Device and method for detecting cortisol in tissue fluid

Abstract

The invention discloses a device and a method for detecting cortisol in tissue fluid. The device comprises a tissue fluid extraction and detection module, a wireless transceiving and control circuit module and a power supply module, the tissue fluid extraction and detection module comprises a tissue fluid extraction module, a detection chamber, a waste liquid chamber, an electrolyte chamber and an injection micropump; the tissue fluid extraction module adopts a hollow microneedle array, and realizes extraction of cortisol in tissue fluid under the action of an injection micropump; an electrochemical immunosensing electrode in the detection chamber is used for detection; and the wireless transceiving and control circuit module is used for sending a detection result to a control end. According to the technical scheme provided by the invention, the cortisol concentration of the human body tissue fluid can be rapidly detected, the operation process is simple and convenient, the consumed time is short, the efficiency of the method and the portability of equipment are improved, an accurate, rapid and minimally invasive tissue fluid cortisol sensing method is realized, and a reliable way is provided for acquiring biochemical information of a human body stress state.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technology, and in particular to a device and method for detecting cortisol in tissue fluid. Background Art

[0002] Cortisol is a glucocorticoid hormone that is involved in many physiological processes and is also a good indicator of animal stress tolerance. The release of cortisol is regulated by the hypothalamus-pituitary adrenal axis, and increased cortisol secretion is considered a stress response to stress. Cortisol can accelerate carbohydrate metabolism in animals and increase the availability of glucose in the blood, which helps to cope with short-term stress. At the same time, cortisol is also an important physiological indicator that reflects the intensity of stress when an individual animal faces various stimuli. The right amount of cortisol secretion helps to cope with stress, but excessive cortisol secretion is not good for the health of the body and can cause mental disorders such as anxiety and depression. Cortisol testing can be used to monitor stress so that measures can be taken to avoid serious consequences.

[0003] At present, some wearable devices based on electrochemical immunosensors are used to detect sweat cortisol, but the concentration of sweat cortisol is much lower than that of blood, and cannot accurately reflect the actual state of the body in a timely manner. Compared with sweat cortisol, tissue fluid cortisol can more accurately reflect the adrenal cortisol secretion caused by stress factors. However, there is currently no public technology for detecting tissue fluid cortisol in living bodies.

[0004] In addition, in special circumstances, such as when testing tissue fluid cortisol in children or animals, the tissue fluid cortisol testing device should have a remote control function due to its "inaccessible" nature. Summary of the invention

[0005] In order to solve the above-mentioned deficiencies and defects, the present application provides a tissue fluid cortisol detection device and method.

[0006] The first aspect of the embodiment of the present invention discloses a tissue fluid cortisol detection device, comprising: a tissue fluid extraction detection module, and a wireless transceiver and control circuit module and a power supply module fixed on the top of the tissue fluid extraction detection module; The tissue fluid extraction detection module comprises a shell; the bottom surface of the shell is provided with a tissue fluid extraction module; the bottom surface represents the side in contact with the skin of the detected object, and the tissue fluid extraction module is attached to the body surface of the detected object during operation.

[0007] A detection chamber, a waste liquid chamber and an electrolyte chamber are arranged inside the shell.

[0008] The detection chamber is connected to the waste liquid chamber through a first microchannel, a first pneumatic film microvalve is arranged on the first microchannel, and the first pneumatic film microvalve is connected to a first injection micropump through a first hose; the first injection micropump is used to provide positive pressure or negative pressure to the first pneumatic film microvalve, and the first pneumatic film microvalve is used to control the opening or closing of the first microchannel, and the first pneumatic film microvalve is initially in a closed state.

[0009] The electrolyte chamber stores electrolyte, the detection chamber is connected to the electrolyte chamber via a second microchannel, a second pneumatic film microvalve is provided on the second microchannel, and the second pneumatic film microvalve is connected to a second injection micropump via a second hose; the second injection micropump is used to provide positive pressure or negative pressure to the second pneumatic film microvalve, the second pneumatic film microvalve is used to control the opening or closing of the second microchannel, and the second pneumatic film microvalve is initially in a closed state.

[0010] An electrochemical immunosensor electrode is arranged on the top of the detection chamber; the electrochemical immunosensor electrode comprises a working electrode, a counter electrode and a reference electrode; the working electrode and the counter electrode are gold electrodes, the reference electrode is a silver / silver chloride electrode, and the surface of the working electrode is modified with cortisol antibodies; a third injection micropump is connected to the top of the detection chamber through a third hose, and the third injection micropump is used to provide positive pressure or negative pressure for the detection chamber.

[0011] The first injection micro pump, the second injection micro pump and the third injection micro pump are fixed on the top of the tissue fluid extraction and detection module.

[0012] The wireless transceiver and control circuit module includes a wireless transceiver module, a microcontroller and a detection control circuit; the microcontroller is electrically connected to the wireless transceiver module, the first injection micropump, the second injection micropump, the third injection micropump and the detection control circuit; the detection control circuit is electrically connected to the electrochemical immunosensor electrode; the wireless transceiver module is used to receive wireless control commands from the control end and transmit detection results to the control end; the control end represents a control terminal used by the operator; the start command represents an instruction to start detection.

[0013] The power supply module is electrically connected to the wireless transceiver and control circuit module, the first injection micropump, the second injection micropump, and the third injection micropump for providing electrical energy.

[0014] As an optional embodiment, in the first aspect of the embodiment of the present invention, the tissue fluid extraction module is a hollow microneedle array; the hollow microneedle array includes M×N hollow microneedles; the needle body of the hollow microneedle is hollow, and the base of the hollow microneedle is connected to the detection chamber; the hollow microneedle array is evenly distributed below the detection chamber; the values ​​of M and N are determined according to actual needs.

[0015] As an optional embodiment, in the first aspect of the embodiment of the present invention, the hollow microneedle is conical, with a bottom width of 450 microns to 550 microns, a height of 950 microns to 1050 microns, and a needle tip spacing of 1.4 to 1.6 mm; the hollow pore diameter of the hollow microneedle is 120 to 130 microns.

[0016] As an optional embodiment, in the first aspect of the embodiment of the present invention, the electrolyte stored in the electrolyte chamber is a PBS (phosphate buffer) solution containing 1 mM potassium ferrocyanide, 1 mM potassium ferrocyanide and 100 mM potassium chloride; the volumes of the detection chamber, the waste liquid chamber and the electrolyte chamber are the same.

[0017] As an optional embodiment, in the first aspect of the embodiment of the present invention, the electrochemical immunosensor electrode is prepared by modifying antibodies on a screen-printed electrode; The working electrode was modified with an antibody using a cross-linking agent DSP (dithiobis(succinimidyl propionate), Lomant's reagent) and then blocked with BSA (bovine serum albumin).

[0018] As an optional embodiment, in the first aspect of the embodiment of the present invention, the housing, tissue fluid extraction module, detection chamber, waste liquid chamber, electrolyte chamber, first microchannel, first pneumatic film microvalve, second microchannel, second pneumatic film microvalve, third microchannel and third pneumatic film microvalve included in the tissue fluid extraction detection module are 3D printed with biocompatible resin and prepared as a whole. The various components of the tissue fluid extraction detection module are prepared as a whole by 3D printing, which ensures the airtightness and reliability of the tissue fluid extraction detection module.

[0019] As an optional implementation, in the first aspect of the embodiment of the present invention, the wireless transceiver module adopts the 433MHz frequency band and the power consumption is not more than 12mW; the microcontroller is a single chip microcomputer with a power consumption of not more than 10mW.

[0020] As an optional implementation, in the first aspect of the embodiment of the present invention, the power module uses a 3.7V lithium battery as a power supply, the capacity of the lithium battery is not less than 300 mAh, and the number of charge and discharge cycles is not less than 100 times; the power module also includes a battery charge and discharge management circuit for charging management and over-discharge prevention management of the lithium battery.

[0021] The second aspect of the embodiment of the present invention discloses a method for detecting cortisol in tissue fluid, which is applied to the device for detecting cortisol in tissue fluid disclosed in the first aspect of the embodiment of the present invention. The method comprises: The tissue fluid cortisol detection device is attached to the smooth body surface of the subject to be detected, so that the tissue fluid extraction module is in contact with the body surface; Applying pressure to pierce the microneedle array of the tissue fluid extraction detection module into the skin; After receiving the start command sent by the control end, the wireless transceiver module transmits the start command to the microcontroller; the control end represents the control terminal used by the operator; the start command represents the start detection instruction; According to the start command, the microcontroller controls the third injection micropump to provide negative pressure to the detection chamber, so as to extract tissue fluid in the skin of the detected object from the tissue fluid extraction module to the detection chamber; After the tissue fluid enters the detection chamber, the cortisol in the tissue fluid binds to the antibody on the working electrode for no less than 30 minutes; The microcontroller controls the first injection micropump to provide negative pressure to the first pneumatic film microvalve, so that the first pneumatic film microvalve opens; the microcontroller controls the third injection micropump to provide positive pressure to the detection chamber, and the remaining tissue fluid in the detection chamber flows into the waste liquid chamber through the first microchannel; The microcontroller controls the first injection micropump to provide positive pressure to the first pneumatic film microvalve, so that the first pneumatic film microvalve is closed; the microcontroller controls the second injection micropump to provide negative pressure to the second pneumatic film microvalve, so that the second pneumatic film microvalve is opened; the microcontroller controls the third injection micropump to provide negative pressure to the detection chamber, and the electrolyte in the electrolyte chamber flows into the detection chamber through the second microchannel; The microcontroller controls the detection control circuit to apply voltage to the electrochemical immunosensor electrode to perform electrochemical impedance spectroscopy (EIS) detection to measure the cortisol concentration in the tissue fluid; The cortisol concentration of the tissue fluid is transmitted to the control end by using a wireless transceiver module.

[0022] Compared with the prior art, the embodiments of the present invention have the following beneficial effects: 1. The present application provides a tissue fluid cortisol detection device and method that can quickly detect the cortisol concentration in human tissue fluid. The operation process is simple, convenient, and time-saving. Compared with the existing sweat cortisol concentration detection, this method can timely and accurately reflect the actual state of the body.

[0023] 2. This application integrates multiple functions into a microcontroller to complete human tissue fluid extraction and cortisol detection in an integrated manner, greatly increasing the efficiency of the method and the portability of the equipment, and realizing an accurate, fast, and minimally invasive tissue fluid cortisol sensing method, providing a reliable way to obtain biochemical information on the human body's stress state.

[0024] 3. The present application utilizes a wireless transceiver and control circuit module to remotely control a tissue fluid cortisol detection device, and can achieve extraction and detection of tissue fluid cortisol without approaching the object being detected.

[0025] 4. The present application adopts a method of extracting tissue fluid by applying negative pressure through a hollow microneedle under controlled conditions, thereby achieving safe extraction of tissue fluid in a trace and local extraction manner. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 A schematic diagram of the structure of a tissue fluid cortisol detection device disclosed in an embodiment of the present invention; Figure 2 It is a schematic diagram of a top view of a tissue fluid cortisol detection device disclosed in an embodiment of the present invention; Figure 3 A schematic diagram of an electrochemical immunosensor electrode structure disclosed in an embodiment of the present invention; Figure 4 A schematic diagram of a tissue fluid extraction module disclosed in an example of the present invention; Figure 5 A flow chart of an electrochemical immunosensor electrode modification disclosed in an example of the present invention; Figure 6 The module workflow diagram of a tissue fluid cortisol detection device disclosed in an example of the present invention.

[0027] Reference numerals and descriptions: 100-housing; 200-wireless transceiver and control circuit module; 300-power module; 101-tissue fluid extraction module; 102-detection chamber; 103-waste liquid chamber; 104-electrolyte chamber; 105-first microchannel; 106-second microchannel; 107-first pneumatic membrane microvalve; 108-second pneumatic membrane microvalve; 109-first injection micropump; 110-second injection micropump; 111-third injection micropump; 112-electrochemical immunosensor electrode; 113-working electrode; 114-counter electrode; 115-reference electrode; 201-wireless transceiver module; 202-microcontroller; 203-detection control circuit. DETAILED DESCRIPTION

[0028] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0029] Embodiment 1 See also Figure 1 , Figure 2 It should be noted that Figure 1 , Figure 2 It is a schematic diagram only and is not drawn to scale.

[0030] like Figure 1 As shown, an interstitial fluid cortisol detection device disclosed in an embodiment of the present invention includes: an interstitial fluid extraction detection module, and a wireless transceiver and control circuit module 200 and a power supply module 300 fixed to the top of the interstitial fluid extraction detection module; the interstitial fluid extraction detection module includes a shell 100; a tissue fluid extraction module 101 is arranged on the bottom surface of the shell 100; the bottom surface represents a side in contact with the skin of the detected object, and the tissue fluid extraction module 101 is attached to the body surface of the detected object during operation.

[0031] like Figure 2 As shown, a detection chamber 102, a waste liquid chamber 103 and an electrolyte chamber 104 are arranged inside the shell 100; the detection chamber 102 is connected to the waste liquid chamber 103 through a first microchannel 105, a first pneumatic film microvalve 107 is arranged on the first microchannel 105, and the first pneumatic film microvalve 107 is connected to a first injection micropump 109 through a first hose (not shown in the figure); the first injection micropump 109 is used to provide positive pressure or negative pressure for the first pneumatic film microvalve 107, and the first pneumatic film microvalve 107 is used to control the opening or closing of the first microchannel 105, and the first pneumatic film microvalve 107 is initially in a closed state.

[0032] The detection chamber 102 is connected to the electrolyte chamber 104 through a second microchannel 106, and a second pneumatic film microvalve 108 is arranged on the second microchannel 106. The second pneumatic film microvalve 108 is connected to a second injection micropump 110 through a second hose (not shown in the figure); the second injection micropump 110 is used to provide positive pressure or negative pressure to the second pneumatic film microvalve 108, and the second pneumatic film microvalve 108 is used to control the opening or closing of the second microchannel 106. The second pneumatic film microvalve 108 is initially in a closed state.

[0033] It should be noted that the automatic collection of tissue fluid is achieved through the coordinated control of the above-mentioned injection micropump, which solves the problem of discomfort caused to the subject when collecting tissue fluid directly with a syringe. The discomfort caused by collecting tissue fluid can be reduced by adjusting the power of the injection micropump.

[0034] The top of the detection chamber 102 is provided with an electrochemical immunosensor electrode 112; Figure 3As shown, the electrochemical immunosensor electrode 112 includes a working electrode 113, a counter electrode 114 and a reference electrode 115; the working electrode and the counter electrode are gold electrodes, the reference electrode is a silver / silver chloride electrode, and the surface of the working electrode is modified with cortisol antibodies.

[0035] The top of the detection chamber 102 is connected to a third injection micropump 111 through a third hose (not shown in the figure), and the third injection micropump 111 is used to provide positive pressure or negative pressure to the detection chamber 102; the first injection micropump 109, the second injection micropump 110 and the third injection micropump 111 are fixed on the top of the tissue fluid extraction detection module.

[0036] The wireless transceiver and control circuit module 200 includes a wireless transceiver module 201, a microcontroller 202 and a detection control circuit 203; the microcontroller 202 is electrically connected to the wireless transceiver module 201, the first injection micropump 109, the second injection micropump 110, the third injection micropump 111 and the detection control circuit 203; the detection control circuit 203 is electrically connected to the electrochemical immunosensor electrode 112; the wireless transceiver module 201 is used to receive the wireless control command of the control end and transmit the detection result to the control end, and the control end represents the control terminal used by the operator; the microcontroller 202 controls the first injection micropump 109, the second injection micropump 110, the third injection micropump 111 and the detection control circuit 203 according to the control command; the control of the wireless transceiver and control circuit module can realize the automated processing of tissue fluid collection and the precise control of the collection amount, thereby enhancing the comfort of the collected object.

[0037] The power module 300 is electrically connected to the wireless transceiver and control circuit module 200 , the first injection micropump 109 , the second injection micropump 110 , and the third injection micropump 111 to provide electrical energy.

[0038] It can be seen that the tissue fluid cortisol detection device described in this embodiment can integrate tissue fluid extraction and cortisol detection, greatly increase the efficiency of the method and the portability of the equipment, realize an accurate and rapid tissue fluid cortisol sensing method, and can quickly detect the body's tissue fluid cortisol. The operation process is simple and convenient, and the time is short, which reduces the discomfort of the collected object, provides a reliable way to obtain biochemical information of the body's stress state, and solves the problem in the prior art that the sweat cortisol detection results cannot timely and accurately reflect the actual state of the body.

[0039] In another optional embodiment, the tissue fluid extraction module 101 is a hollow microneedle array, such as Figure 4As shown; the hollow microneedle array uses M×N hollow microneedles; the needle body of the hollow microneedle is hollow, and the base of the hollow microneedle is connected to the detection chamber 102. The hollow microneedle array is evenly distributed below the detection chamber 102; the value range of M is 4-6, and the value range of N is 10-12.

[0040] In this embodiment, the hollow microneedle array adopts a 5×11 hollow microneedle array, which can meet the amount of tissue fluid required for detection and is not likely to cause discomfort to the subject being collected.

[0041] It can be seen that the tissue fluid cortisol detection device described in this embodiment can reduce the discomfort of the subject being collected by performing minimally invasive collection of tissue fluid cortisol using a hollow microneedle array.

[0042] In another optional embodiment, the hollow microneedle is conical, with a bottom width of 450 to 550 microns, a height of 950 to 1050 microns, and a needle tip spacing of 1.4 to 1.6 mm; the hollow hole diameter of the microneedle is 120 to 130 microns, and the distance between the center of the hole and the center of the hollow microneedle is 120 to 130 microns.

[0043] Preferably, the bottom width of the hollow microneedle is 500 microns, the height is 1000 microns, and the needle tip spacing is 1.5 mm; the hollow hole diameter of the hollow microneedle is 125 microns, and the distance between the hole center and the hollow microneedle center is 125 microns.

[0044] In another optional embodiment, the electrolyte stored in the electrolyte chamber is a PBS (phosphate buffered saline) solution containing 1 mM potassium ferrocyanide, 1 mM potassium ferrocyanide and 100 mM potassium chloride; the detection chamber 102, the waste liquid chamber 103 and the electrolyte chamber 104 have the same volume; As an optional implementation method, the detection chamber 102, the waste liquid chamber 103 and the electrolyte chamber 104 have a length of 18 mm, a width of 8 mm and a height of 500 μm.

[0045] In yet another optional embodiment, the electrochemical immunosensor electrode 112 is prepared by modifying antibodies on a screen-printed electrode.

[0046] For specific procedures, please refer to Figure 5. Add 5μL 10mM DSP (dithiobis(succinimidyl propionate), Lomant reagent) to the working electrode and incubate at room temperature for 30min to 1h. Wash the electrode with DMSO (dimethyl sulfoxide) and PBS (phosphate buffered saline) in sequence, and then immediately add 20μL 400-600μg / mL cortisol antibody to the working electrode and incubate at room temperature for about 2h. Wash the electrode with PBS (phosphate buffered saline) and blow it dry with nitrogen, then add 20μL 0.5% (w / w) BSA (bovine serum albumin) to the working electrode and incubate it at room temperature for 1-2h. Finally, wash the electrode with PBS and blow it dry with nitrogen.

[0047] In an optional embodiment, the shell, tissue fluid extraction module, detection chamber, waste liquid chamber, electrolyte chamber, first microchannel, first pneumatic film microvalve, second microchannel, second pneumatic film microvalve, third microchannel and third pneumatic film microvalve included in the tissue fluid extraction detection module are 3D printed with biocompatible resin and prepared as a whole. The various components of the tissue fluid extraction detection module are prepared as a whole by 3D printing, which ensures the airtightness and reliability of the tissue fluid extraction detection module. The tissue fluid extraction detection module shell prepared with biocompatible resin has good biocompatibility and corrosion resistance, making the detection results more accurate; it has good pressure resistance and bending resistance, and is more suitable for the body of the collected object.

[0048] In an optional embodiment, the wireless transceiver module 201 uses a 433 MHz frequency band and has a power consumption of no more than 12 mW; the microcontroller 202 is a single chip microcomputer with a power consumption of no more than 10 mW.

[0049] It should be noted that the tissue fluid cortisol detection device provided in this application can be used for humans and animals. This application uses a wireless transceiver module to receive, process and output signals, realizes the wireless transceiver function under the 433MHz frequency band, can realize long-distance control, and can collect tissue fluid even when not close to the object being detected. The 433MHz frequency band is selected for communication, which has the characteristics of high reliability, high stability, strong anti-interference ability, strong penetration ability and strong environmental adaptability.

[0050] In an optional embodiment, the power module 300 uses a lithium battery as a power supply, the capacity of the lithium battery is not less than 300 mAh, and the number of charge and discharge cycles is not less than 100 times.

[0051] The power module 300 also includes a battery charge and discharge management circuit for managing the charge and over-discharge prevention of the lithium battery.

[0052] It can be seen that the tissue fluid cortisol detection device described in this embodiment uses a low-power microcontroller and a lithium battery, which achieves miniaturization of the device and prolongs the working time.

[0053] Embodiment 2 This embodiment provides a method for detecting cortisol in tissue fluid, which is applied to the device for detecting cortisol in tissue fluid disclosed in the first embodiment of the present invention. The method includes: The tissue fluid cortisol detection device is attached to the smooth body surface of the subject to be detected, so that the tissue fluid extraction module 101 is in contact with the body surface; Applying pressure to pierce the microneedle array of the tissue fluid extraction module 101 into the skin; like Figure 6 As shown, after receiving the start command from the control end, the wireless transceiver module 201 transmits the start command to the microcontroller 202; the control end represents the control terminal used by the operator; the start command represents the start detection instruction; According to the start command, the microcontroller 202 controls the third injection micropump to provide negative pressure to the detection chamber 102, so as to extract the tissue fluid in the skin of the detected object from the tissue fluid extraction module to the detection chamber 102; After the tissue fluid enters the detection chamber 102, the cortisol in the tissue fluid binds to the antibody on the working electrode for 30 minutes; The microcontroller 202 controls the first injection micropump 109 to provide negative pressure to the first pneumatic film microvalve 107, so that the first pneumatic film microvalve 107 is opened; the microcontroller 202 controls the third injection micropump 111 to provide positive pressure to the detection chamber 102, and the remaining tissue fluid in the detection chamber 102 flows into the waste liquid chamber 103 through the first microchannel 105; The microcontroller 202 controls the first injection micropump 109 to provide positive pressure to the first pneumatic film microvalve 107, so that the first pneumatic film microvalve 107 is closed; the microcontroller 202 controls the second injection micropump 110 to provide negative pressure to the second pneumatic film microvalve 108, so that the second pneumatic film microvalve 108 is opened; the microcontroller 202 controls the third injection micropump 111 to provide negative pressure to the detection chamber 102, and the electrolyte in the electrolyte chamber 104 flows into the detection chamber 102 through the second microchannel 106; The microcontroller 202 controls the detection control circuit 203 to apply voltage to the electrochemical immunosensor electrode 112 to perform electrochemical impedance spectroscopy (EIS) detection, and measures an electrical signal reflecting the cortisol concentration; the detection control circuit 203 transmits the electrical signal to the microcontroller 202 for data processing; the microcontroller 202 transmits the processed concentration signal to the wireless transceiver module 201; the wireless transceiver module 201 transmits the concentration signal to the control end, and the tissue fluid cortisol concentration is calculated by software and presented to the user.

[0054] Embodiment 3 This embodiment provides a detection method of a tissue fluid cortisol detection device.

[0055] The morphology of the prepared cortisol electrochemical detection microneedle array was characterized by scanning electron microscopy, stereo microscope and optical microscope. The microneedles were glued to the sample stage, sprayed with gold and purged with nitrogen. The microneedle morphology was observed and the size was measured using a scanning electron microscope, and the microneedle morphology was further observed under a stereo microscope and an optical microscope.

[0056] Microneedle tissue fluid extraction behavior test. Frozen ex vivo SD rat skin was removed, immersed in physiological saline to return to room temperature, and gauze was used to gently wipe off the moisture on the skin surface. The microneedle was vertically inserted into the rat skin using a universal tensile testing machine, and negative pressure was applied to the detection chamber 102 to extract tissue fluid. The microneedle was removed after 1 minute, and the weight of the microneedle before and after tissue fluid extraction was weighed using a balance to calculate the amount of tissue fluid extracted. The intensity and time of negative pressure application during microneedle application were evaluated by measuring the tissue fluid extraction behavior after the microneedle was inserted into the ex vivo SD rat skin. The microneedle was vertically inserted into the rat skin, and the microneedle was removed after applying negative pressure for 0 to 10 minutes. The weight of the microneedle before and after tissue fluid extraction was weighed using a balance, and the microneedle tissue fluid extraction behavior was evaluated by calculating the weight difference.

[0057] It should be noted that when an element is referred to as being "fixed on", "provided with" or "connected to" another element, it can be directly on the other element or there can be a central element. "Fixed on", "provided on" or "connected to" can be a fixed connection, a threaded connection, an adhesive connection, a snap connection or one or more connection methods selected according to actual conditions. An element can be understood as a component or a part. The terms "vertical", "horizontal", "upper", "lower", "left", "right" and similar expressions used herein are for illustrative purposes only and do not represent the only implementation method.

[0058] Finally, it should be noted that the tissue fluid cortisol detection device and method disclosed in the embodiments of the present invention disclose only the preferred embodiments of the present invention, which are only used to illustrate the technical scheme of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, a person skilled in the art should understand that the technical schemes described in the aforementioned embodiments can still be modified, or some of the technical features thereof can be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical schemes from the spirit and scope of the technical schemes of the embodiments of the present invention.

Claims

1. A tissue fluid cortisol detection device, characterized in that: The device comprises a tissue fluid extraction and detection module, and a wireless transceiver and control circuit module and a power supply module fixed on the top of the tissue fluid extraction and detection module; The tissue fluid extraction detection module includes a shell; a tissue fluid extraction module is arranged on the bottom surface of the shell, and a detection chamber, a waste liquid chamber and an electrolyte chamber are arranged inside; the detection chamber is connected to the waste liquid chamber through a first microchannel, a first pneumatic film microvalve is arranged on the first microchannel, and the first pneumatic film microvalve is connected to a first injection micropump through a first hose; electrolyte is stored in the electrolyte chamber, the detection chamber is connected to the electrolyte chamber through a second microchannel, a second pneumatic film microvalve is arranged on the second microchannel, and the second pneumatic film microvalve is connected to a second injection micropump through a second hose. The first pneumatic film microvalve and the second pneumatic film microvalve are initially in a closed state; an electrochemical immunosensor electrode is arranged on the top of the detection chamber; the electrochemical immunosensor electrode comprises a working electrode, a counter electrode and a reference electrode; the working electrode and the counter electrode are gold electrodes, the reference electrode is a silver / silver chloride electrode, and the surface of the working electrode is modified with cortisol antibody; the top of the detection chamber is connected to a third injection micropump through a third hose; the first injection micropump, the second injection micropump and the third injection micropump are fixed on the top of the tissue fluid extraction detection module; The wireless transceiver and control circuit module includes a wireless transceiver module, a microcontroller and a detection control circuit; the microcontroller is electrically connected to the wireless transceiver module, the first injection micropump, the second injection micropump, the third injection micropump and the detection control circuit; The detection control circuit is electrically connected to the electrochemical immunosensor electrode.

2. The tissue fluid cortisol detection device according to claim 1, characterized in that: The tissue fluid extraction module is a hollow microneedle array; the needle body of the hollow microneedle is hollow, and the base of the hollow microneedle is connected to the detection chamber; The hollow microneedle array is evenly distributed below the detection chamber.

3. The tissue fluid cortisol detection device according to claim 2, characterized in that: The hollow microneedle is conical, with a bottom width of 450 to 550 microns, a height of 950 to 1050 microns, and a needle tip spacing of 1.4 to 1.6 millimeters; the hollow pore diameter of the hollow microneedle is 120 to 130 microns.

4. The tissue fluid cortisol detection device according to claim 1, characterized in that: The electrolyte stored in the electrolyte chamber is a PBS solution containing 1 mM potassium ferrocyanide, 1 mM potassium ferrocyanide and 100 mM potassium chloride; the volumes of the detection chamber, the waste liquid chamber and the electrolyte chamber are the same.

5. The tissue fluid cortisol detection device according to claim 1, characterized in that: The electrochemical immunosensor electrode is prepared by modifying antibodies on a screen-printed electrode; the antibody is modified on the working electrode using a cross-linking agent DSP and then blocked with bovine serum albumin.

6. The tissue fluid cortisol detection device according to claim 1, characterized in that: The tissue fluid extraction detection module comprises a shell, a tissue fluid extraction module, a detection chamber, a waste liquid chamber, an electrolyte chamber, a first microchannel, a first pneumatic film microvalve, a second microchannel, a second pneumatic film microvalve, a third microchannel and a third pneumatic film microvalve, which are 3D printed with biocompatible resin and prepared as a whole.

7. The tissue fluid cortisol detection device according to claim 1, characterized in that: The wireless transceiver module uses a 433MHz frequency band and has a power consumption of no more than 12mW; the microcontroller is a single chip microcomputer with a power consumption of no more than 10mW.

8. The tissue fluid cortisol detection device according to claim 1, characterized in that: The power module uses a 3.7V lithium battery as a power supply. The capacity of the lithium battery is not less than 300 mAh, and the number of charge and discharge cycles is not less than 100 times. The power module also includes a battery charge and discharge management circuit.

9. A method for detecting cortisol in tissue fluid, characterized in that: Applied to the tissue fluid cortisol detection device according to any one of claims 1 to 8, the method comprising: The tissue fluid cortisol detection device is attached to the smooth body surface of the subject to be detected, so that the tissue fluid extraction module is in contact with the body surface; Apply pressure to pierce the tissue fluid extraction module into the skin; After receiving the start command sent by the control end, the wireless transceiver module transmits the start command to the microcontroller; the control end represents the control terminal used by the operator; the start command represents the start detection instruction; According to the start command, the microcontroller controls the third injection micropump to provide negative pressure to the detection chamber, so as to extract tissue fluid in the skin of the detected object from the tissue fluid extraction module to the detection chamber; After the tissue fluid enters the detection chamber, the cortisol in the tissue fluid binds to the antibody on the working electrode for no less than 30 minutes; The microcontroller controls the first injection micropump to provide negative pressure to the first pneumatic film microvalve, so that the first pneumatic film microvalve opens; the microcontroller controls the third injection micropump to provide positive pressure to the detection chamber, and the remaining tissue fluid in the detection chamber flows into the waste liquid chamber through the first microchannel; The microcontroller controls the first injection micropump to provide positive pressure to the first pneumatic film microvalve, so that the first pneumatic film microvalve is closed; the microcontroller controls the second injection micropump to provide negative pressure to the second pneumatic film microvalve, so that the second pneumatic film microvalve is opened; the microcontroller controls the third injection micropump to provide negative pressure to the detection chamber, and the electrolyte in the electrolyte chamber flows into the detection chamber through the second microchannel; The microcontroller controls the detection control circuit to apply voltage to the electrochemical immunosensor electrode to perform electrochemical impedance spectroscopy detection to measure the cortisol concentration in the tissue fluid; The cortisol concentration of the tissue fluid is transmitted to the control end by using a wireless transceiver module.

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