A method of manufacturing a flexible fiber-based biosensor

Abstract

This invention discloses a method for fabricating a flexible fiber-based biosensor, belonging to the field of biosensors. The invention aims to address the problem that existing wearable electrochemical sensors cannot achieve real-time monitoring of in vivo physiological information. The method includes: 1. Fabrication of a working electrode; 2. Fabrication of a reference electrode; 3. Fabrication of a counter electrode; 4. Assembly of the biosensor. This invention is used for the fabrication of flexible fiber-based biosensors.

Description

Technical Field

[0001] This invention belongs to the field of biosensors. Background Technology

[0002] Wearable biosensors are a type of sensor that is directly worn or installed on the surface of the human body, integrated into clothing or accessories, and performs sensing and analysis functions. Compared with traditional sensors, wearable biosensors provide a real-time, in-situ approach to effectively monitor human physiological state information, and have higher mobility, sustainability, and interactivity. They are of great significance for disease screening, early warning, diagnosis, prognosis, and health monitoring.

[0003] For monitoring human health, current wearable biosensors mostly focus on detecting physical and physiological signals such as blood pressure, blood oxygen saturation, temperature, and heart rate. These sensors provide a range of valuable information for assessing human health, but lack analysis of metabolic levels and dynamic processes at the molecular level. Some biomarkers in human body fluids are closely related to bodily states and disease development. Currently, biomarker detection is generally achieved through high-performance liquid chromatography (HPLC) or enzyme-linked immunosorbent assay (ELISA), but these methods often require complex pretreatment processes, specialized technicians, and lengthy detection times, thus limiting their application to the laboratory stage. While some optical detection methods offer high sensitivity, they generally require additional optical input and signal detection equipment, hindering their widespread adoption in wearable applications. Colorimetric analysis offers simplicity and convenience, but is susceptible to environmental interference, especially with colored samples such as urine, resulting in high false positive and false negative rates. Electrochemical sensing technology is a detection technique based on the electrochemical properties of the analyte and converting the chemical quantity of the analyte into an electrical quantity for sensing and detection. It has the characteristics of simple method, low cost, high sensitivity and fast response. It has shown great value in gas detection, body fluid analysis and other fields, and provides a feasible path for building efficient and convenient wearable sensors.

[0004] Therefore, developing high-performance wearable electrochemical sensors is an effective way to realize real-time monitoring of physiological information in vivo, providing rich and useful information for human health assessment and early screening, diagnosis and prognosis of diseases, and has significant research significance and practical value. Summary of the Invention

[0005] This invention aims to address the problem that existing wearable electrochemical sensors cannot achieve real-time monitoring of physiological information in vivo, and thus provides a method for preparing a biosensor based on flexible fibers.

[0006] A method for fabricating a biosensor based on flexible fibers, comprising the following steps:

[0007] I. Preparation of the working electrode:

[0008] ① Carbon nanotube fibers were placed in H2SO4 solution, with platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers as the working electrode. Under the conditions of voltage of -1.5V to 1.5V and scan rate of 0.05V / s to 0.20V / s, the carbon nanotube fibers were cyclically treated for 5 to 20 cycles using cyclic voltammetry to obtain acid-treated carbon nanotube fibers.

[0009] ② The acid-treated carbon nanotube fibers were placed in an electrolyte, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the acid-treated carbon nanotube fibers as the working electrode. Under the conditions of a voltage of -0.2V to 0.9V and a scan rate of 0.05V / s to 0.20V / s, the acid-treated carbon nanotube fibers were cyclically treated for 5 to 30 cycles using cyclic voltammetry to obtain the deposited carbon nanotube fibers.

[0010] The electrolyte is a mixture of ferric chloride hydrochloric acid solution, potassium chloride solution, potassium ferricyanide solution, cortisol solution, pyrrole solution, and PBS buffer. The concentrations of ferric chloride, potassium chloride, potassium ferricyanide, cortisol, HCl, and PBS buffer in the electrolyte are: 0.025 mol / L to 0.125 mol / L; 0.05 mol / L to 0.10 mol / L; 0.02 mol / L to 0.125 mol / L; 0.02 mol / L to 0.10 mol / L; 0.05 mol / L to 0.10 mol / L; and 0.5 mol / L to 2 mol / L.

[0011] ③ The deposited carbon nanotube fibers were placed in PBS buffer, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the deposited carbon nanotube fibers as the working electrode. Under the conditions of voltage of -0.2V to 0.8V and scan rate of 0.05V / s to 0.20V / s, the deposited carbon nanotube fibers were cycled for 10 to 40 cycles using cyclic voltammetry to obtain the MIP / CNT electrode.

[0012] II. Preparation of the reference electrode:

[0013] Ag / AgCl / PVB / CNT electrodes can be prepared using carbon nanotube fibers, or Ag / AgCl / cotton thread electrodes can be prepared using cotton thread and Ag / AgCl conductive paste.

[0014] III. Preparation of the counter electrode:

[0015] Using the dual-potential step method, flexible conductive fibers were plated with platinum in potassium chloroplatinate solution to obtain Pt / conductive fiber electrodes.

[0016] IV. Assembly of Biosensors:

[0017] Using Ag / AgCl / PVB / CNT or Ag / AgCl / cotton thread electrode as reference electrode, Pt / conductive fiber electrode as counter electrode, and MIP / CNT electrode as working electrode, the reference electrode, counter electrode and working electrode are assembled to obtain a biosensor based on flexible fiber.

[0018] The beneficial effects of this invention are:

[0019] 1. This invention uses cortisol as a template and pyrrole as a functional monomer to composite a molecularly imprinted polymer onto carbon nanotube fibers via electropolymerization. Since polypyrrole is an electroactive material, when cortisol enters the molecularly imprinted polymer, it can specifically bind to the binding sites in the cavity, hindering charge transport in the polypyrrole and thus causing a decrease in the current of the it curve. Therefore, cortisol can be detected using the fabricated fibrous working electrode. Furthermore, as the concentration increases, the charge transfer rate decreases, causing the current to decrease. The detection current has a linear relationship with the logarithm of the cortisol concentration, exhibiting a strong correlation, thereby enabling the detection of cortisol concentration.

[0020] 2. When the present invention is detected in lactic acid and glucose solutions, the current does not show a significant decrease, indicating that the molecularly imprinted polymer has good anti-interference properties.

[0021] 3. The biosensor prepared in this invention has a detection limit of 1 nM, a detection range of 1 nM to 1000 nM, and a sensitivity of 3.66 nA / log. 10 (C(Cortisol)), operating temperature is 22℃~37℃, response recovery time is 1min~3min.

[0022] Instruction manual illustrations

[0023] Figure 1 This is a schematic diagram illustrating the use of the flexible fiber-based biosensor of the present invention;

[0024] Figure 2 SEM image of the biosensor prepared in Example 1;

[0025] Figure 3 The it curves before and after detecting cortisol are shown for the biosensor prepared in Example 1.

[0026] Figure 4 The it curves of the biosensor prepared in Example 1 for detecting lactate (LA), glucose (Glu) and cortisol, respectively;

[0027] Figure 5 The it curves for detecting different concentrations of cortisol by the biosensor prepared in Example 1;

[0028] Figure 6 The correlation curve of cortisol concentration and current detected by the biosensor prepared in Example 1;

[0029] Figure 7 The images are scanning electron microscope images. a and d are carbon nanotube fibers with deposited Ag particles prepared in step two of Example 1, b and e are carbon nanotube fibers with deposited Ag / AgCl layers prepared in step two of Example 1, and c and f are Ag / AgCl / PVB / CNT electrodes prepared in step two of Example 1.

[0030] Figure 8 The open-circuit voltage diagrams of the Ag / AgCl / PVB / CNT electrode prepared in step two of Example 1 and the commercial Ag / AgCl electrode in potassium chloride solutions of different concentrations are shown.

[0031] Figure 9 The open-circuit voltage diagrams of the Ag / AgCl / PVB / CNT electrode prepared in step two of Example 1 and the commercial Ag / AgCl electrode in PBS solution are shown.

[0032] Figure 10 This is a scanning electron microscope image of the Pt / CNT electrode prepared in step three of Example 1;

[0033] Figure 11 The voltammetric curve of the Pt / CNT electrode prepared in step three of Example 1 in a potassium ferricyanide solution with a concentration of 0.05 mol / L is shown.

[0034] Figure 12 The images are scanning electron microscope images. a and e are untreated carbon nanotube fibers, b and f are acid-treated carbon nanotube fibers prepared in step 1① of Example 1, c and g are deposited carbon nanotube fibers prepared in step 1② of Example 1, and d and h are MIP / CNT electrodes prepared in step 1③ of Example 1.

[0035] Figure 13 The image shows the voltammetric curve of the MIP / CNT electrode prepared in step 3 of Example 1 in PBS solution. Detailed Implementation

[0036] Specific Implementation Method 1: This implementation method is a method for preparing a biosensor based on flexible fibers, which is carried out according to the following steps:

[0037] I. Preparation of the working electrode:

[0038] ① Carbon nanotube fibers were placed in H2SO4 solution, with platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers as the working electrode. Under the conditions of voltage of -1.5V to 1.5V and scan rate of 0.05V / s to 0.20V / s, the carbon nanotube fibers were cyclically treated for 5 to 20 cycles using cyclic voltammetry to obtain acid-treated carbon nanotube fibers.

[0039] ② The acid-treated carbon nanotube fibers were placed in an electrolyte, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the acid-treated carbon nanotube fibers as the working electrode. Under the conditions of a voltage of -0.2V to 0.9V and a scan rate of 0.05V / s to 0.20V / s, the acid-treated carbon nanotube fibers were cyclically treated for 5 to 30 cycles using cyclic voltammetry to obtain the deposited carbon nanotube fibers.

[0040] The electrolyte is a mixture of ferric chloride hydrochloric acid solution, potassium chloride solution, potassium ferricyanide solution, cortisol solution, pyrrole solution, and PBS buffer. The concentrations of ferric chloride, potassium chloride, potassium ferricyanide, cortisol, HCl, and PBS buffer in the electrolyte are: 0.025 mol / L to 0.125 mol / L; 0.05 mol / L to 0.10 mol / L; 0.02 mol / L to 0.125 mol / L; 0.02 mol / L to 0.10 mol / L; 0.05 mol / L to 0.10 mol / L; and 0.5 mol / L to 2 mol / L.

[0041] ③ The deposited carbon nanotube fibers were placed in PBS buffer, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the deposited carbon nanotube fibers as the working electrode. Under the conditions of voltage of -0.2V to 0.8V and scan rate of 0.05V / s to 0.20V / s, the deposited carbon nanotube fibers were cycled for 10 to 40 cycles using cyclic voltammetry to obtain the MIP / CNT electrode.

[0042] II. Preparation of the reference electrode:

[0043] Ag / AgCl / PVB / CNT electrodes can be prepared using carbon nanotube fibers, or Ag / AgCl / cotton thread electrodes can be prepared using cotton thread and Ag / AgCl conductive paste.

[0044] III. Preparation of the counter electrode:

[0045] Using the dual-potential step method, flexible conductive fibers were plated with platinum in potassium chloroplatinate solution to obtain Pt / conductive fiber electrodes.

[0046] IV. Assembly of Biosensors:

[0047] Using Ag / AgCl / PVB / CNT or Ag / AgCl / cotton thread electrode as reference electrode, Pt / conductive fiber electrode as counter electrode, and MIP / CNT electrode as working electrode, the reference electrode, counter electrode and working electrode are assembled to obtain a biosensor based on flexible fiber.

[0048] This specific embodiment is a three-electrode electrochemical biosensor consisting of a reference electrode, a counter electrode, and a working electrode, with the substrates of all three electrodes being flexible fibers; the carbon nanotube fibers are obtained by direct spinning via floating catalytic CVD.

[0049] This specific embodiment utilizes a combination of molecular imprinting (MIP) and electrochemical techniques to modify carbon nanotube fibers, enabling them to exhibit a highly specific response to analytes in sweat.

[0050] The MIP / CNT working electrode is prepared by using cyclic voltammetry to deposit conductive polymers (such as polypyrrole), redox pairs (Prussian blue), and template molecules onto the surface of carbon nanotube fibers. The template molecules are then removed by electroelution, resulting in a MIP biosensor that responds to the template molecules.

[0051] Preparation of the reference electrode (Ag / AgCl / PVB / CNT electrode): Ag particles were deposited on the CNT surface using electrochemical reduction of silver nitrate (AgNO3). The electrode was then chlorinated in hydrochloric acid solution to form an Ag / AgCl layer on the CNT surface. A PVB solution was then drop-coated onto the CNT surface, and after standing and drying, the reference electrode was obtained.

[0052] Figure 1 This is a schematic diagram illustrating the use of the flexible fiber-based biosensor of this invention. Three fibers are integrated into one to form an electrochemical sensor, which is then woven together with commercially available fibers using a pattern to construct an electrochemical sensing fabric. It is designed in various forms, such as a wristband, to meet different application needs. Due to capillary action, exocrine fluids produced by the human body are directly collected by the sensor and flow to the detection electrode. Micro-nano fabrication technology is used to construct a detection drive circuit and signal output circuit based on a flexible printed circuit board for voltage control input and signal transmission output. Channel design and element assembly are optimized to coordinate the detection performance of the multi-functional module, achieving integrated sensor control. A mobile client is designed using Java, and signal transmission is achieved via a Bluetooth transmission module, enabling real-time data sharing and analysis on the mobile device.

[0053] The beneficial effects of this embodiment are:

[0054] 1. This embodiment uses cortisol as a template and pyrrole as a functional monomer. A molecularly imprinted polymer is composited onto carbon nanotube fibers via electropolymerization. Since polypyrrole is an electroactive material, when cortisol enters the molecularly imprinted polymer, it can specifically bind to the binding sites in the cavity, hindering charge transport in the polypyrrole and thus causing a decrease in the current of the it curve. Therefore, cortisol can be detected using the fabricated fibrous working electrode. Furthermore, as the concentration increases, the charge transfer rate decreases, causing the current to decrease. The detection current has a linear relationship with the logarithm of the cortisol concentration, exhibiting a strong correlation, thereby enabling the detection of cortisol concentration.

[0055] 2. In this embodiment, when tested in lactic acid and glucose solutions, the current did not decrease significantly, indicating that the molecularly imprinted polymer has good anti-interference properties.

[0056] 3. The biosensor prepared in this embodiment has a detection limit of 1 nM, a detection range of 1 nM to 1000 nM, and a sensitivity of 3.66 nA / log. 10 (C(Cortisol)), operating temperature is 22℃~37℃, response recovery time is 1min~3min.

[0057] Specific Implementation Method Two: This implementation method differs from Specific Implementation Method One in that the concentration of the H2SO4 solution mentioned in step one ① is 0.1 mol / L to 0.5 mol / L. Everything else is the same as in Specific Implementation Method One.

[0058] Specific Implementation Method Three: This implementation method differs from Specific Implementation Method One or Two in that the PBS buffer mentioned in Step One ② and Step One ③ is a 10×PBS buffer with a pH of 7.4. Everything else is the same as in Specific Implementation Method One or Two.

[0059] Specific Implementation Method Four: This implementation method differs from Specific Implementation Methods One to Three in that: the ferric chloride hydrochloric acid solution mentioned in step one ② is prepared by mixing ferric chloride with hydrochloric acid at a concentration of 0.5 mol / L to 2.0 mol / L, and the concentration of ferric chloride in the hydrochloric acid solution is 0.025 mol / L to 0.125 mol / L; the potassium chloride solution mentioned in step one ② is prepared by mixing potassium chloride with water, and the concentration of potassium chloride in the potassium chloride solution is 0.05 mol / L to 0.10 mol / L. The potassium ferricyanide solution mentioned in step 1② is prepared by mixing potassium ferricyanide and water, and the concentration of potassium ferricyanide in the solution is 0.025 mol / L to 0.125 mol / L; the cortisol solution mentioned in step 1② is prepared by mixing cortisol and ethanol, and the concentration of cortisol in the solution is 0.02 mol / L to 0.10 mol / L; the pyrrole solution mentioned in step 1② is prepared by mixing pyrrole and water, and the concentration of pyrrole in the solution is 0.05 mol / L to 0.10 mol / L. Other aspects are the same as in specific embodiments one to three.

[0060] Specific Implementation Method Five: This implementation method differs from Specific Implementation Methods One to Four in that the flexible conductive fiber mentioned in step three is a carbon nanotube fiber, a metal wire, or a metal / cotton conductive composite. Everything else is the same as in Specific Implementation Methods One to Four.

[0061] Specific Implementation Method Six: This implementation method differs from Specific Implementation Methods One to Five in that: in step two, the Ag / AgCl / PVB / CNT electrode is prepared using carbon nanotube fibers. Specifically, it is prepared according to the following steps: Carbon nanotube fibers are placed in a KNO3 / AgNO3 solution, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the carbon nanotube fibers as the working electrode. Under conditions of a voltage of -0.9V to 0.9V and a scan rate of 0.05V / s to 0.20V / s, the carbon nanotube fibers are cyclically treated for 5 to 30 cycles using cyclic voltammetry to obtain carbon nanotube fibers with deposited Ag particles. Carbon nanotube fibers deposited with Ag particles were placed in an HCl / KCl solution. Using a platinum wire as the counter electrode and Ag / AgCl as the reference electrode, the carbon nanotube fibers deposited with Ag particles served as the working electrode. Cyclic voltammetry was used to cycle the carbon nanotube fibers deposited with Ag particles for 1 to 8 cycles at a voltage of -0.15V to 1.05V and a scan rate of 0.01V / s to 0.10V / s, resulting in carbon nanotube fibers with a deposited Ag / AgCl layer. A PVB solution was then drop-coated onto the surface of the Ag / AgCl layered carbon nanotube fibers. After standing and drying, an Ag / AgCl / PVB / CNT electrode was obtained. Other procedures are the same as in embodiments one through five.

[0062] Specific Implementation Method Seven: This implementation method differs from Specific Implementation Methods One to Six in that: the concentration of AgNO3 in the KNO3 / AgNO3 solution is 1 mmol / L to 8 mmol / L, and the concentration of KNO3 is 0.2 mol / L to 2.0 mol / L; the concentration of HCl in the HCl / KCl solution is 0.02 mol / L to 0.2 mol / L, and the concentration of KCl is 0.005 mol / L to 0.05 mol / L; the concentration of PVB in the PVB solution is 0.002 mol / L to 0.5 mol / L. Everything else is the same as in Specific Implementation Methods One to Six.

[0063] Specific Implementation Method Eight: This implementation method differs from Specific Implementation Methods One to Seven in that: in step two, the Ag / AgCl / cotton thread electrode is prepared using cotton thread and Ag / AgCl conductive paste. Specifically, it is prepared according to the following steps: the cotton thread is immersed in the Ag / AgCl conductive paste, and then rotated by a motor for 1 to 5 minutes at a speed of 2 to 20 r / min. Then, it is treated at a temperature of 40°C to 60°C for 5 to 10 minutes, and finally dried using an infrared lamp to obtain the Ag / AgCl / cotton thread electrode. The Ag / AgCl conductive paste is composed of 35 to 65 parts Ag particles, 10 to 35 parts AgCl, 20 to 40 parts polyurethane, 5 to 15 parts acrylic acid, 10 to 20 parts polyvinylpyrrolidone, and 5 to 20 parts water by mass. Everything else is the same as in Specific Implementation Methods One to Seven.

[0064] Specific Implementation Method Nine: This implementation method differs from Specific Implementation Methods One to Eight in that the Pt / conductive fiber electrode described in step three is prepared according to the following steps: using a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and flexible conductive fiber as the working electrode, the flexible conductive fiber is subjected to a step treatment in a mixed solution of KCl and K₂PtCl₆ for 5s to 20s at a voltage of 0.2V to 0.8V, and then subjected to a step treatment in a mixed solution of KCl and K₂PtCl₆ for 5s to 20s at a voltage of -0.8V to -0.1V, and the treatment is repeated 10 to 80 times; the concentration of KCl in the mixed solution of KCl and K₂PtCl₆ is 0.05mol / L to 0.20mol / L, and the concentration of K₂PtCl₆ is 0.5mmol / L to 3mmol / L. Everything else is the same as in Specific Implementation Methods One to Eight.

[0065] Specific Implementation Method Ten: This implementation method differs from Specific Implementation Methods One to Nine in that, in step four, the reference electrode, counter electrode, and working electrode are integrated into a single fiber and woven into clothing or garments. Everything else is the same as in Specific Implementation Methods One to Nine.

[0066] The beneficial effects of the present invention are verified using the following embodiments:

[0067] Example 1:

[0068] A method for fabricating a biosensor based on flexible fibers, comprising the following steps:

[0069] I. Preparation of the working electrode:

[0070] ① Carbon nanotube fibers were placed in H2SO4 solution. Platinum wire was used as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers as the working electrode. Under the conditions of voltage of -1.5V to 1.5V and scan rate of 0.05V / s, the carbon nanotube fibers were cyclically treated for 5 cycles using cyclic voltammetry to obtain acid-treated carbon nanotube fibers.

[0071] ② The acid-treated carbon nanotube fibers were placed in an electrolyte, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the acid-treated carbon nanotube fibers as the working electrode. Under the conditions of a voltage of -0.2V to 0.9V and a scan rate of 0.05V / s, the acid-treated carbon nanotube fibers were cyclically treated for 5 cycles using cyclic voltammetry to obtain the deposited carbon nanotube fibers.

[0072] The electrolyte is a mixture of ferric chloride hydrochloric acid solution, potassium chloride solution, potassium ferricyanide solution, cortisol solution, pyrrole solution, and PBS buffer. The concentrations of ferric chloride, potassium chloride, potassium ferricyanide, cortisol, pyrrole, and HCl in the electrolyte are 0.025 mol / L, 0.1 mol / L, 0.025 mol / L, 0.02 mol / L, 0.08 mol / L, and 1.0 mol / L.

[0073] ③ The deposited carbon nanotube fibers were placed in PBS buffer, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the deposited carbon nanotube fibers as the working electrode. The deposited carbon nanotube fibers were cyclically treated for 10 cycles under the conditions of a voltage of -0.2V to 0.8V and a scan rate of 0.05V / s to obtain the MIP / CNT electrode.

[0074] II. Preparation of the reference electrode:

[0075] Carbon nanotube fibers were placed in a KNO3 / AgNO3 solution. Using a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the carbon nanotube fibers as the working electrode, cyclic voltammetry was applied to the carbon nanotube fibers for 10 cycles at a voltage of -0.9V to 0.9V and a scan rate of 0.05V / s to obtain carbon nanotube fibers with deposited Ag particles. These Ag-particle-deposited carbon nanotube fibers were then placed in an HCl / KCl solution. Using a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the Ag-particle-deposited carbon nanotube fibers as the working electrode, cyclic voltammetry was applied to the fibers at a voltage of -0.15V to 1.05V and a scan rate of 0.05V / s. Carbon nanotube fibers with deposited Ag particles were cyclically treated for 4 cycles to obtain carbon nanotube fibers with a deposited Ag / AgCl layer. A PVB solution was then drop-coated onto the surface of the Ag / AgCl layered carbon nanotube fibers. After standing and drying, an Ag / AgCl / PVB / CNT electrode was obtained. The concentrations of AgNO3 and KNO3 in the KNO3 / AgNO3 solution were 2 mmol / L and 0.5 mol / L, respectively; the concentrations of HCl and KCl in the HCl / KCl solution were 0.1 mol / L and 0.02 mol / L, respectively; and the concentration of PVB in the PVB solution was 0.01 mol / L, thus obtaining the Ag / AgCl / PVB / CNT electrode.

[0076] III. Preparation of the counter electrode:

[0077] Using platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers as the working electrode, a dual-potential step method was employed. The carbon nanotube fibers were subjected to a step treatment in a mixed solution of KCl and K₂PtCl₆ for 10 s at a voltage of 0.6 V, followed by a step treatment in the same solution for 10 s at a voltage of -0.6 V. This process was repeated 20 times to obtain the Pt / CNT electrode. The concentration of KCl in the mixed solution was 0.08 mol / L, and the concentration of K₂PtCl₆ was 0.8 mmol / L.

[0078] IV. Assembly of Biosensors:

[0079] Using an Ag / AgCl / PVB / CNT electrode as the reference electrode, a Pt / CNT electrode as the counter electrode, and a MIP / CNT electrode as the working electrode, the reference electrode, counter electrode, and working electrode are integrated into a single fiber to obtain a three-electrode integrated fiber, i.e., a biosensor.

[0080] The concentration of the H2SO4 solution mentioned in step 1① is 0.2 mol / L.

[0081] The PBS buffer mentioned in steps 1.2 and 1.3 is 10×PBS buffer with a pH of 7.4.

[0082] The ferric chloride hydrochloric acid solution mentioned in step 1② is prepared by mixing ferric chloride with hydrochloric acid of concentration 1 mol / L, and the concentration of ferric chloride in the ferric chloride hydrochloric acid solution is 0.05 mol / L; the potassium chloride solution mentioned in step 1② is prepared by mixing potassium chloride with water, and the concentration of potassium chloride in the potassium chloride solution is 0.075 mol / L; the potassium ferricyanide solution mentioned in step 1② is prepared by mixing potassium ferricyanide with water, and the concentration of potassium ferricyanide in the potassium ferricyanide solution is 0.05 mol / L; the cortisol solution mentioned in step 1② is prepared by mixing cortisol with ethanol, and the concentration of cortisol in the cortisol solution is 0.05 mol / L; the pyrrole solution mentioned in step 1② is prepared by mixing pyrrole with water, and the concentration of pyrrole in the pyrrole solution is 0.075 mol / L.

[0083] The carbon nanotube fibers mentioned in Step 1①, Step 2 and Step 3 have a diameter of approximately 160 μm.

[0084] Table 1 Mechanical Performance Test Table

[0085]

[0086] The mechanical property testing standard in Table 1 is ISO 11566:1996. The test conditions are: the distance between the clamps is 25 mm, and the test speed is 1 mm / min. The tensile breaking strength unit cN / dtex in the table is referenced from Science 373, 692-696 (2021).

[0087] Figure 2 This is a SEM image of the biosensor prepared in Example 1. As shown in the image, three carbon nanotube fiber electrodes are spirally formed into one, then woven into a fabric, and together with an FPCB board and a display, constitute a flexible wearable biosensor based on fabric. The system transmits signals via Bluetooth.

[0088] Figure 3 The figure shows the it curves of the biosensor prepared in Example 1 before and after detecting cortisol. As can be seen from the figure, polypyrrole is an electroactive material. When cortisol (1 μM) enters the molecularly imprinted polymer, it hinders the charge transport of polypyrrole, and the current in the it curve decreases.

[0089] Figure 4 The it curves of the biosensor prepared in Example 1 for detecting lactic acid (LA), glucose (Glu) and cortisol are shown in the figure. As can be seen from the figure, the current did not decrease significantly when detected in 5mM lactic acid and 50μM glucose solutions, indicating that the molecularly imprinted polymer has good anti-interference properties.

[0090] The biosensor prepared in Example 1 was used to detect different concentrations of cortisol under operating conditions of 22℃~37℃ and a response recovery time of 3 min (response time 1 min, recovery time 2 min). Figure 5 The figure shows the it curves for detecting different concentrations of cortisol using the biosensor prepared in Example 1. As the concentration increases, the charge transfer rate decreases, leading to a decrease in current, thus enabling the detection of cortisol concentration. Furthermore, the figure shows that the biosensor prepared in Example 1 has a detection limit of 1 nM, a detection range of 1 nM to 1000 nM, and a sensitivity of 3.66 nA / log. 10 (C(Cortisol)).

[0091] Figure 6 The image shows the correlation curve between cortisol concentration and current detected by the biosensor prepared in Example 1. Figure 5 The results were fitted, and the resulting equation was y = -3.66x + 36.82 (where y represents the current and x represents the logarithm of the cortisol concentration), and the regression coefficient R0 was [value missing]. 2 The value is 0.9921. The fitting results show that there is a linear relationship between the detection current and the logarithm of the cortisol concentration, indicating a strong correlation.

[0092] Figure 7 The images are scanning electron microscope (SEM) images. a and d are carbon nanotube fibers with deposited Ag particles prepared in step two of Example 1; b and e are carbon nanotube fibers with deposited Ag / AgCl layers prepared in step two of Example 1; c and f are Ag / AgCl / PVB / CNT electrodes prepared in step two of Example 1. As can be seen from the images, Ag nanoparticles were first deposited on the surface of the carbon nanotube fibers, then chlorinated to form an Ag / AgCl structure, and finally a PVB layer was coated on the surface of the chlorinated carbon nanotube fibers.

[0093] Figure 8 The open-circuit voltage diagrams of the Ag / AgCl / PVB / CNT electrode prepared in step two of Example 1 and the commercial Ag / AgCl electrode in potassium chloride solutions of different concentrations are shown. Figure 9 The figure shows the open-circuit voltage of the Ag / AgCl / PVB / CNT electrode prepared in step two of Example 1 and the commercial Ag / AgCl electrode in PBS solution. As can be seen from the figure, the open-circuit voltage potential deviation between the Ag / AgCl / PVB / CNT electrode prepared in step two of Example 1 and the commercial Ag / AgCl electrode in 10mM, 100mM, 1000mM KCl and PBS solution (10×PBS buffer, pH 7.4) is close to zero, indicating that the electrode has good test stability.

[0094] Figure 10The image shows a scanning electron microscope (SEM) image of the Pt / CNT electrode prepared in step three of Example 1. As can be seen from the image, Pt nanoparticles are uniformly deposited on the surface of the carbon nanotube fibers.

[0095] Figure 11 The image shows the voltammetric curve of the Pt / CNT electrode prepared in step three of Example 1 in a 0.05 mol / L potassium ferricyanide solution. The voltammetric curves of the prepared Pt / CNT electrode almost overlap with those of the commercial Pt wire electrode, indicating that this electrode can replace the commercial Pt wire electrode.

[0096] Figure 12 The images are scanning electron microscope (SEM) images. A and E represent untreated carbon nanotube fibers, B and F represent acid-treated carbon nanotube fibers prepared in step ① of Example 1, C and G represent deposited carbon nanotube fibers prepared in step ② of Example 1, and D and H represent MIP / CNT electrodes prepared in step ③ of Example 1. As shown in Figure f, pores appear in the acid-treated carbon nanotube fibers. As shown in Figure g, electrodeposited particles adhere to the carbon nanotube fibers. As shown in Figure h, the template molecule cortisol was successfully removed after electroelution.

[0097] The MIP / CNT electrode prepared in step one of Example 1 was tested: the electrode was placed in 10×PBS solution for CV detection, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the MIP / CNT electrode as the working electrode. Under conditions of -1.2V to 0.8V and a scan rate of 0.05V / s, the electrode was cycled for 5 times, resulting in a stable redox peak. The test results are as follows: Figure 13 As shown, Figure 13 The figure shows the voltammetric curve of the MIP / CNT electrode prepared in step 1③ of Example 1 in PBS solution; as can be seen from the figure, oxidation and reduction peaks appear between 0.2 and 0.4 V. Figure 12 This indicates that the MIP / CNT electrode was successfully fabricated.

Claims

1. A method for the preparation of a flexible fiber-based biosensor, characterized in that It is done in the following steps: I. Preparation of the working electrode: ① Carbon nanotube fibers were placed in H2SO4 solution, with platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers as the working electrode. Under the conditions of voltage of -1.5V~1.5V and scan rate of 0.05V / s~0.20V / s, the carbon nanotube fibers were cyclically treated for 5~20 cycles using cyclic voltammetry to obtain acid-treated carbon nanotube fibers. The carbon nanotube fibers mentioned above are obtained by direct spinning using floating catalytic CVD. ② The acid-treated carbon nanotube fibers were placed in an electrolyte, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the acid-treated carbon nanotube fibers as the working electrode. Under the conditions of a voltage of -0.2V to 0.9V and a scan rate of 0.05V / s to 0.20V / s, the acid-treated carbon nanotube fibers were cyclically treated for 5 to 30 cycles using cyclic voltammetry to obtain the deposited carbon nanotube fibers. The electrolyte is a mixture of ferric chloride hydrochloric acid solution, potassium chloride solution, potassium ferricyanide solution, cortisol solution, pyrrole solution, and PBS buffer. The concentrations of ferric chloride, potassium chloride, potassium ferricyanide, cortisol, HCl, and PBS buffer in the electrolyte are: 0.025 mol / L to 0.125 mol / L; 0.05 mol / L to 0.10 mol / L; 0.02 mol / L to 0.125 mol / L; 0.02 mol / L to 0.10 mol / L; 0.05 mol / L to 0.10 mol / L; and 0.5 mol / L to 2 mol / L. ③ The deposited carbon nanotube fibers were placed in PBS buffer, with a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and the deposited carbon nanotube fibers as the working electrode. Under the conditions of a voltage of -0.2V to 0.8V and a scan rate of 0.05V / s to 0.20V / s, the deposited carbon nanotube fibers were cycled for 10 to 40 cycles using cyclic voltammetry to obtain the MIP / CNT electrode. II. Preparation of the reference electrode: Ag / AgCl / PVB / CNT electrodes can be prepared using carbon nanotube fibers, or Ag / AgCl / cotton thread electrodes can be prepared using cotton thread and Ag / AgCl conductive paste. III. Preparation of the counter electrode: Using the dual-potential step method, flexible conductive fibers were plated with platinum in potassium chloroplatinate solution to obtain Pt / conductive fiber electrodes. IV. Assembly of Biosensors: Using Ag / AgCl / PVB / CNT or Ag / AgCl / cotton thread electrode as the reference electrode, Pt / conductive fiber electrode as the counter electrode, and MIP / CNT electrode as the working electrode, the reference electrode, counter electrode, and working electrode are integrated into a single fiber to obtain a flexible fiber-based biosensor.

2. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... The concentration of the H2SO4 solution mentioned in step 1① is 0.1mol / L~0.5mol / L.

3. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... The PBS buffer mentioned in steps 1.2 and 1.3 is 10×PBS buffer with a pH of 7.

4.

4. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... The ferric chloride hydrochloric acid solution mentioned in step 1② is prepared by mixing ferric chloride with hydrochloric acid at a concentration of 0.5 mol / L to 2.0 mol / L, and the concentration of ferric chloride in the hydrochloric acid solution is 0.025 mol / L to 0.125 mol / L; the potassium chloride solution mentioned in step 1② is prepared by mixing potassium chloride with water, and the concentration of potassium chloride in the potassium chloride solution is 0.05 mol / L to 0.10 mol / L; the potassium ferricyanide solution mentioned in step 1②... The solution is composed of potassium ferricyanide and water, with the concentration of potassium ferricyanide in the solution being 0.025 mol / L to 0.125 mol / L; the cortisol solution mentioned in step 1② is composed of cortisol and ethanol, with the concentration of cortisol in the solution being 0.02 mol / L to 0.10 mol / L; the pyrrole solution mentioned in step 1② is composed of pyrrole and water, with the concentration of pyrrole in the solution being 0.05 mol / L to 0.10 mol / L.

5. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... The flexible conductive fiber mentioned in step three is a carbon nanotube fiber, a metal wire, or a metal / cotton conductive composite.

6. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... Step two involves preparing an Ag / AgCl / PVB / CNT electrode using carbon nanotube fibers. Specifically, the preparation process includes the following steps: Carbon nanotube fibers are placed in a KNO3 / AgNO3 solution. A platinum wire is used as the counter electrode, Ag / AgCl as the reference electrode, and the carbon nanotube fibers as the working electrode. Under conditions of -0.9V to 0.9V and a scan rate of 0.05V / s to 0.20V / s, the carbon nanotube fibers are cyclically treated for 5 to 30 cycles using cyclic voltammetry to obtain carbon nanotube fibers with deposited Ag particles. The carbon nanotube fibers with deposited Ag particles are then placed in an H2O solution. In a Cl / KCl solution, using a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and carbon nanotube fibers deposited with Ag particles as the working electrode, the carbon nanotube fibers deposited with Ag particles were cyclically treated for 1 to 8 cycles using a voltage of -0.15V to 1.05V and a scan rate of 0.01V / s to 0.10V / s to obtain carbon nanotube fibers with deposited Ag / AgCl layers. PVB solution was then drop-coated onto the surface of the carbon nanotube fibers with deposited Ag / AgCl layers, and after standing and drying, an Ag / AgCl / PVB / CNT electrode was obtained.

7. A method of making a flexible fiber-based biosensor according to claim 6, wherein The concentration of AgNO3 in the KNO3 / AgNO3 solution is 1 mmol / L to 8 mmol / L, and the concentration of KNO3 is 0.2 mol / L to 2.0 mol / L; the concentration of HCl in the HCl / KCl solution is 0.02 mol / L to 0.2 mol / L, and the concentration of KCl is 0.005 mol / L to 0.05 mol / L; the concentration of PVB in the PVB solution is 0.002 mol / L to 0.5 mol / L.

8. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... In step two, an Ag / AgCl / cotton thread electrode is prepared using cotton thread and Ag / AgCl conductive paste. Specifically, the preparation is carried out according to the following steps: the cotton thread is immersed in the Ag / AgCl conductive paste, and then the immersed cotton thread is rotated for 1 min to 5 min at a speed of 2 r / min to 20 r / min using a motor. Then, it is treated at a temperature of 40℃ to 60℃ for 5 min to 10 min, and then dried using an infrared lamp to obtain the Ag / AgCl / cotton thread electrode. The Ag / AgCl conductive paste is composed of 35 parts to 65 parts Ag particles, 10 parts to 35 parts AgCl, 20 parts to 40 parts polyurethane, 5 parts to 15 parts acrylic acid, 10 parts to 20 parts polyvinylpyrrolidone, and 5 parts to 20 parts water by mass.

9. The method for fabricating a flexible fiber-based biosensor according to claim 1, characterized in that... The Pt / conductive fiber electrode described in step three is prepared according to the following steps: using a platinum wire as the counter electrode, Ag / AgCl as the reference electrode, and a flexible conductive fiber as the working electrode, the flexible conductive fiber is subjected to a step treatment in a mixed solution of KCl and K2PtCl6 for 5s to 20s under a voltage of 0.2V to 0.8V. Then, the flexible conductive fiber is subjected to a step treatment in a mixed solution of KCl and K2PtCl6 for 5s to 20s under a voltage of -0.8V to -0.1V, and the treatment is repeated 10 to 80 times. The concentration of KCl in the mixed solution of KCl and K2PtCl6 is 0.05mol / L to 0.20mol / L, and the concentration of K2PtCl6 is 0.5mmol / L to 3mmol / L.

10. The method of claim 1, wherein In step four, the reference electrode, counter electrode, and working electrode are integrated into a single fiber and woven into clothing or garments.

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