glass cover plate on the upper surface of the side wall of the micro-channel to form the micro-channel;

v, assembling early cardiac injury marker antibodies on the surface of the silicon oxide layer in the micro-channel; it is characterized in that the preparation method is characterized in that,

the detection method of the sensor comprises the following steps:

s1, introducing a pure serum sample into the micro-channel, and measuring the resonant frequency of the piezoelectric thin-film resonator;

taking the resonance frequency of the resonator in a stable state as a measured frequency baseline;

s2, sequentially introducing serum standard solutions containing the cardiac injury early markers with different concentrations into the micro-channel, and continuously measuring the resonant frequency of the piezoelectric thin-film resonator under the serum standard solutions of the cardiac injury early markers with different concentrations;

sequentially obtaining a plurality of change curves of the resonant frequency along with time through the processes;

s3, taking the stable value of the resonance frequency in each change curve, then comparing with the frequency base line, and using the frequency shift value as the response of the sensor to obtain the concentration correction curve of the sensor to the early markers of the heart injury;

wherein frequency shift value = frequency baseline value — stable value of resonant frequency;

s4, introducing a serum sample to be detected into the micro-channel;

s5, continuously measuring the resonant frequency of the piezoelectric film resonator to obtain the change curve of the resonant frequency along with time, taking the frequency stable value on the change curve, comparing with the frequency base line, and using the frequency movement value as the sensor response;

s6 the sensor response is obtained in response to the early marker concentration of cardiac injury, against the concentration calibration curve in step s3.

2. The detection method of a sensor according to claim 1,

in the step I, the piezoelectric film sensor includes a diaphragm type piezoelectric film sensor, a solid assembly type piezoelectric film sensor having an acoustic reflection layer, or an air gap type structural piezoelectric film sensor.

3. The detection method of a sensor according to claim 1,

in the step II, the silicon oxide layer is obtained by adopting a magnetron sputtering deposition method.

4. The detection method of a sensor according to claim 1,

in the step III, the side wall of the micro-channel is made of SU8 negative glue, polydimethylsiloxane or polyimide and is manufactured by common photoetching and soft photoetching; the height of the side wall of the micro flow channel is 1-5 mm.

5. The detection method of a sensor according to claim 1,

in step IV, the glass cover plate and the surface of the side wall of the micro flow channel are treated with oxygen-containing particles before the glass cover plate is placed.

6. The detection method of a sensor according to claim 1,

in step V, the assembly process of the early heart injury marker antibody is as follows:

firstly, introducing deionized water and ethanol into the micro-channel to clean the surface;

then introducing an ethanol solution of aminopropyltriethoxysilane, and interacting with hydroxyl of the silicon oxide layer to form an amino surface;

further introducing glutaraldehyde aqueous solution for aldehyde group modification;

then introducing phosphate buffer solution of the early cardiac injury marker antibody to carry out covalent binding of the antibody;

and finally, introducing a phosphate buffer solution of bovine serum albumin to block the unbound aldehyde groups.

7. The detection method of a sensor according to claim 1,

in the step V, the early cardiac injury marker antibody assembled on the surface of the silicon oxide layer in the micro-channel comprises creatine kinase MB isozyme, cardiac troponin, cardiac fatty acid binding protein or B-type natriuretic peptide antibody.

8. The detection method of a sensor according to claim 1,

in step s2, after each time of introducing the serum standard solution of the early cardiac injury marker with different concentrations, sodium dodecyl sulfate is used to recover the sensor after the adsorption of the early cardiac injury marker antibody.

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