A detection system and detection method for real-time coagulation function detection

Abstract

The present invention discloses a detection system and a detection method for real-time coagulation function detection. The detection system of the present invention drives blood droplets according to the dielectric wetting effect to test and analyze blood samples. The present invention proposes for the first time a new method for clinical coagulation function detection, which drives blood droplets through the digital microfluidics technology of the PCB board, dynamically monitors blood samples, calculates the speed and length of the droplets, calculates the coagulation time of the blood and the stiffness of the clot, and the obtained experimental results are correlated and consistent with the measurement results of commercial standard instruments, proving that the method has great application prospects. In addition, the detection system of the present invention has the advantages of miniaturization, portability, low cost, small amount of blood sample reagents, and short detection time. It can be used for immediate diagnosis occasions such as perioperative care, intensive care, and trauma emergency scenes, and has a very large application prospect in point-of-care diagnosis (POCT) occasions.

Description

Technical Field

[0001] The present invention relates to the field of biomedical engineering technology, and in particular to a detection system and a detection method for real-time coagulation function detection. Background Art

[0002] Coagulation monitoring is very important for the diagnosis of hemorrhagic and thrombotic diseases, perioperative care, emergency care, etc. At present, clinical coagulation testing is mainly based on four time-based coagulation items, which are mainly evaluated in two aspects. On the one hand, it is an endpoint test based on coagulation time, such as prothrombin time (PT), thrombin time (TT), and activated partial thromboplastin time (APTT). On the other hand, it involves biochemical functions and the measurement of the concentration of related proteins, such as coagulation factors, fibrinogen (FIB), and D-dimer.

[0003] Although these tests have been applied to routine clinical diagnosis, they still have certain limitations. First, although the mechanical properties of the clot formed during the coagulation process can greatly affect the effectiveness of coagulation performance, this aspect is rarely measured in routine coagulation tests. Studies have shown that some patients have excessive bleeding but normal coagulation due to insufficient clot stability. Secondly, the samples used in the test are plasma, and the effects of cellular components on coagulation function cannot be detected. Thirdly, most of the existing coagulation tests are performed in clinical laboratories, with long test times (>4h) and large sample consumption (~3mL). In many clinical settings, such as critical care, perioperative care, and pediatric care, patients need to be responded to in a shorter time. In addition, since multiple tests are usually performed separately, blood consumption may reach a very high level.

[0004] In order to make up for the shortcomings of the four coagulation tests, researchers proposed thromboelastograph (TEG) for coagulation testing. Although it can effectively evaluate the viscoelasticity of blood, these devices have the disadvantages of high cost, large size, and complex operation. Therefore, the development of a miniaturized, portable, and low-cost coagulation testing device can make up for the shortcomings of TEG testing and be suitable for point-of-care diagnostics (POCT).

[0005] Microfluidic technology has the advantages of small size, low sample consumption and high integration, providing a powerful tool for instant diagnosis of coagulation analysis. However, the microfluidic coagulation devices reported so far work using droplet microfluidics, acoustic waves, optics, microwell arrays, mechanical resonance, etc. Their implementation relies on complex instruments such as syringe pumps, acoustic sensors, and fluorescence microscopes, which limits their practical application. Summary of the invention

[0006] The purpose of the present invention is to overcome the above-mentioned defects and shortcomings in the prior art and to provide a detection system for real-time coagulation function detection.

[0007] The second object of the present invention is to provide application of the detection system in blood coagulation function detection.

[0008] The third object of the present invention is to provide a method for detecting coagulation function based on the detection system.

[0009] The above-mentioned object of the present invention is achieved through the following technical solutions:

[0010] A detection system for coagulation function detection comprises a single-chip control module, a high-voltage drive module, a digital microfluidic chip and a digital microscope; the single-chip control module is connected to a high-voltage drive device for providing a drive signal to the high-voltage drive module; the high-voltage drive module is connected to the digital microfluidic chip for providing a drive voltage to the digital microfluidic chip for driving blood droplets on an electrode array of an electrode layer; the digital microfluidic chip is placed under the digital microscope lens, and comprises, from bottom to top, a substrate, an electrode layer embedded in the substrate, a dielectric layer covering the electrode layer and a hydrophobic layer arranged on the dielectric layer; the digital microscope is used to monitor the movement of blood droplets on the electrode array of the digital microfluidic chip to perform coagulation analysis.

[0011] The detection system of the present invention, which includes a digital microfluidic chip, drives droplets based on the dielectric wetting effect, and can control the reciprocating motion of whole blood droplets on the electrode array through high and low level conversion, so that the blood droplets can be smoothly driven in the system, and the coagulation analysis is performed by monitoring the motion of the blood droplets, and the coagulation time and stiffness of the clot are reflected by the speed and length of the blood droplets. With the start of the coagulation process, the speed of the blood droplets gradually slows down when moving, and the dynamic curve of the speed can reflect the time required for coagulation to occur; after the blood droplets are in a solid state and stop moving, they are stretched by external forces, and the droplets still have a tendency to move, and the length of the droplets stretched can be used to reflect the stiffness of the formed clot.

[0012] Preferably, the system also includes a personal computer, whose output end is connected to the single-chip microcomputer module for sending control commands to the single-chip microcomputer module, and whose input end is connected to the digital microscope for receiving and processing the signals fed back by the digital microscope: reflecting the time required for coagulation through the dynamic curve of the blood droplet movement speed, and calculating the blood coagulation time; reflecting the stiffness of the formed clot through the stretched length of the blood droplet on the electrode array after the blood droplet becomes solid and stops moving, and calculating the stiffness of the clot.

[0013] Preferably, the system further comprises a voltage amplifier connected to the high-voltage driving module, for increasing the signal voltage of the high-voltage driving module.

[0014] Preferably, the substrate is a PCB board.

[0015] Preferably, the dielectric layer is a sealing film.

[0016] Preferably, the hydrophobic layer is made of silicone oil, which can also be used as a lubricant to reduce the driving damping of the droplets, lower the driving voltage, and reduce the evaporation of the droplets.

[0017] Preferably, the digital microfluidic chip further comprises an outermost cover plate.

[0018] Therefore, the present invention also provides the use of any of the above-mentioned detection systems in the detection of blood coagulation function.

[0019] The present invention also provides a method for detecting coagulation function based on any of the above detection systems, comprising the following steps:

[0020] S1. Obtain blood containing sodium citrate anticoagulant and drip the blood sample onto the electrode array of the digital microfluidic chip;

[0021] S2. Sending control commands to the single-chip microcomputer module through a personal computer, and the single-chip microcomputer module controls the high-voltage driving module to provide driving voltage to the electrode array of the digital microfluidic chip, driving the blood droplets to move in a fixed path, adding a coagulant during the movement, and monitoring the movement of the blood droplets on the electrode array of the digital microfluidic chip through a digital microscope to collect images;

[0022] S3. Analyze the collected images, calculate the speed and length of the blood droplets, and calculate the blood coagulation time and the stiffness of the clot.

[0023] Preferably, the method for calculating the blood coagulation time and the stiffness of the clot is: selecting or synthesizing a blank frame of only the digital microfluidic chip electrode array as the background image, and subtracting each frame with the background, and then performing edge detection, small object removal, morphological closure and filling, and shape detection on the generated image to obtain an image of only blood droplets, and then performing data processing on the blood droplets; using a fourth-order high-pass Butterworth filter with a cutoff frequency of 0.8 Hz to perform filtering analysis on the measured velocity data, and then fitting the obtained velocity data with a Boltzamann-sigmoid function, wherein the fitting function is shown in the following formula (1):

[0024]

[0025] Where Vi is the initial velocity, V f is the final velocity, t0 is the center time of the velocity decrease cycle, τ is the time constant, and the reaction time RT is defined as the time when the velocity of the blood droplet begins to slow down to 0.99V. i The time required for the blood droplet to gradually come to a standstill when the velocity is 0.01V i After that, the length of the clot and the deformation of the clot are calculated; by calculating the upper envelope and the lower envelope of the blood droplet length, and calculating the ratio of the difference to the lower envelope as the instantaneous strain, the average value of the strain is defined as the clot deformation to indicate the deformability of the clot.

[0026] Preferably, the driving voltage is 230V and the refresh rate is 1Hz, thereby achieving the purpose of low cost and low power consumption while ensuring the validity of the detection results.

[0027] Preferably, the coagulant is a 0.1-0.2M CaCl2 solution, with a volume ratio of 1:17 to the blood droplets.

[0028] Preferably, the analysis is performed at room temperature (~25°C) to reduce temperature control costs.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] The present invention discloses a detection system and a detection method for real-time coagulation function detection, wherein the detection system drives blood droplets according to the dielectric wetting effect to perform blood sample testing. The present invention proposes a new method for clinical coagulation function detection for the first time, drives blood droplets through the digital microfluidics technology of the PCB board, dynamically monitors the motion information of whole blood droplets on the digital microfluidics, collects images in real time and analyzes them, calculates the speed and length of blood droplets respectively, and reflects the coagulation time of blood and the stiffness of clots, and the obtained experimental results have good correlation and consistency with the measurement results of commercial standard instruments, and the reaction time RT can be used as some key indicators of coagulation detection, and the clot strain can represent the mechanical hardness of the clot, proving that the method has great application prospects. In addition, the detection system of the present invention has the advantages of miniaturization, portability, low cost, small amount of blood sample reagent (only 17 μL is required), short detection time (~15min), etc., and can be used for immediate diagnosis occasions such as perioperative care, intensive care, trauma emergency scene, etc., and has a very large application prospect in the instant diagnosis (POCT) occasion. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 Schematic diagram of a single-board or double-board structure of a digital microfluidic chip based on a PCB board.

[0032] Figure 2 Schematic diagram of the detection system for coagulation function detection.

[0033] Figure 3 Schematic diagram of coagulation detection technology. (a) The effect diagram of the real-time coagulation detection technology based on the digital microfluidic chip on the PCB board; (b) The motion state diagram of the blood droplet on the electrode array, t1 is the early stage of coagulation, t2 is the middle stage of coagulation, and t3 is the end stage of coagulation; (c) The processing process of the blood droplet speed and length, 1 and 3 in the figure are the original data, Figure 2 , 4 is the processed data.

[0034] Figure 4 The effect of voltage and refresh rate on the analysis experiment. (a) The movement speed and reaction time of blood droplets when the voltage is set to 200, 230, 260, and 290 V; (b) Bar graph of reaction time at different voltages; (c) Bar graph of clot deformation with voltage, data represents mean ± standard deviation, n = 3; (d) The movement speed of blood droplets when the refresh rate is set to 0.1, 1, 2, and 4 Hz; (e) Bar graph of reaction time at different refresh rates; (f) Bar graph of clot deformation with refresh rate, data represents mean and standard deviation, n = 3.

[0035] Figure 5The effects of CaCl2 concentration and temperature on the measurement results. (a) The time velocity of blood droplets after adding 0, 0.1, and 0.2M CaCl2; (b) Bar graph of reaction time versus CaCl2 concentration; (c) Bar graph of clot deformation versus CaCl2 concentration, data represent mean ± standard deviation, n = 3; (d) The time velocity of blood droplets measured at 25°C and 37°C; (e) Bar graph of reaction time at 25°C and 37°C; (f) Bar graph of clot deformation versus temperature, data represent mean ± standard deviation, n = 3.

[0036] Figure 6 The effects of procoagulants and anticoagulants on coagulation results. (a) The time velocity of blood droplets treated with procoagulant Kaolin and anticoagulant Heparin; (b) The reaction time of blood droplets treated with procoagulant Kaolin and anticoagulant Heparin; (c) The clot deformation of blood droplets treated with procoagulant Kaolin and anticoagulant Heparin.

[0037] Figure 7 Correlation analysis between experimental data and four coagulation indexes. (a) Correlation analysis between RT (reaction time) and PT (prothrombin time), Pearsom's correlation coefficient is 0.89; (b) Correlation analysis between strain (Stain) and FIB, Pearsom's correlation coefficient is -0.89. DETAILED DESCRIPTION

[0038] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0039] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0040] Example 1 Preparation of digital microfluidic chip based on PCB board

[0041] like Figure 1As shown, a digital microfluidic chip based on a PCB board adopts a single-board or double-board structure; the single-board structure includes, from bottom to top, a substrate, an electrode layer embedded in the substrate, a dielectric layer covering the electrode layer, a hydrophobic layer arranged on the dielectric layer, and a zero electrode in the hydrophobic layer; the double-board structure consists of an upper and lower electrode plate, with a gap between the upper and lower electrode plates; the lower electrode plate includes, from bottom to top, a substrate, an electrode layer embedded in the substrate, a dielectric layer covering the electrode layer, a hydrophobic layer arranged on the dielectric layer, and a zero electrode in the hydrophobic layer; the upper electrode plate includes, from bottom to top, a hydrophobic layer and ITO glass. A PCB board is used as a substrate, an electrode layer is made of immersion gold, a sealing film is used as a dielectric layer, and silicone oil is used as a hydrophobic layer. The digital microfluidic chip sets an electrode layer on the PCB board substrate, and a layer of dielectric material with good hydrophobic properties is coated on the surface of the electrode layer to prevent the electrode from directly contacting the conductive droplets, avoiding electrochemical reactions between the electrode and the droplets, and then setting a hydrophobic layer to ensure the smoothness and stability of the droplet movement process. The PCB-based digital microfluidic chip drives droplets based on the dielectric wetting effect. Voltage is applied to the electrode array of the electrode layer embedded under the dielectric layer to change the wetting characteristics between the dielectric layer and the conductive droplets attached to its surface, so that the liquid-solid contact angle changes, causing asymmetric deformation at both ends of the droplet, prompting a pressure difference to be generated inside the droplet, thereby realizing the movement of the droplet.

[0042] Example 2 Detection system and detection method for coagulation function detection

[0043] The PCB-based digital microfluidic chip of the above-mentioned Example 1 is combined with an image collection and processing platform to build a detection system for coagulation function detection. The blood droplets are driven back and forth on the electrode array by voltage, and the movement of the blood is monitored to perform coagulation analysis. Image processing and data output are performed through software such as MATLAB 2020a and ORIGIN 2020.

[0044] The detection system for coagulation function detection is as follows Figure 2As shown, it includes a personal computer, a single-chip control module, a high-voltage drive module, a high-voltage amplifier, a digital microfluidic chip based on a PCB board, and a digital microscope. The output end of the personal computer is connected to the single-chip module for sending control commands to the single-chip, and the input end of the personal computer is connected to the digital microscope for receiving the signal fed back by the digital microscope for processing; the single-chip is connected to the high-voltage drive module for providing a drive signal to the high-voltage drive module; the high-voltage drive module is connected to the digital microfluidic chip for providing a drive voltage for driving the blood droplets on the electrode array to the digital microfluidic chip; the signal amplifier is connected to the high-voltage drive module for increasing the signal voltage of the high-voltage drive module; the digital microfluidic chip based on the PCB board is placed under the digital microscope lens; the digital microscope is used to monitor the movement of blood droplets on the digital microfluidic chip for coagulation analysis.

[0045] According to the dielectric wetting effect of the detection system, blood droplets are driven to move back and forth on the electrode array of the digital microfluidic chip by voltage, and the movement of blood is monitored to perform coagulation analysis, such as Figure 3 (a) shows that there are three different stages in the movement of blood droplets, such as Figure 3 As shown in (b), it is divided into a flow stage, a transition stage, and a stable stage. In the flow stage, the coagulation reagent CaCl2 has just been added to start the coagulation process. The viscosity of the blood droplets is relatively low, and the blood droplets can move smoothly. After a few minutes of movement, the viscosity of the blood droplets increases and the speed gradually decreases until the blood droplets become clots and stop moving. This stage is called the transition stage. After the blood droplets stop moving, although the blood droplets basically have no displacement, they are still stretched by the electric field force. This stage is called the stable stage. The dynamic curve of the speed can reflect the time required for coagulation to occur; the length of the blood droplets stretched by the electric field force can be used to reflect the stiffness of the formed clot.

[0046] Specifically, the steps of using the detection system to perform detection are as follows:

[0047] S1. Blood containing sodium citrate anticoagulant obtained from the hospital is first dripped onto the electrode array of the digital microfluidic chip;

[0048] S2. Sending control commands to the single-chip microcomputer module through a personal computer, and the single-chip microcomputer module controls the high-voltage driving module to provide driving voltage to the electrode array of the digital microfluidic chip, and then drives the blood droplets to move in a fixed path, adding a coagulant during the movement, and monitoring the movement of the blood droplets on the electrode array of the digital microfluidic chip through a digital microscope to collect images;

[0049] S3. Analyze the collected images, use the program written in MATLAB and DMV software to analyze the experimental video, calculate the speed and length of the droplet, and calculate the blood coagulation time and the stiffness of the clot: In the DMV software, select or synthesize a blank frame of only the digital microfluidic chip electrode array as the background image, and subtract each frame with the background, and then perform edge detection, small object removal, morphological closure and filling, and shape detection on the generated image to obtain an image of only blood droplets, and then perform data processing on the blood droplets; in order to obtain the size of the movement speed, use a fourth-order high-pass Butterworth filter with a cutoff frequency of 0.8Hz to filter and analyze the speed data measured in the DMV. Figure 3 (c) is shown, and then the obtained speed data is fitted with the Boltzamann-sigmoid function, and the fitting function is shown in the following formula (1):

[0050]

[0051] Where V i is the initial velocity, V f is the final velocity, t0 is the center time of the velocity decrease cycle, τ is the time constant, and the reaction time (RT) is defined as the time when the velocity of the blood droplet begins to slow down to 0.99V. i The time required for the blood droplet to gradually come to a standstill when the velocity is 0.01V i After that, the length of the clot and the deformation of the clot are calculated; by calculating the upper envelope and the lower envelope of the blood droplet length, and calculating the ratio of the difference to the lower envelope as the instantaneous strain, the average value of the strain is defined as the clot deformation to indicate the deformability of the clot.

[0052] Example 3 Optimization and verification of coagulation function test method

[0053] Based on the detection system and detection method for coagulation function detection described in Example 2, the hardware part of the detection system and the biochemical parameters of the analysis experiment were optimized, and the effects of driving voltage, refresh rate, temperature, coagulant and anticoagulant on the coagulation experiment were explored.

[0054] 1. Experimental parameter optimization

[0055] (1) Regarding the hardware settings of the digital microfluidic chip on the PCB board, the voltage of the electrode and the refresh rate of the voltage signal are the main parameters affecting the measurement. The voltage will affect the electric field strength, thereby affecting the driving force of the blood droplet movement, which may have a certain impact on the coagulation experiment. The voltage was systematically adjusted and coagulation experiments were carried out. The results showed that when the voltage was lower than 200V, the blood droplets could not move normally on the electrode array due to insufficient driving force. When the voltage was higher than 200V, the droplet velocity increased with the increase of voltage (such as Figure 4 a), which may be because the higher the voltage, the more obvious the dielectric wetting effect. However, according to the reaction time (RT) and clot deformation, the voltage value had no significant effect on the progress of the coagulation experiment and the deformation of the generated clot, which had no significant difference at each voltage (as shown in Figure 4 b, as shown in 4c).

[0056] (2) The refresh rate of the electrode voltage determines the duration of each voltage signal and, therefore, the duration of each blood drop displacement. At higher refresh rates, the blood drop may move faster and, therefore, experience higher shear rates, which may affect the progression of the coagulation reaction and the hardness of the resulting clot. The refresh rate was set to 0.5, 1, 2, and 4 Hz and coagulation experiments were performed. As the voltage switching rate increased, the expected increase in velocity amplitude was seen ( Figure 4 d). There was no significant difference in the reaction time calculated according to each refresh rate, indicating that the coagulation trend may not be significantly affected ( Figure 4 e). However, as the refresh rate increased, an increase in clot deformation was observed ( Figure 4 f). For example, when the refresh rate is 0.5 Hz, the clot deformation is (3.9 ± 0.9)%, which is significantly less than (5.6 ± 0.6)% at 4 Hz. These results indicate that the analysis results may be less dependent on the voltage setting and the voltage refresh rate. For the sake of low cost and low power consumption, a driving voltage of 230 V and a refresh rate of 1 Hz were selected for subsequent experiments.

[0057] (3) Regarding the biochemical setting of the analysis, the concentration of CaCl2 and the experimental temperature are the parameters that directly affect the measurement. The effect of CaCl2 solution on the coagulation of blood droplets was examined by adding 0, 0.1, and 0.2 M CaCl2 solutions. CaCl2 was added to blood droplets at a volume ratio of 1:17 and analyzed. In the absence of CaCl2, blood droplets containing sodium citrate continued to move on the electrode array without coagulation (e.g. Figure 5a). When 0.1M CaCl2 was added, the velocity began to decrease 170s after the start of the experiment, indicating that the interruption of blood droplet motion was caused by blood coagulation. In contrast, the addition of 0.2M CaCl2 subsequently triggered a decrease in velocity with a reaction time of approximately 230s, which is consistent with previously reported work (e.g. Figure 5 b, 5c). In terms of deformability, both 0.1 M and 0.2 M showed about 5% clot deformation, with no significant difference. These results suggest that CaCl2 should be standardized in subsequent experiments to ensure reproducible measurements.

[0058] (4) Temperature affects enzyme activity and thus has a significant impact on the coagulation reaction. The analysis was performed at 25°C and 37°C, representing room temperature and body temperature, respectively. As expected, the coagulation process at 37°C was much faster than at 25°C, with a measured reaction time of 41.3±10.1s, significantly less than 239.7±21s (Fig. Figure 5 e). However, tracking of velocity magnitude showed similar kinetics, indicating that low temperature only delayed the reaction time without changing the trend. In contrast, there was no significant difference in clot deformability (e.g. Figure 5 f), the calculated clot deformation at 37 °C was (5.0 ± 0.7)%, and the calculated clot deformation at 25 °C was (4.6 ± 0.7)%. These results suggest that the analysis can be performed at 25 °C to reduce the cost of temperature control at the trade-off of prolonged analysis time.

[0059] 2. Test results verification

[0060] In order to verify whether the detection system and detection method for coagulation function detection described in Example 2 can distinguish between hypercoagulable and hypocoagulable blood samples, blood samples were pretreated with a coagulant kaolin (Kaolin, 5 mg / mL) or an anticoagulant heparin (Heparin, 8 U / mL) as simulated clinical samples. The results showed that the blood samples pretreated with kaolin showed an accelerated coagulation process (e.g. Figure 6 a, 6b), the reaction time was 53.7±28.0s, which was significantly longer than the control group 171.5±51.6s. Although Kaolin treatment caused changes in the coagulation process, the generated clots showed similar deformability (e.g. Figure 6 c), clot deformation in the Kaolin experimental group and the control group was (7.0±1.2)% and (9.0±4.8)%, respectively. Although Kaolin accelerated clot formation, it had little effect on clot stability, which is consistent with the reported results. In contrast, in the Heparin-treated samples, blood continued to move throughout the analysis without showing deceleration and clot formation, which is consistent with the strong anticoagulant effect of Heparin.

[0061] In order to further verify the clinical relevance of the detection system and detection method for coagulation function detection described in Example 2, a coagulation experiment was performed under the set parameters, and the reaction time (RT) and strain (Strain) measured in the experiment were correlated with PT (prothrombin time) and FIB (fibrinogen concentration) in the four clinical coagulation items. PT evaluates the coagulation trend of the blood sample, which should be related to the RT (reaction time) in the experiment of the present invention. FIB measures the concentration of fibrinogen in the blood and affects the stability of the generated thrombus, which should be related to the Strain measured in the experiment of the present invention. The analysis results are as follows: Figure 7 shown.

[0062] Eleven patient samples were tested using the detection system and detection method of Example 2, and commercial standard instruments were used to measure thrombin time (PT) and fibrinogen concentration (FIB). Correlation analysis showed that the RT value in the experiment had a strong correlation with the PT in the four clinical coagulation items, and the Pearson correlation coefficient was 0.89019, indicating that there was a strong positive correlation between the two (e.g. Figure 7 a), which indicates that the RT measured by the present invention can be used as some key indicators of coagulation detection. At the same time, the strain measured has a strong correlation with the FIB of the four clinical coagulation items, and the Pearson's correlation coefficient is -0.89652, indicating that there is a strong negative correlation between the two (such as Figure 7 b), the clot strain measured in the analysis of the present invention can indeed represent the mechanical hardness of the clot, indicating that the present invention has successfully developed a new clinical instant coagulation diagnostic technology with the advantages of low cost, portability, and rapid detection time, and can be used in perioperative care, intensive care, trauma emergency scenes and other instant diagnosis occasions.

Claims

1. A method for detecting coagulation function, characterized in that: The steps include: S1. Obtain blood containing sodium citrate anticoagulant and drip the blood sample onto the electrode array of the digital microfluidic chip; S2. Sending control commands to the single-chip microcomputer module through a personal computer, and the single-chip microcomputer module controls the high-voltage driving module to provide driving voltage to the electrode array of the digital microfluidic chip, driving the blood droplets to move in a fixed path, adding a coagulant during the movement, and monitoring the movement of the blood droplets on the electrode array of the digital microfluidic chip through a digital microscope to collect images; S3. analyzing the acquired images, calculating the velocity and length of the blood droplets, and thereby calculating the blood coagulation time and the stiffness of the clot; The method for calculating the coagulation time of blood and the stiffness of clots is as follows: selecting or synthesizing a blank frame of only the digital microfluidic chip electrode array as a background image, and subtracting each frame with the background, and then performing edge detection, small object removal, morphological closure and filling, and shape detection on the generated image to obtain an image of only blood droplets, and then performing data processing on the blood droplets; using a fourth-order high-pass Butterworth filter with a cutoff frequency of 0.8 Hz to perform filtering analysis on the measured speed data, and then fitting the obtained speed data with a Boltzamann-sigmoid function, wherein the fitting function is shown in the following formula (1): Where V i is the initial velocity, V f is the final velocity, t0 is the center time of the velocity decrease cycle, τ is the time constant, and the reaction time RT is defined as the time when the velocity of the blood droplet begins to slow down to 0.99V. i The time required for the blood droplet to gradually come to a standstill when the velocity is 0.01V i After that, the length of the clot and the deformation of the clot are calculated; by calculating the upper envelope and the lower envelope of the blood drop length, and calculating the ratio of the difference to the lower envelope as the instantaneous strain, the average value of the strain is defined as the clot deformation to indicate the deformability of the clot; The detection system used in the coagulation function detection method includes a single-chip control module, a high-voltage drive module, a digital microfluidic chip and a digital microscope; the single-chip control module is connected to the high-voltage drive module to provide a drive signal to the high-voltage drive module; the high-voltage drive module is connected to the digital microfluidic chip to provide a drive voltage to the digital microfluidic chip to drive blood droplets on the electrode array of the electrode layer; the digital microfluidic chip is placed under the digital microscope lens, and from bottom to top it includes a substrate, an electrode layer embedded in the substrate, a dielectric layer covering the electrode layer, and a hydrophobic layer arranged on the dielectric layer; the digital microscope is used to monitor the movement of blood droplets on the electrode array of the digital microfluidic chip to perform coagulation analysis.

2. The method for detecting coagulation function according to claim 1, characterized in that: The substrate is a PCB board.

3. The method for detecting coagulation function according to claim 1, characterized in that: The dielectric layer adopts a sealing film.

4. The method for detecting coagulation function according to claim 1, characterized in that: The hydrophobic layer adopts silicone oil.

5. The method for detecting coagulation function according to claim 1, characterized in that: The driving voltage is 230V and the refresh rate is 1Hz.

6. The method for detecting coagulation function according to claim 1, characterized in that: The coagulant is a 0.1-0.2M CaCl2 solution, and the volume ratio of the solution to the blood droplets is 1:17.

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