A p-type organic electrochemical transistor biosensor and preparation method and application

Abstract

The application belongs to the technical field of organic electrochemical transistors, and discloses a p-type organic electrochemical transistor biosensor, a preparation method and application. The sensor comprises a first substrate and a second substrate. A source electrode is arranged on one side of the first substrate. A polymer semiconductor channel is arranged on the source electrode. A drain electrode is arranged on the polymer semiconductor channel. A first floating gate is arranged on the other side of the first substrate. The first substrate, the drain electrode and the first floating gate are covered with a first electrolyte layer. A gate electrode is arranged on one side of the second substrate. A gate modification layer is arranged on the gate electrode. A second floating gate is arranged on the other side of the second substrate. The second substrate, the gate modification layer and the second floating gate are covered with a second electrolyte layer. The first floating gate and the second floating gate are connected through a wire. The application realizes the detection of an ultralow concentration (10 ‑20 mol / L) H1N1 virus marker and realizes the detection of 10 ‑17 mol / L H1N1 virus marker in an ultrashort time.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of organic electrochemical transistors, and particularly relates to a p-type organic electrochemical transistor biosensor and a preparation method and application thereof. BACKGROUND

[0002] With the improvement of social living standards and the continuous progress of medical technology, people pay more and more attention to health. Real-time detection of biomarkers has become an effective means to obtain information about individual health status, disease development and treatment effect. Pathogen markers play an important role in life, and changes in their expression levels often reflect the situation of physiological or pathological changes in the body. By monitoring the changes in pathogen levels in body fluids, medical personnel can more accurately assess the disease state and develop personalized treatment plans to provide more accurate medical services for patients. This application based on pathogen markers brings new possibilities for disease management and health monitoring, and helps to improve the level of medical diagnosis and treatment, and provides more comprehensive attention and care for individual health.

[0003] In recent years, with the development of organic semiconductor materials and the optimization of processing technology, high-performance organic electrochemical transistors (OECT) have been widely studied and rapidly developed. Due to their excellent working stability in liquid phase, signal conversion and amplification function, and good biocompatibility, OECT-based biosensors have shown great potential in rapid and sensitive biomarker detection applications. However, pathogen biomarkers have low intrinsic content, and there are a large number of interfering substances such as ions and molecules in human body fluids (blood, sweat, saliva, etc.), so it is urgent to develop a biosensor with high stability, strong anti-interference ability and high sensitivity. SUMMARY

[0004] In view of the above problems in the prior art, the purpose of the present application is to provide a p-type organic electrochemical transistor biosensor and a preparation method and application thereof. The p-type organic electrochemical transistor sensor can work under positive voltage, avoiding the problem of small and unstable voltage modulation range of the device under negative voltage, and has the advantages of rapid, sensitive and stable detection.

[0005] In order to achieve the above purpose, the technical scheme adopted by the present application is as follows:

[0006] A p-type organic electrochemical transistor biosensor, comprising a first substrate and a second substrate.

[0007] The first substrate is provided with a source electrode on one side, a polymer semiconductor channel on the source electrode, and a drain electrode on the polymer semiconductor channel; the first substrate is provided with a first floating gate on the other side, and the first substrate, the drain electrode and the first floating gate are provided with a first electrolyte layer;

[0008] The second substrate is provided with a gate electrode on one side, a gate electrode modification layer on the gate electrode, and a second floating gate on the other side; the second substrate, the gate electrode modification layer and the second floating gate are provided with a second electrolyte layer; the first floating gate and the second floating gate are connected by a wire;

[0009] A drain voltage is applied in a loop composed of the source electrode, the polymer semiconductor channel and the drain electrode, and the source electrode is grounded;

[0010] A gate voltage is applied in a loop composed of the source electrode, the first floating gate, the second floating gate, the wire, the first electrolyte layer, the second electrolyte layer, the gate electrode and the gate electrode modification layer;

[0011] The gate electrode modification layer is a gate electrode modification layer containing a target probe molecule layer of a target to-be-measured substance.

[0012] Further, the materials of the first substrate and the second substrate are silicon oxide;

[0013] The materials of the source electrode, the drain electrode and the gate electrode are Au.

[0014] Further, the first electrolyte layer is prepared from a NaCl aqueous solution, and the second electrolyte layer is prepared from a PBS buffer solution.

[0015] Further, the material of the polymer semiconductor channel is poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene];

[0016] The first floating gate and the second floating gate are prepared from Ag / AgCl paste.

[0017] Further, the thickness of the source electrode is 60-70 nm; the thickness of the drain electrode is 60-70 nm; the thickness of the polymer semiconductor channel is 130-150 nm; the thickness of the gate electrode is 60-70 nm; the thickness of the gate electrode modification layer is 5-10 nm; and the thickness of the first floating gate and the second floating gate is 5-10 μm.

[0018] A preparation method of a p-type organic electrochemical transistor biosensor, comprising the following steps:

[0019] Depositing Cr on the first substrate to form a Cr layer, and then depositing Au on the Cr layer to form a source electrode; and depositing Cr on the second substrate to form a Cr layer, and then depositing Au on the Cr layer to form a gate electrode;

[0020] A chloroform solution of poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] is spin-coated on the source electrode surface to obtain a polymer semiconductor channel;

[0021] Au is deposited on the surface of the polymer semiconductor channel to form a drain electrode;

[0022] A first floating gate is arranged on the first substrate, and a second floating gate is arranged on the second substrate;

[0023] A PBS buffer solution containing a target analyte probe is applied to the surface of the gate electrode, and is left to stand to form a target analyte probe molecule layer; a 6-mercapto-1-hexanol solution is added dropwise to the surface of the gate electrode to cover the surface of the gate electrode that is not covered by the target analyte probe molecule layer, and is left to stand to form a 6-mercapto-1-hexanol blocking layer, and the target analyte probe molecule layer and the 6-mercapto-1-hexanol blocking layer form a gate modification layer.

[0024] A first electrolyte layer is arranged on the first substrate, the drain electrode and the first floating gate, and a second electrolyte layer is arranged on the second substrate, the gate modification layer and the second floating gate, and the first floating gate and the second floating gate are connected by a wire to obtain a p-type organic electrochemical transistor biosensor.

[0025] Further, the first floating gate is arranged on the first substrate, and the second floating gate is arranged on the second substrate, and the method comprises the following steps:

[0026] Ag / AgCl paste is applied to the first substrate and the second substrate, and then annealing is performed to form the first floating gate on the first substrate and the second floating gate on the second substrate.

[0027] Further, the annealing is performed under an inert gas, and the temperature of the annealing is 100 DEG C, and the time of the annealing is 30 min.

[0028] A p-type organic electrochemical transistor biosensor in the detection of biomarkers.

[0029] Compared with the prior art, the present application has at least the following beneficial effects:

[0030] To meet the high sensitivity and high selectivity detection requirements of the OECT sensor, the application utilizes the interaction between the target to be measured and the probe molecules to modulate the electrical properties of the device, and realizes high sensitivity sensing response. The setting of the first floating gate and the second floating gate enables the p-type organic electrochemical transistor biosensor to always operate under optimal conditions and not to be contaminated by the to-be-measured substance, and can be repeatedly used, thereby reducing the test cost. Experimental results show that the p-type organic electrochemical transistor biosensor has good cycle operation stability and sensitivity to voltage changes, and can complete the detection of 10 amol / L (10 -17 mol / L) H1N1 virus markers within 5 minutes, the minimum detection limit is as low as 10 zmol / L (10 - 20 mol / L) level, and the current response has a good linear change relationship with the detection concentration, showing the advantages of high sensitivity and rapid detection, and showing extraordinary potential in the field of instant on-site diagnosis.

[0031] Further, the application selects poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] as the channel material, and selects a vertical device structure, so that the prepared p-type organic electrochemical transistor biosensor has an ultra-high transconductance value and signal amplification capability. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 is a structure schematic diagram of a p-type organic electrochemical transistor biosensor of the application;

[0033] Figure 2 is a current change graph of OECT long-time cycle test when Au is used as the floating gate material in embodiment 1 of the application;

[0034] Figure 3 is a current change graph of OECT long-time cycle test when Ag / AgCl is used as the floating gate material in embodiment 1 of the application;

[0035] Figure 4 is a graph of reading out the actual gate voltage through the second floating gate when a positive gate voltage (0~+0.6 V) is applied in embodiment 1 of the application;

[0036] Figure 5 is a graph of reading out the actual gate voltage through the second floating gate when a negative gate voltage (0~-0.6 V) is applied in embodiment 1 of the application;

[0037] Figure 6 is a graph of reading out the voltage of the second floating gate when the gate voltage V g = 0 V in the feasibility verification of the effective gate voltage test method in embodiment 1 of the application;

[0038] Figure 7 The gate voltage is applied when the feasibility of the effective gate voltage test method in Example 1 of the present invention is verified. V g = 0.3 V when the voltage reading of the second floating gate is;

[0039] Figure 8 Initial setting of gate voltage V g = 0.3 V, the voltage readout value of the second floating gate and the polymer semiconductor channel current value;

[0040] Figure 9 The graph shows the voltage readout of the second floating gate and the polymer semiconductor channel current value after the device was placed in air for 1 hour under the same setting conditions;

[0041] Figure 10 To set the gate voltage V g = 0 V, the voltage readout value of the second floating gate and the polymer semiconductor channel current value;

[0042] Figure 11 To set the gate voltage V g = 0.05 V, the voltage readout of the second floating gate and the polymer semiconductor channel current value;

[0043] Figure 12 Schematic diagram of the gate of the p-type organic electrochemical transistor biosensor with a gate modification layer in Example 1 of the present invention;

[0044] Figure 13 Schematic diagram of the gate after the gate modification layer is specifically combined with the H1N1 virus marker;

[0045] Figure 14 for Figure 12 Schematic diagram of ion doping in the polymer semiconductor channel under the conditions shown in ;

[0046] Figure 15 for Figure 13 Schematic diagram of ion doping in the polymer semiconductor channel under the conditions shown in ;

[0047] Figure 16 To set the gate voltage for the p-type organic electrochemical transistor biosensor in Example 2 of the present invention V g = 0.3 V, the voltage readout value of the second floating gate and the polymer semiconductor channel current value;

[0048] Figure 17The second floating gate voltage readout and the polymer semiconductor channel current were plotted after the gate of the modified target probe molecule was incubated in PBS buffer for 1 hour while maintaining the original electrical test parameter settings.

[0049] Figure 18 The gate of the modified target probe molecule is at a concentration of 10 -20 After incubation in 1 mol / L H1N1 PBS buffer for 1 h, the original electrical test parameter settings were maintained, and the second floating gate voltage readout and polymer semiconductor channel current value were plotted;

[0050] Figure 19 for Figure 16 Transfer curve test data of the device module before the effective gate voltage test;

[0051] Figure 20 for Figure 17 Transfer curve test data of the device module before the effective gate voltage test;

[0052] Figure 21 for Figure 18 Transfer curve test data of the device module before the effective gate voltage test;

[0053] Figure 22 for Figure 16 The baseline current obtained from Figure 17 The noise current obtained from Figure 18 Schematic diagram of the comparison of the binding response current obtained in;

[0054] Figure 23 To set the gate voltage for the p-type organic electrochemical transistor biosensor in Example 2 of the present invention V g = 0.3 V, the voltage readout value of the second floating gate and the polymer semiconductor channel current value;

[0055] Figure 24 This is a graph showing the second floating gate voltage readout and the polymer semiconductor channel current value obtained after the gate of the modified targeting probe molecule in Example 2 was incubated in PBS buffer for 5 minutes while maintaining the original electrical test parameter settings;

[0056] Figure 25 The gate of the modified target probe molecule in Example 2 is at a concentration of 10 -17 After incubation in 10 mol / L H1N1 PBS buffer for 5 min, the original electrical test parameter settings were maintained, and the second floating gate voltage readout and polymer semiconductor channel current value were plotted.

[0057] Figure 26 for Figure 23The transfer curve test data graph of the device module before the effective gate voltage test;

[0058] Figure 27 For Figure 24 The transfer curve test data graph of the device module before the effective gate voltage test;

[0059] Figure 28 For Figure 25 The transfer curve test data graph of the device module before the effective gate voltage test;

[0060] Figure 29 For Figure 23 The baseline current derived in the middle, Figure 24 The noise current derived in the middle, Figure 25 The combined response current size comparison schematic diagram derived in the middle;

[0061] Figure 30 The current response change fitting curve derived in the H1N1 virus detection in the embodiment 2 of the application, after the target probe molecule is combined with the target to be detected for 5 min, for different concentrations of target to be detected;

[0062] Figure 31 The gate voltage of the p-type organic electrochemical transistor biosensor in the embodiment 4 of the application V g The voltage readout value of the second floating gate and the polymer semiconductor channel current value when = 0.3 V;

[0063] Figure 32 The second floating gate voltage readout value and the polymer semiconductor channel current value derived after the gate of the modified target probe molecule is incubated in PBS buffer for 5 min, and the original electrical test parameter setting is maintained;

[0064] Figure 33 The second floating gate voltage readout value and the polymer semiconductor channel current value derived after the gate of the modified target probe molecule is incubated in 10 -17 mol / L H1N1 PBS buffer for 5 min, and the original electrical test parameter setting is maintained;

[0065] Figure 34 For Figure 31 The transfer curve test data graph of the device module before the effective gate voltage test;

[0066] Figure 35 For Figure 32 The transfer curve test data graph of the device module before the effective gate voltage test;

[0067] Figure 36 For Figure 33Transfer curve test data of the device module before the effective gate voltage test;

[0068] Figure 37 for Figure 31 The baseline current obtained from Figure 32 The noise current obtained from Figure 33 Schematic diagram of the comparison of the binding response current obtained in;

[0069] Figure 38 To set the gate voltage for the p-type organic electrochemical transistor biosensor in Example 6 of the present invention V g = 0.3 V, the voltage readout value of the second floating gate and the polymer semiconductor channel current value;

[0070] Figure 39 The second floating gate voltage readout value and the polymer semiconductor channel current value are plotted after the gate of the modified targeting probe molecule of Example 6 is incubated in PBS buffer for 5 minutes while maintaining the original electrical test parameter settings;

[0071] Figure 40 The gate of the modified targeting probe molecule of Example 6 is at a concentration of 10 -17 After incubation in 10 mol / L H1N1 PBS buffer for 5 min, the original electrical test parameter settings were maintained, and the second floating gate voltage readout and polymer semiconductor channel current value were plotted.

[0072] Figure 41 for Figure 38 Transfer curve test data of the device module before the effective gate voltage test;

[0073] Figure 42 for Figure 39 Transfer curve test data of the device module before the effective gate voltage test;

[0074] Figure 43 for Figure 40 Transfer curve test data of the device module before the effective gate voltage test;

[0075] Figure 44 for Figure 38 The baseline current obtained from Figure 39 The noise current obtained from Figure 40 Schematic diagram of the comparison of the binding response current obtained in;

[0076] In the figure, 1 is the first substrate, 2 is the source, 3 is the polymer semiconductor channel, 4 is the drain, 5 is the gate, 6 is the first electrolyte layer, 7 is the second electrolyte layer, 8 is the gate modification layer, 9 is the first floating gate, 10 is the wire, 11 is the second substrate, and 12 is the second floating gate. DETAILED DESCRIPTION

[0077] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments.

[0078] The PBS buffer solution in the present invention is a standard PBS buffer solution with a pH of 7.2-7.4 and a concentration of 10 mmol / L.

[0079] See also Figure 1 A p-type organic electrochemical transistor biosensor of the present invention includes a first substrate 1 and a second substrate 11; a source electrode 2 is arranged on one side of the first substrate 1, a polymer semiconductor channel 3 is arranged on the source electrode 2, a drain electrode 4 is arranged on the polymer semiconductor channel 3, and a first floating gate 9 is arranged on the other side of the first substrate 1, the source electrode 2 and the first floating gate 9 are arranged at intervals, and a first electrolyte layer 6 is covered on the first substrate 1, the drain electrode 4 and the first floating gate 9; a gate electrode 5 is arranged on one side of the second substrate 11, a gate modification layer 8 is arranged on the gate electrode 5, and a second floating gate 12 is arranged on the other side of the second substrate 11, the gate 5 and the second floating gate 12 are arranged at intervals, and a second electrolyte layer 7 is covered on the second substrate 11, the gate modification layer 8 and the second floating gate 12; the first floating gate 9 and the second floating gate 12 are connected by a wire 10.

[0080] The first substrate 1 and the second substrate 11 are made of silicon oxide.

[0081] The source electrode 2 , the drain electrode 4 and the gate electrode 5 are made of Au.

[0082] The first electrolyte layer 6 is a NaCl aqueous solution, and the second electrolyte layer 7 is a PBS buffer solution.

[0083] The material of the polymer semiconductor channel 3 is poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] (CAS No. 1974310-62-7, abbreviated as P(g2T-T)).

[0084] The gate modification layer 8 is a target probe molecule layer of the target analyte randomly and staggeredly bonded to the surface of the gate 5 and a 6-mercapto-1-hexanol blocking layer.

[0085] The first floating gate 9 and the second floating gate 12 are electrodes made of Ag / AgCl slurry.

[0086] The wire 10 is a copper wire.

[0087] The length of the source electrode 2 is 250-300 μm, the width is 95-105 μm, and the thickness is 60-70 nm; the length of the drain electrode 4 is 1200-1400 μm, the width is 95-105 μm, and the thickness is 60-70 nm; the length of the polymer semiconductor channel 3 is 100-110 μm, the width is 100-110 μm, and the thickness is 130-150 nm; the length of the gate electrode 5 is 9-10 mm, the width is 5-6 mm, and the thickness is 60-70 nm; the length of the gate modification layer 8 is 9-10 mm, the width is 5-6 mm, and the thickness is 5-10 nm; the length of the first floating gate 9 and the second floating gate 12 is 11-13 mm, the width is 6-7 mm, and the thickness is 5-10 μm.

[0088] The preparation method of the p-type organic electrochemical transistor biosensor as described above comprises the following steps:

[0089] A Cr layer is formed on the clean first substrate 1 by mask vacuum thermal evaporation deposition of metal Cr at a deposition rate of 0.2 Å / s to improve the adhesion of Au to the first substrate 1, and then an Au layer is formed on the Cr layer by mask vacuum thermal evaporation deposition of metal Au at a deposition rate of 1 Å / s-1.5 Å / s to form the source electrode 2. A Cr layer is formed on the clean second substrate 11 by mask vacuum thermal evaporation deposition of metal Cr at a deposition rate of 0.2 Å / s to improve the adhesion of Au to the second substrate 11, and then an Au layer is formed on the Cr layer by mask vacuum thermal evaporation deposition of metal Au at a deposition rate of 1 Å / s-1.5 Å / s to form the gate electrode 5.

[0090] A chloroform solution of poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] (molecular weight 20000) is spin-coated on the surface of the source electrode 2 to form the polymer semiconductor channel 3; the concentration of the chloroform solution of poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] is 20 mg / ml, the spin-coating speed is 5000 r / min, the spin-coating time is 25-30 s, the volume is 25 μL, and the thickness of the polymer semiconductor channel 3 is 130-150 nm.

[0091] An Au layer is formed on the surface of the polymer semiconductor channel 3 by mask vacuum thermal evaporation deposition of metal Au at a deposition rate of 1 Å / s-1.5 Å / s to form the drain electrode 4.

[0092] The poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] on the first substrate 1 except the polymer semiconductor channel 3 between the source electrode 2 and the drain electrode 4 is removed by a reactive ion etching method; specifically, etching is performed using the drain electrode 4 as a mask plate, the etching gas is argon, the flow rate is 15 sccm, the etching power is 25 W, and the etching time is 250 s.

[0093] The Ag / AgCl slurry (the manufacturer of the Ag / AgCl slurry is Shanghai Julong Electronics Technology Co., Ltd., and the product name is JLL20) is coated on the first substrate 1 and the second substrate 11 by using a mask plate; the coated first substrate 1 and the second substrate 11 are heated at 100 ℃ for 30 min in an inert gas environment for annealing, so as to obtain the first floating gate 9 and the second floating gate 12, the first floating gate 9 is located at one end of the first substrate 1, and the second floating gate 12 is located at the other end of the second substrate 11.

[0094] The gate electrode 5 is surrounded by a PDMS (Polydimethylsiloxane) groove, PBS (Phosphate Buffered Saline) buffer solution added with a target probe molecule (in the present application, the target probe molecule of the target analyte is an H1N1 molecule) is dropped on the surface of the gate electrode 5 surrounded by the PDMS groove, and the surface is modified at 4 ℃ for 15 h to form a target probe molecule layer of the target analyte; the surface of the gate electrode 5 combined with the target probe molecule layer is washed with PBS buffer solution, then 1 mmol / L MCH (6-Mercapto-1-hexanol) solution is added to the target probe molecule layer of the target analyte to fill the blank area of the target probe molecule layer of the target analyte not combined with the gate electrode 5, and the surface is left to stand at room temperature for 1 h to form a 6-mercapto-1-hexanol blocking layer, finally, the surface of the gate electrode 5 is washed with PBS buffer solution, and the target probe molecule layer of the target analyte and the 6-mercapto-1-hexanol blocking layer form a gate modification layer 8.

[0095] The first electrolyte is dropped on the first substrate 1, the drain electrode 4 and the first floating gate 9 to form a first electrolyte layer 6, the second electrolyte is dropped on the second substrate 11, the gate modification layer 8 and the second floating gate 12 to form a second electrolyte layer 7, and the first floating gate 9 and the second floating gate 12 are connected by using a wire 10 to obtain a p-type organic electrochemical transistor biosensor.

[0096] The application of the p-type organic electrochemical transistor biosensor as described above is: the effective gate voltage test method is used for detection of pathogen biomarkers.

[0097] The application realizes the super-high transconductance of 438 mS by selecting the vertical structure OECT device and optimizing the performance. By setting the first floating gate 9 and the second floating gate 12, the gate control ability of the gate 5 is greatly increased, and by using the effective gate voltage test method, the stable device stability is ensured, and the super-sensitive small voltage offset response is realized, which lays a foundation for subsequent realization of ultra-low concentration pathogen detection. The application can realize 10 -20 mol / L H1N1 marker detection, and complete 10 - 17 mol / L H1N1 marker detection, and the current response has a good linear change relationship with the concentration of the detected substance. The preparation method of the p-type organic electrochemical transistor biosensor is simple, and has a wide application prospect in the field of rapid and portable detection of trace biomarkers.

[0098] In the application, the floating gate structure is introduced to separate the test part from the device part, prevent the p-type organic electrochemical transistor from being contaminated by the sample to be detected, ensure that the p-type organic electrochemical transistor biosensor is in the optimal test condition, and adopt the effective gate voltage test method, so that the p-type organic electrochemical transistor sensor can work under a positive voltage, avoiding the problem of small and unstable voltage modulation range of the device under negative voltage, and realizing rapid, sensitive and stable detection.

[0099] Embodiment 1

[0100] The embodiment relates to a preparation method of a p-type organic electrochemical transistor biosensor, and specific steps are as follows:

[0101] Step 1, cleaning the rigid silicon oxide substrate: sequentially placing the substrate in detergent water, ultrapure water, acetone and isopropyl alcohol for ultrasonic cleaning for 30 min.

[0102] Step 2, blowing the substrate surface with an air gun, and then placing it in an ultraviolet ozone cleaning machine for cleaning for 15 min to improve the wettability of the substrate surface.

[0103] Step 3, vacuum evaporation deposition on the cleaned first substrate 1 through a mask, first depositing a 2 nm thick Cr layer as an adhesion layer at a speed of 0.2 Å / s to enhance the bonding force of gold and the substrate, and then depositing a 60 nm thick Au layer at a speed of 1 Å / s~1.5 Å / s to form a source electrode 2; vacuum evaporation deposition on the cleaned second substrate 11 through a mask, first depositing a 2 nm thick Cr layer as an adhesion layer at a speed of 0.2 Å / s to enhance the bonding force of gold and the substrate, and then depositing a 60 nm thick Au layer at a speed of 1 Å / s~1.5 Å / s to form a gate electrode 5; wherein the length of the source electrode 2 is 250 μm, the width is 95 μm, the length of the gate electrode 5 is 9 mm, and the width is 5 mm.

[0104] Step 4, poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] (molecular weight 20000) was added to chloroform, stirred for 15 h, after poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] was fully dissolved, 20 mg / ml poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] chloroform solution was obtained, the first substrate 1 was accelerated to 5000 rpm, 25 μL of poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] chloroform solution was added dropwise on the first substrate 1, and the solvent was completely volatilized in a vacuum environment to form a polymer semiconductor channel 3, the length of the polymer semiconductor channel 3 was 100 μm, the width was 110 μm, and the thickness was 130 nm.

[0105] Step 5, a 60 nm thick Au layer was deposited as a drain 4 on the surface of the polymer semiconductor channel 3 by vacuum evaporation using a mask at a speed of 1 Å / s~1.5 Å / s; wherein the length of the drain 4 was 1200 μm, the width was 95 μm.

[0106] Step 6, the polymer semiconductor channel 3 was patterned by reactive ion etching (RIE) using the drain 4 as a mask plate, and the excess poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] was removed to avoid invalid doping; the etching gas in the reactive ion etching was inert gas argon, the flow rate was 15 sccm, the etching power was 25 w, and the etching time was 250 s.

[0107] Step 7, the first floating gate 9 was formed by coating Ag / AgCl paste on the first substrate 1 using a pre-customized mask plate, and then placing the first substrate 1 in an inert gas atmosphere and performing thermal annealing at 100 ℃ for 30 min, and the second floating gate 12 was formed by coating Ag / AgCl paste on the second substrate 11 using a pre-customized mask plate, and then placing the second substrate 11 in an inert gas atmosphere and performing thermal annealing at 100 ℃ for 30 min, wherein the length of the first floating gate 9 and the second floating gate 12 was 11 mm, the width was 6 mm, and the thickness was 5 μm.

[0108] Step 8, 10 μL of tris (2-carboxyethyl) phosphine (TCEP) aqueous solution with a concentration of 1 mmol / L was added into 100 μL of PBS buffer solution of the target analyte targeting probe molecules with a concentration of 1 μmol / L to obtain a mixed solution. The purpose of adding tris (2-carboxyethyl) phosphine (TCEP) aqueous solution is to open the disulfide bond of the target analyte targeting probe molecules to better bind to the surface of the gate 5. The mixed solution was dropped into the PDMS tank for fixing the modification area, and the second substrate 11 was placed in an environment at 4°C for 15 h for modification to form a layer of the target analyte targeting probe molecules.

[0109] Step 9, the modified surface of the gate 5 was washed with PBS buffer solution, and the surface was blown dry with an ear bulb. In the PDMS tank in step 8, 100 μL of MCH solution with a concentration of 1 mmol / L was added and left to stand at room temperature for 1 h to form a 6-mercapto-1-hexanol blocking layer. The purpose of standing for 1 h is to make the MCH solution cover the surface of the gate 5 which is not covered by the layer of the target analyte targeting probe molecules, and to make the probe molecules arrange regularly to prevent non-specific adsorption on the surface of the gate 5. Then, the surface of the gate 5 was washed with PBS buffer solution to form a gate modification layer 8 with a length of 9 mm, a width of 5 mm and a thickness of 5 nm.

[0110] Step 10, 0.1 mol / L NaCl aqueous solution was dropped on the first substrate 1, the polymer semiconductor channel 3 and the first floating gate 9 as the first electrolyte layer 6; PBS buffer solution was dropped on the second substrate 11, the gate modification layer 8 and the second floating gate 12 as the second electrolyte layer 7, and the first floating gate 9 was connected to the second floating gate 12 by the copper wire 10 to complete the preparation of the p-type organic electrochemical transistor biosensor.

[0111] Among them, the first substrate 1, the source 2, the polymer semiconductor channel 3, the drain 4, the first electrolyte layer 6 and the first floating gate 9 are the organic electrochemical transistor (OECT) device module, and the second substrate 11, the gate 5, the gate modification layer 8, the second electrolyte layer 7 and the second floating gate 12 are the sensing module. The OECT device module is used to amplify the signal perceived by the sensing module, and the two modules together constitute the p-type organic electrochemical transistor biosensor. A drain voltage (Vd) is applied in the loop composed of the source 2, the polymer semiconductor channel 3 and the drain 4 (Vd), and the source 2 is grounded. V DS A gate voltage (Vg) is applied in the loop composed of the source 2, the first floating gate 9, the second floating gate 12, the wire 10, the first electrolyte layer 6, the second electrolyte layer 7, the gate 5 and the gate modification layer 8 (Vg), and the source 2 is grounded. V GS ​

[0112] The performance improvement mechanism of the p-type organic electrochemical transistor biosensor of the present embodiment is as follows:

[0113] 1) Effect of floating gate material on device performance: First, the performance of the OECT device module is optimized, the source 2, the first floating gate 9 and the first electrolyte layer 6 form a loop, and the first floating gate 9 is applied with a gate voltage, and the source 2 is grounded; the source 2, the polymer semiconductor channel 3 and the drain 4 form a loop, and the drain 4 is applied with a drain voltage, and the source 2 is grounded. First, use the polarizable electrode Au as the electrode material of the first floating gate 9 to test the long-term stability of the device module, as shown in Figure 2 , the maximum on-state current of the device module changes irregularly during the triangular wave cyclic scanning process, and this poor cyclic stability will bring problems of excessive noise and baseline drift in the subsequent device module test. Subsequently, use the non-polarizable electrode Ag / AgCl as the electrode material of the first floating gate 9 to test the cyclic stability, as shown in Figure 3 , the device module has good cyclic stability, and the performance of the device module almost does not decay within 3000 s of testing time, which meets the testing requirements of the sensor on performance stability. Therefore, it is speculated that the reason for the poor test stability when Au is used as the gate material is that it is a polarizable electrode, and charges accumulate on the surface during the test, causing the electrolyte and gate interface capacitance to change constantly, which in turn affects the potential at the polymer semiconductor channel, which is manifested as the source-drain current constantly drifting from the perspective of device electrical performance. However, the gate modification layer 8 can only bind with Au, so the present application uses Ag / AgCl paste as the floating gate to read the surface potential change of the gate 5, thereby realizing the conduction of the device module and greatly improving the stability of the device.

[0114] 2) Effect of different gate voltage application on device performance: The second floating gate 12 reads the surface potential change of the gate 5, and the test results are shown in Figure 4 and Figure 5 , when a positive voltage is applied to the surface of the gate 5, the second floating gate 12 reads out a voltage that is linearly related to the applied triangular wave voltage, but when a negative voltage is applied to the surface of the gate 5, the read-out voltage has a large loss compared to the applied voltage, and the voltage modulation range is small, and as the number of scanning cycles increases, charges further accumulate on the surface of the gate 5, and the read-out voltage of the second floating gate 12 decreases due to the increase in the surface capacitance of the gate 5. Therefore, in the PBS solution environment, the adsorption and desorption processes of anions on the surface of the gate 5 are reversible under positive gate voltage, but the adsorption and desorption processes of cations are basically irreversible under negative gate voltage, resulting in a small range of actual gate voltage change and weak modulation effect on the device current. Therefore, when testing, a positive gate voltage should be applied to the gate 5 to ensure that the device has a high sensitivity to small voltage changes and produces a significant current response.

[0115] 3) The influence of effective gate voltage test on device performance: Based on the above influence of different gate voltages on device performance, the present application proposes to change the traditional transfer curve test method of applying constant negative drain voltage and scanning negative gate voltage to applying constant 0.1 V positive gate voltage on the gate, and simultaneously applying 0~0.6 V triangular wave voltage scanning on the drain 4. In this process, when the drain 4 voltage is greater than 0.1 V, the gate potential is less than the drain 4 potential, at this time a positive field of the drain 4 pointing to the gate will be generated, and the anions will move to the channel under the action of the electric field force, realizing the doping of the p-type channel under the positive voltage.

[0116] The feasibility of the effective gate voltage test method is analyzed below to determine the modulation range and stability of the applied voltage. As shown in Figure 6 and Figure 7 , in the case of applying a positive voltage, the actual voltage reading of the gate is linear and stable, indicating that this effective gate voltage test method is feasible in principle. The sensing and detection principle is that after specific binding between the measured object and the probe, the electrolyte and the gate interface capacitor C G changes, thereby affecting the actual gate voltage division. Therefore, before being formally applied to actual pathogen detection, the stability of the device under this mode and the sensitivity to changes in gate voltage need to be tested.

[0117] As shown in Figure 8 , Figure 9 , Figure 10 and Figure 11 , the initial drain current I d = 9.27 mA, after one hour of standing I d = 9.37 mA, with a change of about 1 %, which meets the stability requirements of the device test. Subsequently, the specific binding process at the gate is simulated, as shown in Figure 10 and Figure 11 , the biased gate voltage is changed from 0 V to 0.05 V, and the device I d changes from 10.21 mA to 9.27 mA, and a voltage change of 50 mV causes a current response of about 1 mA, proving the feasibility of the OECT (Organic Electrochemical Transistor) device using the effective gate voltage test method for pathogen marker detection.

[0118] Based on the above test results, it can be seen that the setting of the floating gate structure greatly improves the stability of the device during long-term operation, avoiding problems such as excessive noise and baseline drift during subsequent sensing tests. When Au is used as the gate electrode material, ion adsorption and accumulation on the surface occur due to its polarization characteristics during the test process, and this process is only reversible under positive gate voltage. A new effective gate voltage test method is proposed, in which the bias gate voltage and the drain voltage range of the triangular wave scan are set reasonably, and under the driving of the electric field force, anions are doped into the channel and coupled out of the holes, realizing the conduction of the device. Under this test method, the performance of the device only shifts by 1% under 1 h of static conditions, and for a voltage shift of 50 mV, it produces a current response change of nearly 1 mA, greatly confirming the feasibility and superiority of this new test method.

[0119] Example 2

[0120] This embodiment relates to the application of a p-type organic electrochemical transistor biosensor; the p-type organic electrochemical transistor sensor can realize the detection of trace pathogen markers (such as H1N1 virus RNA), and the specific method is as follows:

[0121] Step 1, 10 PBS solutions with a concentration gradient of 10 -12 mol / L~10 -21 mol / L H1N1 RNA were prepared by gradient dilution method.

[0122] Step 2, place the PDMS tank for testing on the surface of the gate 5 of the p-type organic electrochemical transistor biosensor prepared in Example 1, and use a pipette to add 250 μL of PBS buffer solution in the PDMS tank, apply a triangular wave scanning voltage of 0.2 V~ -0.4 V to the first floating gate 9, and apply a bias voltage of -0.6 V between the source 2 and the drain 4, and perform a conventional transfer curve test; then apply a triangular wave scanning voltage of 0~0.6 V between the source 2 and the drain 4, and apply a bias voltage of 0.3 V to the gate, and perform an effective gate voltage test, repeat 10 scanning periods, and take the average value of the maximum current of each period as the baseline current.

[0123] Step 3, after the test is completed, the second substrate 11 with the gate 5 used for testing is taken out, the surface thereof is rinsed using PBS buffer solution, the surface of the gate 5 is blown dry and clean using an ear bulb, then PBS buffer solution is added dropwise on the gate modification layer 8 for incubation, and the time is consistent with the time required for specific binding in subsequent step 4. After the incubation is completed, the surface thereof is rinsed using PBS buffer solution, the surface of the gate 5 is blown dry and clean using an ear bulb, the PDMS tank used for testing is placed on the surface of the gate 5, 250 μL of PBS buffer solution is added into the PDMS tank using a pipette, a triangular wave scanning voltage of 0.2 V~ -0.4 V is applied to the first floating gate 9, a bias voltage of -0.6 V is applied between the source 2 and the drain 4, and a conventional transfer curve test is performed; then a triangular wave scanning voltage of 0~0.6 V is applied between the source 2 and the drain 4, a bias voltage of 0.3 V is applied to the gate 5, an effective gate voltage test is performed, 10 scanning periods are repeated, and the average value of the maximum current of each period is taken as the noise current.

[0124] Step 4, after the test is completed, the second substrate 11 is taken out, the surface thereof is rinsed using PBS buffer solution, the surface of the gate 5 is blown dry and clean using an ear bulb, then a test concentration of PBS solution is added dropwise on the gate modification layer 8 for incubation, and the time can be adjusted according to the test purpose (for example, the binding time required for the minimum detection limit in this embodiment is 1 h, and the binding time required for the fastest detection speed is 5 min). After the incubation is completed, the surface thereof is rinsed using PBS buffer solution, the surface of the gate is blown dry and clean using an ear bulb, the PDMS tank used for testing is placed on the surface of the gate, 250 μL of PBS buffer solution is added into the PDMS tank using a pipette, a triangular wave scanning voltage of 0.2 V~ -0.4 V is applied to the first floating gate 9, a bias voltage of -0.6 V is applied between the source 2 and the drain 4, and a conventional transfer curve test is performed; then a triangular wave scanning voltage of 0~0.6 V is applied between the source 2 and the drain 4, a bias voltage of 0.3 V is applied to the gate 5, an effective gate voltage test is performed, 10 scanning periods are repeated, and the average value of the maximum current of each period is taken as the response current.

[0125] The principle of the p-type organic electrochemical transistor biosensor for detecting H1N1 according to the present application is shown in Figure 12 、 Figure 13 、 Figure 14 and Figure 15 . The specific RNA sequence of the H1N1 virus can specifically bind to the single-stranded DNA probe modified at the gate 5, and after the binding, the ion transfer to the surface of the gate 5 increases the impedance, reduces the electrolyte and gate interface capacitance C G . Since the electrolyte and channel capacitance C ChThe series relationship is thus established, and the partial pressure of the electrolyte and the channel interface is divided after specific binding occurs V G,Ch The amount of cations driven into the channel by the electric field is reduced, and the source-drain current changes accordingly. Since the amount of change in the effective gate voltage V G,Ch is directly proportional to the concentration of H1N1, the amount of change in the source-drain current is also directly proportional to the concentration of H1N1.

[0126] To verify the ability of the p-type organic electrochemical transistor biosensor to detect low concentrations of biomarkers, a test of ultra-low concentration (10 -20 mol / L) H1N1 was performed using the p-type organic electrochemical transistor biosensor of Example 1, with a binding time of 1 h for the H1N1 marker and the target probe molecule. The data results are shown in FIGS. Figure 16 、 Figure 17 、 Figure 18 、 Figure 19 、 Figure 20 、 Figure 21 and Figure 22 . The baseline current I d,基线 = 7.50±0.03 mA. The noise current I d,噪声 = 7.45±0.02 mA. The shift in the second test data compared to the baseline is taken as the test noise, and the concentration corresponding to a three-fold response of the test noise is defined as the detection limit (LoD). Here, the three-fold noise = 3( I d,基线 - I d,噪声 ) = 136.4 μA. Finally, the response current I d,响应 = 7.13±0.03 mA. In summary, the amount of change in the current Δ I d = I d,噪声 - I d,响应 = 7.45-7.13 = 329.8 μA> 136.4 μA, and the detection is successful. At the same time, a transfer curve test was performed before the effective gate voltage test, to determine whether the performance of the device had degraded. As can be seen from FIGS. Figure 19 、 Figure 20 and Figure 21 , the performance of the p-type organic electrochemical transistor biosensor did not degrade during the test, meaning that the 10 -20 mol / L H1N1 virus was successfully detected.

[0127] In practical application, the minimum detection limit of the device is important, but the rapid current detection in a short time is more in line with the real application scenario. Therefore, the lowest detection limit that can be achieved in combination with the time of 5 min is further explored. The concentration of H1N1 used is 10 -17 mol / L, and the data results are shown in Figure 23 、 Figure 24 、 Figure 25 、 Figure 26 、 Figure 27 、 Figure 28 and Figure 29 . The baseline current I d,基线 = 4.44±0.01 mA. The noise current I d,噪声 = 4.42±0.01 mA, three times the noise = 3( I d,基线 - I d,噪声 ) =51.9 μA, the response current I d,响应 = 4.34±0.04 mA. In summary, the current change amount Δ I d = I d,噪声 - I d,响应 = 4.42-4.34= 80.9 μA>51.9 μA, the detection is successful. At the same time, it can be seen from Figure 26 、 Figure 27 and Figure 28 that the performance of the device does not decay during the test, which means that the 10 -17 mol / L H1N1 virus is successfully detected in the combination time of 5 min.

[0128] In order to realize the quantitative analysis of the concentration of H1N1 in the sample to be tested, H1N1 variable concentration test is carried out in PBS buffer, and the concentration range is from 10 -16 mol / L to 10 -12 mol / L. It can be seen from Figure 30 that the current change response has a good linear relationship with the concentration of H1N1, which reflects the superiority of the performance and test method of the device, and the quantitative detection of H1N1 can be realized.

[0129] Example 3

[0130] The same as example 1, except that in step 3, the length of source electrode 2 is 300 μm, the width is 105 μm, and the thickness is 65 nm; the length of gate electrode 5 is 10 mm, the width is 6 mm, and the thickness is 70 nm;

[0131] In step 4, the length of the polymer semiconductor channel 3 is 100 μm, the width is 105 μm, and the thickness is 140 nm;

[0132] In step 5, the length of the drain 4 is 1400 μm, the width is 105 μm, and the thickness is 65 nm;

[0133] In step 7, the length of the first floating gate 9 and the second floating gate 12 is 11 mm, the width is 6 mm, and the thickness is 10 μm;

[0134] In step 9, the length of the gate modification layer 8 is 10 mm, the width is 6 mm, and the thickness is 8 nm.

[0135] Example 4

[0136] To verify the ability of the p-type organic electrochemical transistor biosensor to detect low concentrations of biomarkers, a p-type organic electrochemical transistor biosensor of Example 3 was used to perform a test of ultra-low concentration (10 -17 mol / L) H1N1, with the binding time of the H1N1 marker and the target probe molecule set to 5 min. The data results are shown in FIGS. 1-3. Figure 31 、 Figure 32 、 Figure 33 、 Figure 34 、 Figure 35 、 Figure 36 and Figure 37 . The baseline current I d,基线 = 4.82±0.11 mA. The noise current I d,噪声 = 4.83±0.08 mA. The shift of the second test data compared with the baseline is taken as the test noise, and the concentration corresponding to a three-fold response of the test noise is defined as the detection limit (LoD). Here, the three-fold noise = 3( I d,基线 - I d,噪声 ) = 30 μA. Finally, the response current I d,响应 = 4.62±0.13 mA. In summary, the amount of current change Δ I d = I d,噪声 - I d,响应 = 4.83-4.62 = 210 μA>30 μA, and the detection is successful. At the same time, a transfer curve test was performed before the effective gate voltage test, to determine whether the performance of the device has been attenuated. From Figure 34 、 Figure 35 and Figure 36It can be seen that the performance of the p-type organic electrochemical transistor biosensor did not decay during the test, meaning that the p-type organic electrochemical transistor biosensor of the present application can be used for a long time. -20 mol / L H1N1 virus was successfully detected.

[0137] Example 5

[0138] In the same manner as in Example 1, except that in Step 3, the length of the source electrode 2 was 280 μm, the width was 100 μm, and the thickness was 70 nm; the length of the gate electrode 5 was 9 mm, the width was 5 mm, and the thickness was 65 nm;

[0139] In Step 4, the length of the polymer semiconductor channel 3 was 110 μm, the width was 100 μm, and the thickness was 150 nm;

[0140] In Step 5, the length of the drain electrode 4 was 1300 μm, the width was 100 μm, and the thickness was 70 nm;

[0141] In Step 7, the length of the first floating gate 9 and the second floating gate 12 was 13 mm, the width was 7 mm, and the thickness was 5 μm;

[0142] In Step 9, the length of the gate modification layer 8 was 9 mm, the width was 5 mm, and the thickness was 10 nm.

[0143] Example 6

[0144] To verify the ability of the p-type organic electrochemical transistor biosensor to detect low concentrations of biomarkers, a p-type organic electrochemical transistor biosensor of Example 5 was used to test H1N1 at an ultra-low concentration (10 -17 mol / L), and the time for which the H1N1 marker was combined with the target probe molecule was set to 5 min. The data results are shown in Figure 38 、 Figure 39 、 Figure 40 、 Figure 41 、 Figure 42 、 Figure 43 and Figure 44 . The baseline current I d,基线 = 6.68±0.03 mA. The noise current I d,噪声 = 6.73±0.01 mA. The shift in the data of the second test compared to the baseline was taken as the test noise, and the concentration corresponding to a three-fold response of the test noise was defined as the limit of detection (LoD). Here, the three-fold noise = 3( I d,基线 - I d,噪声 ) = 150 μA. Finally, the response current I d,响应= 6.56±0.01 mA. In summary, the current variation ΔI I d = I d,噪声 - I d,响应 = 6.73-6.56 = 170.4 μA > 150 μA, the detection is successful. At the same time, the transfer curve test is carried out before the effective gate voltage test, so as to judge whether the performance of the device is attenuated. It can be seen from the following formula that the performance of the p-type organic electrochemical transistor biosensor does not attenuate in the test process, which means that the 10 Figure 41 、 Figure 42 and Figure 43 mol / L H1N1 virus is successfully detected. - 17 mol / L H1N1 virus is successfully detected.

[0145] The above merely describes the preferred embodiments of the present application, and is not intended to limit the technical solutions of the present application in any way. Those skilled in the art should understand that the technical solutions can be simply modified and replaced without departing from the spirit and principles of the present application, and these modifications and replacements also belong to the protection scope covered by the claims.

Claims

1. A p-type organic electrochemical transistor biosensor, characterized in that: Comprising a first substrate (1) and a second substrate (11); A source electrode (2) is provided on one side of the first substrate (1), a polymer semiconductor channel (3) is provided on the source electrode (2), a drain electrode (4) is provided on the polymer semiconductor channel (3), a first floating gate (9) is provided on the other side of the first substrate (1), and a first electrolyte layer (6) is provided on the first substrate (1), the drain electrode (4) and the first floating gate (9); A gate (5) is provided on one side of the second substrate (11), a gate modification layer (8) is provided on the gate (5), a second floating gate (12) is provided on the other side of the second substrate (11), and a second electrolyte layer (7) is provided on the second substrate (11), the gate modification layer (8) and the second floating gate (12); the first floating gate (9) and the second floating gate (12) are connected via a wire (10); A drain voltage is applied to a loop consisting of a source electrode (2), a polymer semiconductor channel (3) and a drain electrode (4), and the source electrode (2) is grounded; A gate voltage is applied to a circuit consisting of a source electrode (2), a first floating gate (9), a second floating gate (12), a wire (10), a first electrolyte layer (6), a second electrolyte layer (7), a gate electrode (5) and a gate modification layer (8); The gate modification layer (8) is a gate modification layer containing a target probe molecule layer of a target analyte.

2. The p-type organic electrochemical transistor biosensor according to claim 1, characterized in that The materials of the first substrate (1) and the second substrate (11) are silicon oxide; The source electrode (2), the drain electrode (4) and the gate electrode (5) are made of Au.

3. The p-type organic electrochemical transistor biosensor according to claim 1, characterized in that The first electrolyte layer (6) is prepared from a NaCl aqueous solution, and the second electrolyte layer (7) is prepared from a PBS buffer solution.

4. The p-type organic electrochemical transistor biosensor according to claim 1, characterized in that: The material of the polymer semiconductor channel (3) is poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene]; The first floating gate (9) and the second floating gate (12) are prepared from Ag / AgCl slurry.

5. The p-type organic electrochemical transistor biosensor according to claim 1, characterized in that: The thickness of the source electrode (2) is 60-70 nm; the thickness of the drain electrode (4) is 60-70 nm; the thickness of the polymer semiconductor channel (3) is 130-150 nm; the thickness of the gate electrode (5) is 60-70 nm; the thickness of the gate modification layer (8) is 5-10 nm; and the thickness of the first floating gate (9) and the second floating gate (12) is 5-10 μm.

6. A method for preparing a p-type organic electrochemical transistor biosensor according to any one of claims 1 to 5, characterized in that: The following steps are involved: Cr is deposited on a first substrate (1) to form a Cr layer, and then Au is deposited on the Cr layer to form a source electrode (2), Cr is deposited on a second substrate (11) to form a Cr layer, and then Au is deposited on the Cr layer to form a gate electrode (5); Spin-coating a chloroform solution of poly[thiophene-bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-2,2'-bithiophene] on the surface of the source electrode (2) to obtain a polymer semiconductor channel (3); Depositing Au on the surface of the polymer semiconductor channel (3) to form a drain electrode (4); A first floating gate (9) is provided on a first substrate (1), and a second floating gate (12) is provided on a second substrate (11); A PBS buffer solution containing the target analyte probe is applied to the surface of the gate (5) and allowed to stand to form a target probe molecular layer of the target analyte; a 6-mercapto-1-hexanol solution is added dropwise to the surface of the gate (5) to cover the surface of the gate (5) not covered by the target probe molecular layer of the target analyte, and allowed to stand to form a 6-mercapto-1-hexanol blocking layer. The target probe molecular layer of the target analyte and the 6-mercapto-1-hexanol blocking layer form a gate modification layer (8); A first electrolyte layer (6) is provided on a first substrate (1), a drain electrode (4) and a first floating gate (9); a second electrolyte layer (7) is provided on a second substrate (11), a gate modification layer (8) and a second floating gate (12); the first floating gate (9) and the second floating gate (12) are connected via a wire (10) to obtain a p-type organic electrochemical transistor biosensor.

7. The method for preparing a p-type organic electrochemical transistor biosensor according to claim 6, characterized in that: A first floating gate (9) is provided on a first substrate (1), and a second floating gate (12) is provided on a second substrate (11), comprising the following steps: Ag / AgCl slurry is coated on a first substrate (1), Ag / AgCl slurry is coated on a second substrate (11), and then annealing is performed to form a first floating gate (9) on the first substrate (1) and a second floating gate (12) on the second substrate (11).

8. The method for preparing a p-type organic electrochemical transistor biosensor according to claim 7, characterized in that: Annealing was carried out under inert gas at a temperature of 100 °C for 30 min.

9. Use of the p-type organic electrochemical transistor biosensor according to any one of claims 1 to 5 in biomarker detection.

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