Cerebral arterial thrombosis diagnosis and prognosis evaluation kit based on TFAM

Abstract

The invention discloses a TFAM-based cerebral arterial thrombosis diagnosis and prognosis evaluation kit, and relates to the technical field of medical detection, in particular to a TFAM-based cerebral arterial thrombosis diagnosis and prognosis evaluation kit, which comprises a kit body, a base, an isolation plate, a rotatable reagent rack (containing reagent tubes) arranged on one side of the isolation plate, and a liquid medicine rack (containing liquid medicine bottles) arranged on one side of the isolation plate, an elisa plate is arranged on the other side; automatic ejection of reagent tubes and liquid medicine bottles is realized through structures such as a motor, a cam and a push rod, a telescopic rod and a connecting rod mechanism control opening and closing of a reagent tube sealing cover, and an elisa plate can be driven by an ejector rod to move up and down to uniformly mix liquid in a hole; a control panel on the outer side of the box body is electrically connected with the motors and the telescopic rods, and centralized control is achieved. The kit is high in integration and automation degree, convenient to operate and suitable for rapid diagnosis and prognosis evaluation of cerebral arterial thrombosis.

Description

Technical Field

[0001] This invention belongs to the field of medical testing technology, specifically a TFAM-based diagnostic and prognostic kit for ischemic stroke. Background Technology

[0002] With the increasing aging population and changes in lifestyle, the incidence of ischemic stroke is rising year by year. Early and accurate diagnosis and prognostic assessment are crucial for patient treatment and rehabilitation. Existing stroke detection kits often suffer from problems such as dispersed structure and cumbersome operation. Reagents need to be handled manually, which introduces uncertainty in reagent and drug usage, easily leading to misuse or non-use. At the same time, opening and closing the sealed cap can easily cause contamination, resulting in poor liquid mixing and low detection efficiency and insufficient accuracy. On the other hand, during testing, personnel need to repeatedly shake and mix the test solution and reagents in the wells of the enzyme-labeled plate to ensure reaction. However, the shaking operation by employees is difficult to fully mix, which can lead to low reliability in ensuring diagnosis and prognostic assessment and fails to meet the actual needs of rapid clinical diagnosis and prognostic assessment. Therefore, this invention proposes a TFAM-based diagnostic and prognostic assessment kit for ischemic stroke. Summary of the Invention

[0003] This invention provides a TFAM-based diagnostic and prognostic kit for ischemic stroke, which solves the problems mentioned in the background.

[0004] To achieve the above objectives, the present invention provides the following technical solution: A TFAM-based diagnostic and prognostic kit for ischemic stroke includes a box body. An isolation plate is installed in the middle of the box body, and a base is installed at the bottom of the box body. A reagent rack is symmetrically and rotatably connected to the top of the base on one side of the isolation plate. The reagent rack has an array of through holes, in which reagent tubes are placed. Partition 1 and Partition 2 are installed between the reagent racks and on the right side of the reagent rack, respectively. An annular slider is installed on the outer surface of the reagent rack, rotatably connected to the side walls of the isolation plate, the box body, Partition 1, and Partition 2. An L-shaped block is installed on the top of the base on one side below the reagent rack, and a push rod is slidably connected to the L-shaped block. The push rod can slide to push the reagent tubes in the through holes upwards. A sealing cap is sealed on the top of the reagent tube. A linkage mechanism is rotatably connected to the top of the sealing cap, and a telescopic rod is rotatably connected to the other end of the linkage mechanism. The telescopic rod can move the sealing cap closer to or away from the top of the reagent tube through the linkage mechanism. An ELISA plate is disposed on the other side of the isolation plate. The ELISA plate can move up and down inside the box body and can mix the liquid in the wells of the ELISA plate.

[0005] Preferably, a rotating shaft is installed in the middle of the bottom of the reagent rack, a helical gear is installed at the lower end of the rotating shaft, a motor is installed on one side of the helical gear, and a worm gear is fixedly connected to the output shaft of the motor, the worm gear meshing with the helical gear.

[0006] Preferably, a side plate is installed on one side of the push rod at the bottom of the L-shaped block. Fixing blocks are vertically and symmetrically installed on both sides of the side plate against the side wall of the push rod. The fixing blocks have vertical slots. A limit block is installed at the lower end of the push rod. The limit block is placed in the slot to limit the push rod's up and down sliding. A second spring is placed between the top of the limit block and the bottom of the L-shaped block. A third motor is installed on the top of the base on the other side of the side plate. The output shaft of the third motor passes through the side plate and is fixedly connected to a first cam. The outline of the first cam can roll into contact with the bottom of the push rod.

[0007] Preferably, a medicine rack is installed on the other side of the partition plate 2. The top of the medicine rack has an array of circular holes, and the bottom of the circular holes has a through hole. A medicine bottle is placed in the circular hole. A push rod is provided directly below the through hole. The push rod can be moved to the bottom of each through hole and slide up and down to push the medicine bottle in the circular hole outward.

[0008] Preferably, the bottom of the push rod is provided with a circular groove, and two symmetrical open inclined grooves are provided on the sidewalls of the circular groove. Protrusions are installed on both sides of the rotating shaft, and the rotating shaft can extend into the circular groove. The rotating shaft can rotate in the circular groove and the top of the protrusions periodically pushes the push rod upward along the inclined groove trajectory. A motor is provided below the rotating shaft, and the output shaft of the motor is fixedly connected to the bottom of the rotating shaft. A slider is provided below the motor, and a sliding sleeve is installed on the top of the slider. The motor is installed in the middle of the top of the slider. The top of the push rod passes through the top of the sliding sleeve, and the push rod can move axially inside the sliding sleeve.

[0009] Preferably, the top of the base has a groove, and a lead screw is rotatably connected inside the groove. The lead screw is threadedly connected to the slider. A second motor is installed on one side of the groove, and the output shaft of the second motor passes through the side wall of the groove and is fixedly connected to the lead screw.

[0010] Preferably, a sleeve is provided at the lower corner of the ELISA plate, and the bottom of the sleeve is fixedly connected to the base. A spring three is installed inside the sleeve. A sliding rod is installed at the bottom corner of the ELISA plate, and the sliding rod passes through the sleeve and is fixedly connected to the spring three. A top rod two is installed in the middle of the bottom of the ELISA plate, and a fixing frame is slidably connected to the top rod two. The bottom of the fixing frame is fixedly connected to the base. A motor four is installed on one side of the top rod two at the top of the base. A cam two is fixedly connected to the output shaft of the motor four. The top rod two passes through the fixing frame and can roll into contact with the contour of the cam two. A spring four is installed between the bottom end of the top rod two and the fixing frame. The up and down movement of the top rod two can drive the ELISA plate to mix the liquid in its wells.

[0011] Preferably, a reading card is placed inside the box on the other side of the medicine holder.

[0012] Preferably, a control panel is installed on the outer wall of the box, and the control panel is electrically connected to motor one, motor two, motor three, motor four, telescopic rod, and motor five.

[0013] The present invention has the following beneficial effects: 1. This TFAM-based diagnostic and prognostic kit for ischemic stroke includes a reagent rack symmetrically and rotatably connected to the top of the base on one side of an isolation plate. The reagent rack has an array of through-holes, each containing a reagent tube. Partitions one and two are installed between the reagent racks and on the right side of the racks, respectively. A ring-shaped slider is mounted on the outer surface of the racks, rotatably connected to the side walls of the isolation plate, the housing, and partitions one and two. An L-shaped block is installed on the top of the base on one side below the reagent racks, with a push rod slidably connected to it. The push rod can push the reagent tubes in the through-holes upwards. A sealing cap is sealed at the top of the reagent tube, and a linkage mechanism is rotatably connected to the top of the sealing cap. A telescopic rod is rotatably connected to the other end of the linkage mechanism. The telescopic rod, through the linkage mechanism, can move the sealing cap closer to or away from the top of the reagent tube. The kit utilizes the symmetrical arrangement of reagent tubes on both sides... The reagent rack allows for the placement of different reagents, with partitions dividing the rack into independent, unique areas. Each reagent is labeled with a unique number near the top of its through-hole, linking the reagent tubes. The rack rotates to bring the tubes closer to the operator, and a push rod ejects the tubes to their designated positions, preventing uncertainty in reagent retrieval caused by manual handling. A control panel records used reagents, improving testing efficiency and accuracy. When the rack brings the target tube to the operator, a telescopic rod and linkage mechanism open the sealing cap. After use, the cap closes as the tube moves out of its target position, completely preventing human error, reducing reagent contamination risks, and improving diagnostic efficiency.

[0014] 2. The TFAM-based diagnostic and prognostic kit for ischemic stroke has an ELISA plate on the other side of the isolation plate. The ELISA plate can move up and down inside the box and can mix the liquid in the wells of the ELISA plate. By moving the ELISA plate up and down, the liquid in the wells of the ELISA plate is ensured to be mixed and the reaction is sufficient, which further ensures the reliability of diagnosis and prognostic assessment and improves detection efficiency and accuracy. Attached Figure Description

[0015] Figure 1 This is a schematic diagram of the structure of the present invention; Figure 2 This is a schematic diagram of the internal structure of the present invention; Figure 3 This is a schematic diagram of the mixing structure of the present invention; Figure 4 This is a schematic diagram of the opening structure of the present invention; Figure 5 This is a schematic diagram of the reagent rack driving structure of the present invention; In the diagram: 1. Box body; 2. Control panel; 3. Microplate; 4. Partition 1; 5. Reagent tube; 6. Sealing cap; 7. Reagent rack; 8. Partition 2; 9. Reagent rack; 10. Reagent bottle; 11. Interpretation card; 12. Base; 13. Reagent rack; 14. Top rod 1; 15. Spring 1; 16. Sliding sleeve; 17. Rotating shaft; 18. Lead screw; 19. Motor 1; 20. Motor 2; 21. Sliding block; 22. Annular slide. 23. Push rod; 24. Spring 2; 25. Side plate; 26. Fixed block; 27. L-shaped block; 28. Motor 3; 29. ​​Cam 1; 30. Slide rod; 31. Sleeve; 32. Spring 3; 33. Fixed frame; 34. Spring 4; 35. Top rod 2; 36. Cam 2; 37. Motor 4; 38. Telescopic rod; 40. Linkage mechanism; 41. Rotating shaft; 42. Worm gear; 43. Helical gear; 44. Motor 5. Detailed Implementation

[0016] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0017] Example 1 Please see Figures 1 to 4 This embodiment provides a TFAM-based diagnostic and prognostic kit for ischemic stroke, comprising a box 1, an isolation plate installed in the middle of the box 1, a base 12 installed at the bottom of the box 1, a reagent rack 7 symmetrically rotatably connected to one side of the isolation plate on the top of the base 12, an array of through holes on the reagent rack 7, in which reagent tubes 5 are placed, partitions 4 and 8 are installed between the reagent racks 7 and on the right side of the reagent rack 7, respectively, and an annular slider 22 is installed on the outer surface of the reagent rack 7, the annular slider 22 being positioned between the side walls of the isolation plate, the box 1, partitions 4 and 8. The reagent rack 7 is connected to an L-shaped block 27 on the top of the base 12. A push rod 23 is slidably connected to the L-shaped block 27. The push rod 23 can push the reagent tube 5 in the through hole upward. The top of the reagent tube 5 is sealed with a sealing cap 6. The top of the sealing cap 6 is rotatably connected to a linkage mechanism 40. The other end of the linkage mechanism 40 is rotatably connected to a telescopic rod 38. The telescopic rod 38 can move the sealing cap 6 closer to or away from the top of the reagent tube 5 through the linkage mechanism 40. An enzyme labeling plate 3 is set on the other side of the isolation plate. The enzyme labeling plate 3 can move up and down inside the box 1 and can mix the liquid in the well of the enzyme labeling plate 3.

[0018] Based on the above, the housing 1 is divided into a reagent and drug solution area and an enzyme-labeled plate 3 reaction area by a separator plate, thus achieving functional zoning; the reagent rack 7 can rotate 360° using a ring slider 22, which can transfer the target reagent tube 5 to the retrieval position; the push rod 23 slides to automatically eject the reagent tube 5, and the telescopic rod 38 controls the opening and closing of the sealing cap 6 through the rotational transmission of the linkage mechanism 40; the enzyme-labeled plate 3 moves up and down, using liquid inertia and impact force to achieve uniform mixing of the liquid in the wells. The enzyme-labeled plate 3 is 96 The wells are pre-coated with an ELISA plate; one reagent rack 7 holds 5 tubes of TFAM standard with concentrations of 0.1 ng / mL, 0.5 ng / mL, 2 ng / mL, 10 ng / mL, and 50 ng / mL, 1 mL / tube each; the other reagent rack 7 holds 5 tubes of 8-OHdG standard with concentrations of 0.1 ng / mL, 0.5 ng / mL, 2 ng / mL, 10 ng / mL, and 50 ng / mL, 1 mL / tube each, and contains 1% BSA. The linkage mechanism 40 consists of an active rod, a driven rod, an actuating linkage, and a fixed frame. The main drive rod is a T-shaped rod whose long side is rotatably connected to the telescopic rod 38, one end of its short side is rotatably connected to the fixed frame, and the other end is rotatably connected to the driven rod. The other end of the driven rod is rotatably connected to the top of the sealing cap 6. The middle of the driven rod is rotatably connected to one of the actuating linkages. The end of this actuating linkage is rotatably connected to another fixed frame, and the other actuating linkage is rotatably connected to the sealing cap 6. Furthermore, the functional areas are clearly defined to avoid cross-contamination between different stages; the reagent rack 7 rotates and the push rod 23 pushes out the sealing cap 6 for automatic opening and closing, eliminating the need for employees to touch the reagent tubes 5 throughout the process, reducing employee operational errors; the enzyme-labeled plate 3 automatically mixes instead of being shaken by employees, ensuring sufficient liquid reaction and improving detection accuracy.

[0019] Please see Figures 1 to 5 A rotating shaft 41 is installed in the middle of the bottom of the reagent rack 7. A helical gear 43 is installed at the lower end of the rotating shaft 41. A motor 44 is installed on one side of the helical gear 43. A worm gear 42 is fixedly connected to the output shaft of the motor 44. The worm gear 42 is meshed with the helical gear 43.

[0020] Based on the above, the motor 44 outputs power to drive the worm gear 42 to rotate. By utilizing the meshing transmission relationship between the worm gear 42 and the helical gear 43, the rotational power of the motor is transmitted to the rotating shaft 41. The rotating shaft 41 drives the reagent rack 7 to rotate synchronously, ultimately achieving the precise positioning of the reagent rack 7 (rotating to the pick-up position of the target reagent tube 5). Furthermore, the meshing transmission of worm gear 42 and helical gear 43 has the characteristics of stable transmission ratio and high positioning accuracy, which can avoid deviation or jamming when reagent rack 7 rotates; the motor drive replaces manual rotation, reduces manual operation, and at the same time ensures the consistency of positioning for each rotation, improving the accuracy of reagent dispensing.

[0021] Please see Figures 1 to 2 A side plate 25 is installed on one side of the push rod 23 at the bottom of the L-shaped block 27. Fixing blocks 26 are vertically symmetrically installed on both sides of the side plate 25 against the side wall of the push rod 23. The fixing blocks 26 have vertical slots. A limit block is installed at the lower end of the push rod 23. The limit block is placed in the slot to limit the up and down sliding of the push rod 23. A spring 24 is placed between the top of the limit block and the bottom of the L-shaped block 27. A motor 28 is installed on the top of the base 12 on the other side of the side plate 25. The output shaft of the motor 28 passes through the side plate 25 and is fixedly connected to a cam 29. The contour of the cam 29 can roll into contact with the bottom of the push rod 23.

[0022] Based on the above, motor 3 28 drives cam 1 29 to rotate, and the irregular contour of cam 1 29 will periodically push the bottom of push rod 23; with the cooperation of the slot of fixed block 26 and limit block, push rod 23 can only slide vertically (to avoid lateral displacement); when the highest point of cam 1 29 pushes push rod 23, push rod 23 slides upward to push out reagent tube 5; when the lowest point of cam 1 29 contacts push rod 23, the elastic restoring force of spring 2 24 pulls push rod 23 downward to reset, waiting for the next push-out action; Furthermore, the slot limit of the fixing block 26 ensures the precise sliding direction of the push rod 23, preventing the reagent tube 5 from tilting when it is ejected; the second spring 24 automatically resets without additional power, simplifying the structure while ensuring the continuity of the push rod 23's movement; the cam drive enables the push rod 23 to be ejected periodically and stably, replacing manual removal of the reagent tube 5, improving operational efficiency and preventing damage to the reagent tube 5.

[0023] Please see Figures 1 to 2 On the other side of the partition 2 8, a medicine rack 9 is installed. The top of the medicine rack 9 has an array of round holes, and the bottom of the round holes has a through hole. A medicine bottle 10 is placed in the round hole. A top rod 14 is set directly below the through hole. The top rod 14 can be moved to the bottom of each through hole and slide up and down to push the medicine bottle 10 in the round hole outward.

[0024] Based on the above, the drug solution bottles 10 are placed in the round holes of the drug solution rack 9 according to the detection requirements. The round holes serve to fix the drug solution bottles 10. The push rod 14 can be aligned with the through holes at the bottom of different round holes by moving laterally, and then slides vertically up and down to pass through the through holes and push the bottom of the drug solution bottle 10, finally pushing the drug solution bottle 10 out of the round hole, realizing the automatic ejection of the drug solution bottle 10. The drug solution bottles 10 in the drug solution rack 9 are: one bottle of anti-human 8-OHdG monoclonal antibody (1mg / mL, containing 0.02% sodium azide preservative), one bottle of HRP-labeled secondary antibody (10mL, dilution ratio 1:5000, anti-mouse IgG), one bottle of TMB substrate solution (10mL, containing 0.4mmol / L TMB, 0.01% H2O2), one bottle of stop solution (10mL, 2mol / L H2SO4), and one bottle of sample diluent (50mL, containing 50mmol / L Tris-HCl, 0.15mol / L H2SO4). One bottle of NaCl, 0.05% Tween-20, 1% BSA, 1 mmol / L PMSF; one bottle of washing solution (100 mL, containing 1 mol / L Tris-HCl, 3 mol / L NaCl, 1% Tween-20); Furthermore, the array of circular holes allows for the categorization and placement of different liquids, preventing confusion. The lateral movement and longitudinal ejection of the push rod 14 can accommodate all the liquid bottles in the circular holes, eliminating the need for manual handling. The through hole provides guidance for the push rod 14, ensuring that the liquid bottle 10 can be ejected smoothly.

[0025] Please see Figures 1 to 2 The bottom of the push rod 14 has a circular groove, and the side walls of the circular groove have symmetrically opened inclined grooves on both sides. The rotating shaft 17 has protrusions installed on both sides, and the rotating shaft 17 can extend into the circular groove. The rotating shaft 17 can rotate in the circular groove and cause the top of the protrusions to periodically push the push rod 14 upward along the inclined groove trajectory. The motor 19 is set below the rotating shaft 17. The output shaft of the motor 19 is fixedly connected to the bottom of the rotating shaft 17. The slider 21 is set below the motor 19. The top of the slider 21 is equipped with a sliding sleeve 16. The top of the slider 21 is equipped with the motor 19. The top of the push rod 14 passes through the top of the sliding sleeve 16, and the push rod 14 can move axially inside the sliding sleeve 16.

[0026] Based on the above, the motor 19 outputs power to drive the rotating shaft 17 to rotate. The rotation of the rotating shaft 17 will periodically push the bottom of the push rod 14, causing the push rod 14 to move upward along the radial guide of the sliding sleeve 16 (only allowing up and down sliding), so as to push out the medicine bottle 10. The motor 19 and the push rod 14 are mounted on the slider 21 as a whole, and can move laterally synchronously with the slider 21 to ensure that the push rod 14 can be aligned with the through hole at different positions. Furthermore, the radial guide of the sliding sleeve 16 prevents the push rod 14 from shifting when sliding up and down, ensuring accurate ejection action; the periodic push of the rotating shaft 17 replaces manual pressing, realizing automatic ejection of the medicine bottle 10.

[0027] Please see Figures 1 to 2 The base 12 has a groove on its top, and a lead screw 18 is rotatably connected inside the groove. The lead screw 18 is threadedly connected to the slider 21. A motor 20 is installed on one side of the groove, and the output shaft of the motor 20 passes through the side wall of the groove and is fixedly connected to the lead screw 18.

[0028] Based on the above, the second motor 20 outputs power to drive the lead screw 18 to rotate in the groove of the base 12. By utilizing the threaded meshing relationship between the lead screw 18 and the slider 21, the rotational motion of the lead screw 18 is converted into the lateral linear motion of the slider 21, which in turn drives the top rod 14 and the motor 19 at the top of the slider 21 to move laterally in sync, thereby aligning the top rod 14 with the target through hole. Furthermore, the lead screw 18 transmission features high precision and smooth operation, ensuring accurate positioning of the slider 21 during lateral movement and preventing misalignment between the push rod 14 and the through hole; the motor drive replaces manual pushing of the slider 21, reducing manual operation steps while ensuring consistency of lateral positioning each time, thus improving the efficiency of liquid dispensing.

[0029] Please see Figures 1 to 3 A sleeve 31 is provided at the lower corner of the ELISA plate 3. The bottom of the sleeve 31 is fixedly connected to the base 12. A spring 32 is installed inside the sleeve 31. A slide rod 30 is installed at the bottom corner of the ELISA plate 3. The slide rod 30 passes through the sleeve 31 and is fixedly connected to the spring 32. A top rod 35 is installed in the middle of the bottom of the ELISA plate 3. A fixing frame 33 is slidably connected to the top rod 35. The bottom of the fixing frame 33 is fixedly connected to the base 12. A motor 4 37 is installed on one side of the top rod 35 at the top of the base 12. A cam 2 36 is fixedly connected to the output shaft of the motor 4 37. The top rod 35 passes through the fixing frame 33 and can roll into contact with the contour of the cam 2 36. A spring 4 34 is installed between the bottom end of the top rod 35 and the fixing frame 33. The up and down movement of the top rod 35 can drive the ELISA plate 3 to mix the liquid in its wells.

[0030] Based on the above, motor 4 37 drives cam 2 36 to rotate, and the contour of cam 2 36 periodically pushes the bottom of push rod 2 35. Push rod 2 35 slides up and down along the guide of fixed frame 33. When push rod 2 35 moves upward, it drives the enzyme labeling plate 3 to move upward synchronously. The slide rod 30 at the corner of enzyme labeling plate 3 slides upward along sleeve 31 and compresses spring 3 32. When cam 2 36 rotates to a point other than the highest point, push rod 2 35 slides downward under the restoring force of spring 4 34, and enzyme labeling plate 3 moves downward accordingly. At the same time, the elastic force of spring 3 32 assists enzyme labeling plate 3 to reset. Through the continuous up and down reciprocating motion of enzyme labeling plate 3, the inertia and collision of the liquid in the well are used to achieve thorough mixing. Furthermore, spring 32 and spring 4 34 work together to ensure the smooth reset of the enzyme-labeled plate 3, avoiding liquid spillage caused by violent impacts during up-and-down movement; the fixing frame 33 and sleeve 31 provide dual guidance to ensure consistent horizontality of the enzyme-labeled plate 3 during up-and-down movement, preventing liquid from sloshing out of the wells; automatic mixing replaces manual shaking, solving the problems of uneven mixing force and insufficient mixing during manual mixing, and improving the stability of the detection reaction.

[0031] Please see Figure 1 On the other side of the medicine holder 9, inside the box 1, there is a reading card 11.

[0032] Based on the above, after the liquid reaction is completed in the ELISA plate 3, the detection signals (such as color changes, fluorescence intensity, etc.) in the wells of the ELISA plate 3 are read through the interpretation card 11. Combined with the relevant TFAM detection standards, the diagnosis result or prognosis of ischemic stroke is determined. The interpretation card 11 is marked with the following critical values: TFAM < 2 ng / mL indicates a poor prognosis; 8-OHdG > 5 ng / mL indicates a risk of hemorrhagic transformation. Furthermore, the interpretation card 11 is integrated with the reagent kit 1, eliminating the need for additional carrying or searching, thus improving the convenience of the testing process; the dedicated interpretation card 11 is compatible with the testing system of the reagent kit 1, ensuring the accuracy of result reading and avoiding errors caused by using a general interpretation card 11.

[0033] Please see Figures 1 to 5 A control panel 2 is installed on the outer wall of the box 1. The control panel 2 is electrically connected to motor 19, motor 20, motor 38, motor 47, telescopic rod 38, and motor 544.

[0034] Based on the above, the control panel 2, as the core control unit, sends electrical signals to each motor and telescopic rod 38 through manual operation commands to control their start, stop, speed, and telescopic range. This, in turn, links the actions of the reagent rack 7 to rotate, the reagent tube 5 to eject the medicine bottle 10, the sealing cap 6 to open and close, and the enzyme-labeled plate 3 to mix, thus achieving fully automated control of the entire process. At the same time, it can record the status of the medicine usage for easy traceability. Furthermore, centralized electronic control replaces decentralized manual operation, reducing human intervention, lowering operational difficulty, improving the accuracy of testing, and enhancing the reliability of diagnosis and prognosis assessment; the coordinated control of various components ensures the orderly conduct of the testing process, avoiding errors in the sequence of manual operations; and it improves the universality and clinical suitability of the reagent kit.

[0035] Based on the aforementioned TFAM-based diagnostic and prognostic kit for ischemic stroke, the following testing procedure is now provided: (1) Sample collection and processing: Collect 2 mL of venous blood from the patient (EDTA anticoagulation), centrifuge at 3000 rpm for 10 minutes to separate serum; if it is cerebrospinal fluid sample, take 0.5 mL directly and dilute with sample diluent at 1:10 (to avoid protease degradation of TFAM). (2) Adding samples: Add 50 μL of diluted sample / standard to each well of the microplate, and add 50 μL of sample diluent to the blank well. Incubate at 37°C for 30 minutes. (3) Washing: Discard the liquid in the well, add 200 μL of washing solution (diluted with distilled water 1:20) to each well, let stand for 30 seconds and then discard, repeat washing 3 times, and pat dry the last time; (4) Secondary antibody incubation: Add 50 μL of HRP-labeled secondary antibody to each well, incubate at 37°C for 20 minutes, and repeat step 3 for 5 washes; (5) Color development and termination: Add 50 μL of TMB substrate solution to each well, incubate at 37°C in the dark for 15 minutes, and add 50 μL of stop solution to terminate the reaction (the solution changes from blue to yellow). (6) Reading and calculation: Use an ELISA reader to read the absorbance (OD value) of each well at a wavelength of 450 nm. Plot a standard curve with the concentration of the standard as the x-axis and the OD value as the y-axis. Substitute the OD value of the sample to calculate the concentrations of TFAM and 8-OHdG. (7) Result interpretation: According to the interpretation card, TFAM≥2ng / mL and 8-OHdG≤5ng / mL indicate a good prognosis; TFAM<2ng / mL or 8-OHdG>5ng / mL indicate that stronger treatment intervention is needed.

[0036] Based on the above detection process, the following effects can be achieved: (1). Integrated diagnosis and prognosis: For the first time, TFAM and 8-OHdG were detected together, and the AUC for diagnosing ischemic stroke reached 0.92. At the same time, prognosis and bleeding risk can be assessed without the need for multiple tests; (2). High altitude adaptability: The stability of TFAM is maintained by PMSF (protease inhibitor) in the sample dilution solution. The critical value is optimized for the low expression characteristics of TFAM in high altitude patients (the critical value of TFAM in plain patients is 1.5 ng / mL, which is adjusted to 2 ng / mL in this invention), and the detection accuracy is improved to 91%. (3) Grassroots friendly: No special equipment is required (only ordinary ELISA reader is needed, and the coverage rate of grassroots hospitals is >90%), the testing time is <2 hours, and the cost of a single test is <50 yuan (only 3.3% of the cost of MRI examination). (4) High stability: The reagent kit can be stored at 4℃ for 6 months and the CV value of the standard is <8%; when placed at room temperature (25℃) for 72 hours, the deviation of the test results is <10%, which is suitable for transportation and storage in high-altitude areas.

[0037] Based on the above testing process, the following testing experiments and results are provided: 1. Reagent kit performance validation (1) Sensitivity: When the lowest concentration of TFAM standard is 0.1 ng / mL, the OD value is 0.12±0.01 (blank OD value is 0.03±0.005), the signal-to-noise ratio (S / N) is 4.0, and the sensitivity reaches 0.1 ng / mL; the lowest detection limit of 8-OHdG is the same as that of TFAM; (2) Specificity: When 10 ng / mL of interfering substances (such as IL-6, TNF-α, CRP, homocysteine) were added to the sample, the deviation rates of TFAM and 8-OHdG were 3.2% and 2.8%, respectively, with no cross-reactivity. (3) Repeatability: Ten repeated tests were performed on the same high, medium and low concentration (TFAM: 0.5 ng / mL, 2 ng / mL, 10 ng / mL; 8-OHdG: 0.5 ng / mL, 5 ng / mL, 20 ng / mL) samples. The intra-assay CV value was <5% and the inter-assay CV value was <8%. (4) Stability: After the kit was stored at 4℃ for 6 months, the OD value of the test standard deviated from the initial value by <10%; after being placed at room temperature for 72 hours, the deviation was <12%. 2. Clinical sample test results (1) Diagnostic efficacy analysis (Table 1): Group Number of examples TFAM concentration (ng / mL, x±s) 8-OHdG concentration (ng / mL, x±s) TFAM alone detects AUC (95% CI). AUC (95% CI) of 8-OHdG alone. Combined detection AUC (95% CI) High-altitude stroke group 120 1.82±0.53 6.25±1.87 0.86（0.80-0.92） 0.84（0.78-0.90） 0.93（0.89-0.97） Plain Stroke Group 80 2.56±0.71 5.12±1.53 0.84（0.76-0.92） 0.82（0.73-0.91） 0.91（0.85-0.97） healthy control group 50 4.38±1.05 1.23±0.45 - - - Conclusion: The combined detection AUC was significantly higher than that of the individual detection (P<0.05), and the combined detection AUC was the highest in the plateau group (0.93), which is suitable for the diagnosis and treatment needs in plateau areas.

[0038] (2) Prognostic correlation analysis: 200 stroke patients were followed up for 90 days and divided into a good prognostic group (mRS score 0-2, n=112) and a poor prognostic group (mRS score 3-6, n=88) according to their mRS scores. The results showed that: The TFAM concentration in the good prognosis group (2.98±0.85ng / mL) was significantly higher than that in the poor prognosis group (1.15±0.32ng / mL), and the difference was statistically significant (t=18.63, P<0.001). The concentration of 8-OHdG in the poor prognosis group (7.85±2.12ng / mL) was significantly higher than that in the good prognosis group (3.26±1.08ng / mL), and the difference was statistically significant (t=16.95, P<0.001). Using TFAM < 2 ng / mL combined with 8-OHdG > 5 ng / mL to predict poor prognosis, the sensitivity was 89%, the specificity was 85%, the positive predictive value was 87%, and the negative predictive value was 88%.

[0039] (3) Analysis of the plateau vs. plain subgroup: The TFAM concentration in the plateau stroke group was 28.9% lower than that in the plain stroke group (t=6.82, P<0.001), and the 8-OHdG concentration was 22.1% higher (t=4.35, P<0.001), which is consistent with the pathological characteristics of increased mitochondrial damage caused by high altitude hypoxia. After adjusting the interpretation threshold for the high-altitude group, the detection accuracy of this kit reached 91%, which is 9 percentage points higher than before the adjustment (82%) (χ²=6.25, P<0.05).

[0040] Three county-level hospitals in Pingliang City, Gansu Province (Zhuanglang County People's Hospital, Jingning County People's Hospital, and Chongxin County People's Hospital) were selected. This kit was used to test 50 suspected stroke patients. The results were 94% consistent with those of the tertiary hospital (Affiliated Hospital of Gansu Medical College) (Kappa=0.88, P<0.001). The proficiency score of primary care physicians (4.5±0.5 points, full score 5) was significantly higher than that of traditional 8-OHdG detection (2.8±0.7 points) (t=12.36, P<0.001).

[0041] This kit integrates early diagnosis and prognostic assessment of ischemic stroke through the combined detection of "TFAM+8-OHdG" dual biomarkers. After optimization for plateau regions, the detection accuracy reaches 91%. It is easy to operate, low in cost, and suitable for widespread use in primary hospitals, thus solving the pain points of existing detection technologies.

[0042] In summary, this TFAM-based diagnostic and prognostic kit for ischemic stroke involves first adding the diluted sample to the three wells of an ELISA plate. Then, pressing the corresponding number button on the control panel 2 sends a signal to motor 5 44, which drives the worm gear 42 to rotate. The worm gear meshes with the helical gear 43 at the bottom of the reagent holder 7, causing the rotating shaft 41 and reagent holder 7 to rotate synchronously (the annular slider 22 ensures smooth rotation) until the target reagent tube 5 is rotated to the dispensing position. Simultaneously, the telescopic rod 38 extends and retracts, pulling the sealing cap 6 upwards and opening it via the linkage mechanism 40. Then, motor 3 28 starts, driving cam 1 29 to rotate. Cam 1 29 pushes the bottom of push rod 23, which is then limited by the slot of the fixing block 26. Slide upwards to push reagent tube 5 out of the through hole of reagent rack 7; the operator can directly take the relevant TFAM standard and 8-OHdG standard reagents. After taking them, reagent rack 7 rotates to reset, and telescopic rod 38 moves in the opposite direction to close sealing cap 6, preventing reagent contamination; press the corresponding number button on control panel 2 to send a command to motor 20. Motor 2 drives lead screw 18 to rotate in the groove of base 12. Sliding slider 21, which is threaded to lead screw 18, moves laterally along the groove, driving top push rod 14, sliding sleeve 16 and motor 19 to move directly below the through hole of target medicine bottle 10; then motor 19 starts, driving rotating shaft 17 to rotate. Rotating shaft 17 pushes push rod 14, pushing... Rod 14, guided radially upward along sliding sleeve 16, passes through the through hole, pushing out the medicine bottle 10 in the round hole of medicine holder 9, thereby retrieving the relevant medicine, realizing convenient medicine retrieval and recording the number of times the medicine is retrieved and the type of medicine; after the retrieved reagent and medicine are added to the well of ELISA plate 3 in proportion, the control panel sends a signal to motor 4 37, motor 4 drives cam 2 36 to rotate, cam 2 36 pushes the push rod 2 35 to slide upward along the fixed frame 33, driving ELISA plate 3 to move upward synchronously (the sliding rod 30 at the corner of the ELISA plate slides along sleeve 31, and spring 3 32 buffers the impact); when cam 2 36 rotates to a point other than the highest point, push rod 2 35 moves downward under the restoring force of spring 4 34, and ELISA plate 3 falls back smoothly. Through continuous up-and-down reciprocating motion, the liquid in the wells is thoroughly mixed and the reaction is complete. After the reaction is complete, the integrated interpretation card 11 in the box 1 is taken out, and the enzyme-labeled plate 3 is compared with the interpretation card 11 to read the signal changes in the liquid in the wells (such as color depth, fluorescence intensity, etc.). If TFAM ≥ 2 ng / mL and 8-OHdG ≤ 5 ng / mL, it indicates a good prognosis. If TFAM < 2 ng / mL or 8-OHdG > 5 ng / mL, it indicates that stronger treatment intervention is needed. The diagnosis of ischemic stroke or the patient's prognosis assessment conclusion can be obtained quickly. The entire detection process does not require complicated manual operation, has a high degree of automation, and the detection results are accurate, which improves the reliability of diagnosis and prognosis assessment.

[0043] In the description of this invention, it should be noted that the terms "center," "upper," "lower," "left," "right," "vertical," "horizontal," "inner," and "outer," etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are used only for the convenience of describing the invention and for simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on the invention. The terms "first," "second," and "third" are used for descriptive purposes only and should not be construed as indicating or implying relative importance. Furthermore, unless otherwise explicitly specified and limited, the terms "installed," "connected," and "linked" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal connection of two components. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances. Moreover, the terms “comprising,” “including,” or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.

[0044] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A TFAM-based ischemic stroke diagnosis and prognosis evaluation kit, comprising a box body (1), an isolation plate is installed in the middle of the inside of the box body (1), and a base (12) is installed at the bottom of the box body (1), characterized in that: The isolation plate one side is symmetrically connected with reagent rack (7) on the top of base (12), reagent rack (7) is provided with array arrangement through hole, reagent tube (5) is placed in the through hole, reagent rack (7) between and reagent rack (7) right side is respectively installed with baffle one (4), baffle two (8), the outer surface of reagent rack (7) is installed with annular slide (22), annular slide (22) is placed in the side wall of isolation plate, box body (1), baffle one (4), baffle two (8) and is rotatably connected, the lower side of reagent rack (7) is installed with L type block (27) on the top of base (12), L type block (27) is slidably connected with push rod (23), the push rod (23) can be slid to push reagent tube (5) in the through hole upwards, the top of reagent tube (5) is sealed with sealing cover (6), the top of sealing cover (6) is rotatably connected with connecting rod mechanism (40), the other end of connecting rod mechanism (40) is rotatably connected with telescopic rod (38), the telescopic rod (38) can drive sealing cover (6) to be close to or away from the top of reagent tube (5) through connecting rod mechanism (40), the other side of isolation plate is provided with enzyme labeled plate (3), the enzyme labeled plate (3) can be moved up and down in the box body (1) and can mix the liquid in the hole of enzyme labeled plate (3).

2. The TFAM based diagnostic and prognostic evaluation kit for ischemic stroke according to claim 1, wherein: The bottom of the reagent rack (7) is provided with a rotating shaft (41), the lower end of the rotating shaft (41) is provided with a helical gear (43), one side of the helical gear (43) is provided with a motor five (44), the output shaft of the motor five (44) is fixedly connected with a worm (42), and the worm (42) is meshed with the helical gear (43). 3.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 2, characterized in that: The side plate (25) is installed on one side of the push rod (23) and located at the bottom of the L-shaped block (27), and the vertical symmetric fixed blocks (26) are installed on the side walls of the push rod (23) on both sides of the side plate (25). The fixed blocks (26) are vertically provided with notches, the lower end of the push rod (23) is provided with a limiting block, the limiting block is placed in the notches to limit the up and down sliding of the push rod (23), a spring two (24) is placed between the top of the limiting block and the bottom of the L-shaped block (27), the other side of the side plate (25) is provided with a motor three (28) on the top of the base (12), the output shaft of the motor three (28) penetrates through the side plate (25) and is fixedly connected with a cam one (29), and the profile of the cam one (29) is in rolling contact with the bottom of the push rod (23). 4.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 3, characterized in that: The other side of the baffle two (8) is provided with a liquid medicine rack (9), the top of the liquid medicine rack (9) is provided with an array of round holes, the bottom of the round hole is provided with a through hole, and the liquid medicine bottle (10) is placed in the round hole, the top rod one (14) is arranged below the through hole and can be moved below each through hole and slide up and down to push the liquid medicine bottle (10) in the round hole outward. 5.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 4, characterized in that, The top rod one (14) bottom is provided with a circular groove, and two symmetrical openings are provided on the side wall of the circular groove. The rotating shaft (17) is provided with a protrusion on both sides, and the rotating shaft (17) can extend into the circular groove. The rotating shaft (17) can rotate in the circular groove and make the top of the protrusion periodically push the top rod one (14) upward along the inclined groove track. The motor one (19) is arranged below the rotating shaft (17), and the output shaft of the motor one (19) is fixedly connected with the bottom of the rotating shaft (17). The sliding block (21) is arranged below the motor one (19), and the sliding sleeve (16) is arranged on the top of the sliding block (21). The motor one (19) is arranged on the top of the sliding block (21), and the top rod one (14) passes through the top of the sliding sleeve (16) and can move axially in the sliding sleeve (16). 6.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 5, characterized in that: The bottom of the base (12) is provided with a groove, and a lead screw (18) is rotatably connected in the groove. The lead screw (18) is threadedly connected with the sliding block (21), and the motor two (20) is arranged on one side of the groove. The output shaft of the motor two (20) penetrates the side wall of the groove and is fixedly connected with the lead screw (18).

7. The TFAM-based diagnostic and prognostic evaluation kit for ischemic stroke according to claim 6, characterized in that: The sleeve (31) is arranged at the lower corner of the enzyme-labeled plate (3), and the bottom of the sleeve (31) is fixedly connected with the base (12). The spring three (32) is arranged in the sleeve (31). The sliding rod (30) is arranged at the bottom corner of the enzyme-labeled plate (3), and the sliding rod (30) penetrates the sleeve (31) and is fixedly connected with the spring three (32). The top rod two (35) is arranged at the bottom of the enzyme-labeled plate (3). The fixed frame (33) is slidably connected with the top rod two (35), and the bottom of the fixed frame (33) is fixedly connected with the base (12). The motor four (37) is arranged on one side of the top rod two (35) and at the top of the base (12). The output shaft of the motor four (37) is fixedly connected with the cam two (36). The top rod two (35) penetrates the fixed frame (33) and is in rollable contact with the contour of the cam two (36). The spring four (34) is arranged between the bottom end of the top rod two (35) and the fixed frame (33). The up-and-down movement of the top rod two (35) can drive the enzyme-labeled plate (3) to mix the liquid in the holes. 8.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 7, characterized in that: The other side of the liquid rack (9) is placed with a judgment card (11) in the box body (1). 9.The TFAM-based ischemic stroke diagnosis and prognosis evaluation kit of claim 7, characterized in that: The control panel (2) is arranged on the outer side wall of the box body (1), and the control panel (2) is electrically connected with the motor one (19), the motor two (20), the motor three (28), the motor four (37), the telescopic rod (38) and the motor five (44).

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