A whole blood quantitative platelet aggregation ability detection reagent card

Abstract

The present invention belongs to the field of medical examination, and particularly relates to a whole blood quantitative platelet aggregation ability detection reagent card, the reagent card comprises an injection port, a temporary storage bin, a microfluidic channel, a main liquid separation guide groove, a sample flow channel and more than two detection bins; the injection port is used to fix the blood collection tube, the blood collection needle passes through the blood collection tube from the bottom, and guides the sample to the temporary storage bin through the connecting tube at the tail of the blood collection needle; each detection bin is provided with a sample flow channel; a blood-absorbing cotton is provided at a specified height on the inner side wall of the temporary storage bin; the sample to be tested in the temporary storage bin passes through the microfluidic channel, the main liquid separation guide groove and the sample flow channel in turn into each detection bin, and the detection reagent in the detection bin dissolves in the sample to be tested and reacts immediately. The present invention can meet the clinical rapid detection; the sample flow is accurately controlled; the detection process is simplified, and only one tube of whole blood is drawn, and the results of different activation pathways of platelet aggregation function can be obtained without pre-treatment.

Description

Technical Field

[0001] The invention belongs to the field of medical testing, and particularly relates to a whole blood quantitative platelet aggregation ability detection reagent card. Background Art

[0002] Most of the platelet aggregation detection reagents currently used in the market require centrifugation before testing using platelet-containing plasma. Because the density difference between platelets and red blood cells is small, and the platelet content varies greatly among different populations (125-350×10 9 / L), it is difficult to achieve standardized platelet-poor plasma and platelet-rich plasma through centrifugation, which in turn affects the corresponding test results.

[0003] The reagent cards currently on the market include a sample chamber, a temporary storage chamber and less than four detection chambers. Due to the small number of detection chambers, in order to deal with different activation pathways on platelets, multiple blood draws or a large amount of blood samples need to be drawn at one time. Patients with cardiovascular diseases are more concerned about the amount of blood drawn at one time, and for operators, the tediousness of their work increases. In addition, the preparation and detection processes take a long time and cannot meet the clinical immediacy requirements; and the blood sample flow cannot be accurately controlled. Summary of the invention

[0004] In view of the defects of the prior art, the present invention proposes a whole blood quantitative platelet aggregation ability detection reagent card. The patent proposes integration, multiple detection chambers and the application of microfluidic technology to platelet aggregation ability detection, which can realize the direct detection of whole blood samples and the blood sample flow is accurately controlled; the detection process is simplified, and only one tube of blood is needed to obtain the desired results of different activation pathways of platelet aggregation function.

[0005] The present invention is achieved in that:

[0006] A whole blood quantitative platelet aggregation ability detection reagent card, the reagent card comprises a sample inlet, a temporary storage chamber, a microfluidic channel, a main liquid separation guide groove, a sample flow channel and N detection chambers, N is a positive integer, and N ≥ 2;

[0007] The sample inlet is used to fix the blood collection tube, the blood collection needle passes through the blood collection tube from the bottom, and guides the sample to the temporary storage bin through the connecting tube at the tail of the blood collection needle; each detection bin is provided with a sample flow channel; a blood-absorbing cotton is provided at a specified height on the inner wall of the temporary storage bin;

[0008] The microfluidic channel is provided with a first and a second vent hole, the first vent hole is connected to a one-way valve; the second vent hole is respectively connected to a No. 1 solenoid valve and a No. 2 solenoid valve through an air path, the No. 1 solenoid valve and the No. 2 solenoid valve are connected in parallel, and the No. 1 solenoid valve is connected to an air inlet of a vacuum pump, and the No. 2 solenoid valve is connected to an air outlet of the vacuum pump;

[0009] All the test chambers have a common ventilation slot on the top, and a main exhaust hole connected to the outside is provided at the center of the ventilation slot; each test chamber is connected to the ventilation slot through an exhaust slot, and exhaust holes and test cotton facing the exhaust holes are provided at the connection between the exhaust slot and the ventilation slot; there are gaps between the test cotton and the top and bottom of the exhaust slot for air circulation;

[0010] One of the temporary storage chamber or the N detection chambers is a blank control, and no lyophilized agent is placed therein; one or more of the remaining detection chambers are placed with a designated lyophilized agent, and the lyophilized agent includes a platelet activator and a detection result amplifier;

[0011] The samples to be tested in the temporary storage chamber enter into each detection chamber in turn through the microfluidic channel, the main liquid separation guide groove and the sample flow channel, and the detection reagent in the detection chamber dissolves in the samples to be tested and then reacts.

[0012] Furthermore, light detection channels are provided on opposite sides of the temporary storage bin and each detection bin.

[0013] Furthermore, the method for detecting platelet aggregation ability by the reagent card is:

[0014] 1) Calculate the platelet aggregation rate in each test chamber containing platelet activator and test result amplifier using the supporting software;

[0015] 2) The blank control is used as a transmittance reference, which is used for pre-sampling of the instrument optical path and as a reference indicator for calculation in step 1);

[0016] The supporting software algorithm adopts the mathematical model fitting method: first, the platelet aggregation reaction process is established with time t as the independent variable and light transmittance as the dependent variable. According to the Lambert-Beer law and the change of microsphere concentration, the algorithm formula is fitted.

[0017] Furthermore, the concentration of the ADP curve is recorded as C ADP ,C ADP =a*lg(I Blood / I ADP );

[0018] Its maximum concentration is recorded as C ADP.MAX ,C ADP.MAX =a*lg(I Blood / I ADP.MIN );

[0019] The concentration of the TRAP curve is recorded as C TRAP ,C TRAP =a*lg(I Blood / I TRAP );

[0020] Its maximum concentration is recorded as C TRAP.MAX ,C TRAP.MAX =a*lg(I Blood / I TRAP.MIN );

[0021] The concentration of PLA+PLC curve is recorded as C 2P , C 2P =a*lg(I Blood / I 2P );

[0022] Its maximum concentration is recorded as C 2P.MAX ,C 2P.MAX =a*lg(I Blood / I 2P.MIN );

[0023] The concentration of the COL curve is recorded as C COL ,C COL =a*lg(I Blood / I COL );

[0024] Its maximum concentration is recorded as C COL.MAX ,C COL.MAX =a*lg(I Blood / I COL.MIN );

[0025] The concentration of COX-1 curve is recorded as C AA ,C AA =a*lg(I Blood / I AA );

[0026] Its maximum concentration is recorded as C AA.MAX ,C AA.MAX =a*lg(I Blood / I AA.MIN );

[0027] The concentration of blood is recorded as C Blood =a*lg(I Blood / I Blood )=0;

[0028] The concentration of Blood+Mic is recorded as C Blood+Mic =a*lg(I Blood / I Blood+Mic );

[0029] The temporary storage or testing chamber where the blood is located corresponds to the maximum transmittance I Blood , which has the lowest concentration;

[0030] The transmittance of the detection chambers where TRAP, ADP, PLA+PLA, COL and COX-1 are located is I TRAP ,I ADP ,I 2P ,I COL and I AA The minimum transmittance of each detection chamber is I TRAP.MIN ,I ADP.MIN ,I 2P.MIN ,I COL.MIN and I AA.MIN ;

[0031] The minimum light transmittance of the Blood+Mic detection chamber is I Blood+Mic , which has the highest concentration;

[0032] Based on the above, the reagents are tested according to the configuration of the reagent card test chamber:

[0033] Platelet aggregation response unit PU of P2Y12 receptor activation pathway P2Y12 :

[0034] PU P2Y12 = a*[(C MAX -C ADP ) / (C MAX -C Blood )]+b;

[0035] C MAX = C Blood+Mic or C ADP.MAX ;

[0036] Platelet aggregation response unit PU BASE :

[0037] PU BASE =a*[(C MAX -C TRAP ) / (C MAX -C Blood )]+b;

[0038] C MAX = C Blood+Mic or C TRAP.MAX ;

[0039] Platelet aggregation response unit AU of cyclooxygenase-1 activation pathway AA :

[0040] AU AA =a*[(C MAX -C AA ) / (C MAX -C Blood )]+b;

[0041] C MAX = C Blood+Mic or C AA.MAX ;

[0042] Platelet aggregation response unit CU COL :

[0043] CU COL =a*[(C MAX -C COL ) / (C MAX -C Blood )]+b;

[0044] C MAX = C Blood+Mic or C COL.MAX ;

[0045] Platelet aggregation response unit PU of phospholipase A and phospholipase C activation pathway 2P :

[0046] PU 2P =a*[(C MAX -C 2P ) / (C MAX -C Blood )]+b;

[0047] C MAX = C Blood+Mic or C 2P.MAX ;

[0048] In the above formula, a and b are constant terms; C MAX It is a variable, which indicates the maximum concentration reference parameter used when calculating platelet aggregation reaction units, as described below:

[0049] 1) One of the test chambers of the reagent card is only equipped with a test result amplifier, and its test corresponding concentration is C Blood+Mic ; When calculating the platelet aggregation units of other test chambers, this is used as the maximum concentration reference parameter C MAX ;

[0050] 2) All test chambers of the reagent card are equipped with platelet activators and test result amplifiers at the same time; at this time, the platelet aggregation unit of the test chamber is calculated, and the maximum test concentration of each test chamber is used as the reference parameter C MAX .

[0051] Furthermore, a stirring device for stirring samples is placed in each detection chamber.

[0052] Furthermore, the stirring device includes a limit baffle fixed horizontally on the inner wall of the detection chamber and a steel ball at the bottom thereof.

[0053] Furthermore, there is a gap between the limit baffle and the inner wall of the detection chamber to facilitate the flow of samples; the width of the gap is smaller than the diameter of the steel ball.

[0054] Furthermore, the platelet activator is selected from arachidonic acid, adenosine diphosphate, collagen, adrenaline, 5-hydroxytryptamine, prostaglandin, thromboxane A 2、 Any one of ristocetin, phospholipase A, phospholipase C, thrombin, isothrombin, and isothrombin activating peptide;

[0055] The detection result amplifier is selected from dyed latex microspheres, the surface of which is coated with fibrinogen.

[0056] Furthermore, the reagent card is a 4-channel reagent card, that is, N=4; any one of the detection chambers is a blank control, without any lyophilized agent placed therein; and the remaining 1 to 3 detection chambers are placed with lyophilized agents.

[0057] Furthermore, the platelet activator in the lyophilized agent is selected from any one of arachidonic acid, adenosine diphosphate and isothrombin activating peptide.

[0058] The technical effects of the present invention are as follows:

[0059] 1. The reagent card of the present invention comprises N (N≥2) detection chambers, each of which contains a platelet activator different from that in other detection chambers, so that a small amount of blood can be drawn at one time to complete the activity detection of multiple platelet activation channels, meeting clinical needs and providing a more comprehensive, accurate, rapid and convenient guidance scheme for clinical diagnosis and treatment.

[0060] 2. The liquid path structure of the reagent card of the present invention is combined with the vacuum pump, gas path and corresponding solenoid valve of the platform to ensure that the sample to be tested can enter the test chamber synchronously and remain in an unactivated state, so as to achieve precise control of the flow rate;

[0061] 3. Scan the information code label on the reagent card through the identification module of the instrument, and then select the detection type and obtain the corresponding standard curve based on the information obtained;

[0062] 4. Automatic extraction and addition of blood samples greatly improves the automation of detection and reduces the errors and time costs caused by human participation;

[0063] 5. A stirring device is set inside the detection chamber. The magnet on the output shaft of the detection motor drives the steel ball to move horizontally, so that the reaction liquid in the detection chamber can be evenly stirred, thereby improving the detection accuracy.

[0064] In summary, the reagent card of the present invention can be used in conjunction with a platelet aggregation ability detector to achieve rapid, efficient and accurate detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0065] Figure 1 The schematic diagram of the platelet aggregation ability detection platform of the present invention;

[0066] Figure 2 It is a structural schematic diagram of a four-channel reagent card of the present invention;

[0067] Figure 3 The algorithm model diagram used by the detector of the present invention;

[0068] Figure 4 This is a schematic diagram of the working principle of the reagent card identification module of the present invention;

[0069] Figure 5 This is a schematic diagram of the working principle of the automatic sampling module of the present invention;

[0070] Figure 6 This is a schematic diagram of the working principle of the first infrared sensor receiving module of the present invention;

[0071] Figure 7 This is a schematic diagram of the working principle of the second infrared sensor receiving module of the present invention;

[0072] Figure 8 It is a schematic diagram of the working principles of the temperature control module, the motor stirring module and the optical path detection module of the present invention;

[0073] Fig. 9 is a side view of the reagent card of the present invention;

[0074] Fig.10 Schematic diagram of the working principle of the calibration module of the present invention.

[0075] In the figure: 1. Reagent card; 2. Temporary storage chamber; 2a. Blood-absorbing cotton; 2b. First vent; 2c. Second vent; 3. Microfluidic channel; 4. Detection chamber; 4a. Detection cotton; 4b. Limit baffle; 4c. Steel ball; 5. Blood collection tube; 6. Information code label; 7. Sample injection port; 8. Light detection channel; 9. Sample flow channel; 10. Main liquid separation guide groove; 11. Ventilation groove; 12. Main exhaust hole; 13. Exhaust groove; 14. Exhaust hole. DETAILED DESCRIPTION

[0076] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0077] like Figures 1 to 10As shown, the present invention discloses a reagent card for quantitative platelet aggregation ability detection of whole blood, wherein the reagent card 1 comprises a sample inlet 7, a temporary storage chamber 2, a microfluidic channel 3, a main liquid separation guide groove 10, a sample flow channel 9 and N detection chambers 4, where N is a positive integer and N≥2.

[0078] The sample inlet 7 is used to fix the blood collection tube 5. The blood collection needle passes through the blood collection tube 5 from the bottom, and guides the sample to the temporary storage chamber 2 through the connecting tube at the tail of the blood collection needle. Each detection chamber 4 is provided with a sample flow channel 9. The sample in the temporary storage chamber 2 enters the detection chamber 4 in turn through the microfluidic channel 3, the main liquid separation guide groove 10 and the sample flow channel 9.

[0079] The microfluidic channel 3 is provided with a first and a second vent hole 2c, wherein the first vent hole 2b is connected to a one-way valve; the second vent hole 2c is respectively connected to a No. 1 solenoid valve and a No. 2 solenoid valve through an air path, the No. 1 solenoid valve and the No. 2 solenoid valve are connected in parallel, and the No. 1 solenoid valve is connected to the air inlet of the vacuum pump, and the No. 2 solenoid valve is connected to the air outlet of the vacuum pump;

[0080] A blood-absorbing cotton 2a is provided at a specified height on the inner wall of the temporary storage bin 2. When the blood-absorbing cotton 2a contacts the sample, it will change color. The identification module detects this change and sends a signal to the MCU slave processing module, and then controls the opening and closing of the vacuum pump, the No. 1 solenoid valve, the No. 2 solenoid valve and the one-way valve through the control circuit. When a negative pressure environment is required in the temporary storage bin 2 to extract the sample in the blood collection tube 5, the one-way valve is closed, and no air flows into the first vent 2b; the No. 2 solenoid valve is closed, the No. 1 solenoid valve is turned on, and the vacuum pump controls the No. 1 solenoid valve to extract the air in the temporary storage bin 2 from the second vent 2c; when the sample in the temporary storage bin 2 reaches the height of the blood-absorbing cotton, the infrared sensor detected by the identification module will send a detection signal to the MCU slave processing module, the No. 1 solenoid valve is closed, the No. 2 solenoid valve is turned on, and the air enters the temporary storage bin 2 through the second vent 2c; the one-way valve is opened, and as the sample is injected into each detection bin 4 through the microfluidic channel 3, the air is discharged through the first vent 2b.

[0081] A ventilation groove 11 is provided on the top of all the detection chambers 4, and a general exhaust hole 12 connected to the outside is provided at the center of the ventilation groove 11; each detection chamber is connected to the ventilation groove 11 through an exhaust groove 13, and an exhaust hole 14 and a detection cotton 4a facing the exhaust hole are provided at the connection between the exhaust groove 13 and the ventilation groove 11; there is a gap between the detection cotton and the top and bottom of the exhaust groove for air circulation;

[0082] The air in each detection chamber is discharged to the outside through the exhaust holes 14 and the main exhaust holes 12. When the sample in the detection chamber is full and the air is exhausted, the sample will enter the exhaust groove until it contacts the detection cotton 4a. The bottom of the detection cotton will expand rapidly when it contacts the sample, closing the exhaust holes. The detection chamber cannot exhaust the gas and cannot continue to inject samples, but there is a gap between the top of the detection cotton and the exhaust groove, which does not affect the gas flow of other detection chambers. Therefore, even if the middle detection chamber is filled first, it will not affect the gas discharge of other detection chambers. At the same time, the color of the expanded detection cotton will change; when the detection cotton of all detection chambers is expanded and discolored, the infrared sensor detected by the identification module will send the signal of the detection cotton discoloration to the MCU slave processing module, stop the power supply of the vacuum pump, the No. 1 solenoid valve and the No. 2 solenoid valve, and complete the injection of all detection chambers. Since the internal volume of each detection chamber is the same and it will be filled with samples, the flow rate of each detection chamber is ensured to be the same.

[0083] In order to facilitate uniform mixing of the sample and the platelet activator and thorough reaction, a stirring device is placed in each detection chamber 4. A magnet is provided on the transmission shaft of the detection motor on the detector. When the motor rotates, it can drive the stirring device in the detection chamber 4 to move along a certain trajectory, thereby stirring the reaction.

[0084] Preferably, light detection channels 8 are provided on opposite sides of the temporary storage bin 2 and each detection bin 4 to facilitate the passage of light.

[0085] Preferably, an information code label 6 is provided on the outer surface of the reagent card 1 to facilitate the detector to scan and obtain relevant information of the reagent card 1 .

[0086] The following is a detailed introduction of the reagent card 1 through multiple embodiments:

[0087] Example 1

[0088] like Figure 2 As shown, this embodiment discloses a four-channel reagent card 1 , which includes four detection chambers Ⅰ to Ⅳ that are independently arranged in sequence and connected to a sample temporary storage chamber 2 .

[0089] 1) Method 1: Use detection chamber III as background channel:

[0090] The detection chamber III does not contain the detection reagent, which serves as a blank control; the detection reagents contained in the remaining three detection chambers I, II and IV are exactly the same as the corresponding detection chambers in the following "Method 2". The sample to be tested enters the sample flow channel 9 of the reagent card 1 through the injection port, flows along the sample flow channel 9 to the four detection chambers 4, and the detection reagents in the detection chambers I, II and IV are dissolved in the sample, and then react with the detection components in the sample, generating a signal change relative to the blank control in the sample detection chamber III, and the sample is tested by the size of the signal change.

[0091] 2) Method 2 uses temporary storage 2 as the background channel:

[0092] The temporary storage chamber 2 does not contain any detection reagent and serves as a blank control; each detection chamber 4 contains a detection result amplifier and a platelet activator; the platelet activator used in each detection chamber is different; the detection result amplifier selects dyed latex microspheres, whose surface is coated with fibrinogen; when the latex microspheres are not aggregated, the turbidity of the detection chamber 4 is high; on the contrary, when they adhere to and aggregate with the activated platelets, the turbidity of the detection chamber 4 becomes low; the main function of the platelet activator is to activate platelets so as to aggregate the remaining latex microspheres mentioned above and change the sample turbidity of the detection chamber 4.

[0093] The detection chamber I contains channel I detection reagent, and the channel I detection reagent contains a lyophilized agent made of a detection result amplifier; the detection chamber II contains channel II detection reagent, and the channel II detection reagent contains a lyophilized agent made of a platelet activator and a detection result amplifier; the detection chamber III contains channel III detection reagent, and the channel III detection reagent contains a lyophilized agent made of a platelet activator and a detection result amplifier; the detection chamber IV contains channel IV detection reagent, and the channel IV detection reagent contains a lyophilized agent made of a platelet activator and a detection result amplifier. The sample to be tested enters the sample flow channel 9 of the reagent card 1 through the sample inlet, flows along the sample flow channel 9 to the four detection chambers 4, and the detection reagents in the detection chambers I, II, III and IV are dissolved in the sample, and then react with the detection components in the sample, generating a signal change relative to the blank control in the sample temporary storage chamber 2, and the detection of the sample is completed by the size of the signal change.

[0094] Preferably, the platelet activator is selected from arachidonic acid, adenosine diphosphate, collagen, adrenaline, 5-hydroxytryptamine, prostaglandin, thromboxane A 2、 Any one of ristocetin, phospholipase A, phospholipase C, thrombin, isothrombin, and isothrombin activating peptide;

[0095] Introduction to the principle of platelet aggregation:

[0096] The role of adenosine diphosphate (ADP): When blood vessels are damaged, the damaged blood vessel walls and surrounding tissues release ADP. ADP can trigger platelet activation and aggregation by binding to receptors on the surface of platelets. The binding of ADP leads to the release of calcium ions in platelets and the activation of intracellular signaling pathways, thereby promoting changes in the morphology and function of platelets, making them more adherent and aggregated.

[0097] The role of thrombin: During the process of vascular injury, the damaged vascular wall will release tissue factor, which activates the formation of thrombin. Thrombin can convert fibrinogen into fibrin to form a thrombus. At the same time, thrombin can also directly stimulate the activation of platelets, and promote platelet aggregation and morphological changes by binding to receptors on the surface of platelets.

[0098] The role of phospholipase A and phospholipase C (PLA and PLC): Phospholipase is an enzyme that plays an important role in the process of platelet activation. When platelets are activated, phospholipase is activated and transferred to the surface of platelets, thereby initiating the metabolism and changes of phospholipids. These changes can affect the properties of the cell membrane, making platelets more adherent and aggregated.

[0099] The role of arachidonic acid: Arachidonic acid can be metabolized by enzymes in platelets into a series of biologically active substances, such as prostaglandins, thromboxanes, etc. These substances can promote platelet activation and aggregation by activating intracellular signaling pathways, thereby participating in the process of thrombosis.

[0100] Role of collagen (COL): Collagen is an important extracellular matrix protein of vascular endothelial cells. It is exposed in the blood and binds to receptors on the surface of platelets, thereby initiating platelet activation and aggregation. Collagen activates platelets by binding to receptors GPVI and α2β1 on the surface of platelets and induces them to release platelet-activating factors, such as platelet kinase and 5-hydroxytryptamine, thereby promoting platelet aggregation and morphological changes. In addition, collagen can further promote platelet activation and aggregation by activating platelet signal transduction pathways, such as phospholipase C and protein kinase C.

[0101] The role of thromboxane A2: It is a biologically active substance produced by arachidonic acid metabolism. Thromboxane A2 mainly promotes platelet activation and aggregation by binding to the receptor TP on the surface of platelets. Thromboxane A2 activates the signaling pathway within platelets, such as increasing the intracellular calcium ion concentration and activating phospholipase C, leading to changes in platelet morphology and the release of platelet activation factors, thereby promoting platelet aggregation and thrombosis.

[0102] In general, adenosine diphosphate, thrombin, phospholipase, arachidonic acid, collagen, and thromboxane A2 can all activate platelets through different pathways and cause changes in their morphology and function, thereby participating in the process of thrombosis and hemostasis. The interaction and regulation of these biochemical reactions are important mechanisms of platelet activation and thrombosis.

[0103] The blood collection tube 5 contains an anticoagulant sample, which is undiluted whole blood, plasma or diluted whole blood, all of which are mixed with a sodium citrate solution in a volume ratio of 1:9, and the concentration of the sodium citrate solution is 0.109 mol / L or 0.129 mol / L.

[0104] Preferably, the detection chamber I contains the detection result amplifier, and the sample does not adhere to or aggregate with the latex microspheres in the detection chamber, so that the turbidity of the sample in the detection chamber I is the largest;

[0105] Preferably, the detection chamber II contains one of the platelet activators, namely, isothrombin activating peptide, which avoids activating platelets through a mixture of phospholipase A and phospholipase C or arachidonic acid or adenosine diphosphate pathways as much as possible. The isothrombin activating peptide directly activates platelets to cause a cascade reaction, causing the sample to adhere to and aggregate with the latex microspheres in the detection chamber, thereby changing the turbidity of the sample in the detection chamber II, thereby obtaining the maximum aggregation of platelets under the isothrombin pathway, and obtaining the corresponding results through the cooperation of specific hardware and software in the device;

[0106] Preferably, the detection chamber III refers to the following two methods:

[0107] Method 1: Use detection chamber III as background channel: There is no detection reagent in detection chamber III, which serves as blank control;

[0108] Method 2: Using temporary storage chamber 2 as the background channel: The detection chamber III contains one of the platelet activators - a mixture of phospholipase A and phospholipase C or arachidonic acid; the mixture of phospholipase A and phospholipase C or arachidonic acid activates platelets and releases more platelet aggregation factors, thereby causing a cascade reaction, causing the sample to adhere to and aggregate with the latex microspheres in the detection chamber, thereby changing the turbidity of the sample in the detection chamber III. If the person being tested has previously taken drugs such as cyclooxygenase-1 inhibitors (such as: small doses of 81-325 mg aspirin, indobufen, etc.), the activation of platelets will be inhibited, reducing the rate of change of turbidity in the detection chamber, and the corresponding results obtained through the cooperation of specific hardware and software in the device will also be reduced;

[0109] Preferably, the detection chamber IV contains one of the platelet activators, adenosine diphosphate. Adenosine diphosphate will bind to the adenosine diphosphate receptors on the surface of the platelet membrane, activate the platelets and release substances such as thrombin and adenosine diphosphate, thereby causing a cascade reaction, causing the sample to adhere to and aggregate with the latex microspheres in the detection chamber, thereby changing the turbidity of the sample in the detection chamber IV. If the person being tested has previously taken drugs such as ADP inhibitors (such as clopidogrel, etc.), the activation of platelets will be inhibited, reducing the rate of change of turbidity in the detection chamber, and the corresponding results obtained through the cooperation of specific hardware and software in the device will also be reduced.

[0110] The reagent card 1 of the present invention can detect the platelet aggregation ability, and the detection method is as follows:

[0111] 1) Use the supporting software to calculate the platelet aggregation rate AggI in the detection chamber I;

[0112] 2) Use the supporting software to calculate the platelet aggregation rate AggⅡ in the detection chamber Ⅱ;

[0113] 3) Use the supporting software to calculate the platelet aggregation rate AggⅢ in the detection chamber III;

[0114] 4) Use the supporting software to calculate the platelet aggregation rate AggⅣ in the detection chamber IV.

[0115] The software algorithm is as follows:

[0116] like Figure 3 As shown, the software algorithm uses a mathematical model fitting method. According to the differences in the detection reagents set in each detection chamber, the platelet aggregation reaction process is first established with time t as the independent variable and light transmittance as the dependent variable. According to the Lambert-Beer law and the change in microsphere concentration, the algorithm formula is fitted.

[0117] Figure 3 The meanings of the curves in are as follows:

[0118] Blood whole blood sample reaction line: There are no latex microspheres and activators in the detection chamber or temporary storage chamber. The platelets are not activated and there are no latex microspheres and activators to block the light from passing through the sample, so the light transmittance is the highest;

[0119] Blood+Mic is the whole blood sample + latex microsphere reaction line: after the sample enters the detection chamber, it is stirred and mixed with the latex microspheres. Since there is no activator, the platelets are not activated and will not adhere to the latex microspheres. At the same time, there is a dye on the surface of the latex microspheres, which hinders the detection light, so the transmittance of this detection chamber is the lowest and can be used as a low-value blank control;

[0120] TRAP, ADP, PLA+PLA, COL and COX-1 are in parallel relationship and are used as different types of activators in the detection chamber. Platelets can be activated through their corresponding pathways to make them adhere and aggregate with the latex microspheres in the detection chamber, so as to calculate their light transmittance intensity.

[0121] in:

[0122] TRAP: Thrombin receptor activating peptide activation response curve;

[0123] ADP: adenosine diphosphate activator activation response curve;

[0124] PLA+PLC: phospholipase A and phospholipase C activation reaction curve;

[0125] AA: arachidonic acid activation response curve;

[0126] COL: collagen activation response curve;

[0127] COL-1: Cyclooxygenase-1 inhibitor curve;

[0128] The temporary storage or testing chamber where the blood is located corresponds to the maximum transmittance I Blood , so its concentration is the lowest;

[0129] The transmittance of the detection chambers where TRAP, ADP, PLA+PLA, COL and COX-1 are located is I TRAP ,I ADP ,I 2P ,I COL and I AA ;

[0130] The minimum light transmittance of the Blood+Mic detection chamber is I Blood+Mic , so its concentration is the highest.

[0131] The concentration of the ADP curve is recorded as C ADP ,C ADP =a*lg(I Blood / I ADP );

[0132] Its maximum concentration is recorded as C ADP.MAX ,C ADP.MAX =a*lg(I Blood / I ADP.MIN );

[0133] The concentration of the TRAP curve is recorded as C TRAP ,C TRAP =a*lg(I Blood / I TRAP );

[0134] Its maximum concentration is recorded as C TRAP.MAX ,C TRAP.MAX =a*lg(I Blood / I TRAP.MIN );

[0135] The concentration of PLA+PLC curve is recorded as C 2P , C 2P =a*lg(I Blood / I 2P );

[0136] Its maximum concentration is recorded as C 2P.MAX ,C 2P.MAX =a*lg(I Blood / I 2P.MIN );

[0137] The concentration of the COL curve is recorded as C COL ,C COL =a*lg(I Blood / I COL );

[0138] Its maximum concentration is recorded as C COL.MAX ,C COL.MAX =a*lg(I Blood / I COL.MIN );

[0139] The concentration of COX-1 curve is recorded as C AA ,C AA =a*lg(I Blood / I AA );

[0140] Its maximum concentration is recorded as C AA.MAX ,C AA.MAX =a*lg(I Blood / I AA.MIN );

[0141] The concentration of blood is recorded as C Blood =a*lg(I Blood / I Blood )=0;

[0142] The concentration of Blood+Mic is recorded as C Blood+Mic =a*lg(I Blood / I Blood+Mic );

[0143] The temporary storage or testing chamber where the blood is located corresponds to the maximum transmittance I Blood , which has the lowest concentration;

[0144] The transmittance of the detection chambers where TRAP, ADP, PLA+PLA, COL and COX-1 are located is I TRAP ,I ADP ,I 2P ,I COL and I AA The minimum transmittance of each detection chamber is I TRAP.MIN ,I ADP.MIN ,I 2P.MIN ,I COL.MIN and I AA.MIN ;

[0145] The minimum light transmittance of the Blood+Mic detection chamber is I Blood+Mic , which has the highest concentration;

[0146] Based on the above, the reagents are tested according to the configuration of the reagent card test chamber:

[0147] Platelet aggregation response unit PU of P2Y12 receptor activation pathway P2Y12 :

[0148] PU P2Y12 = a*[(C MAX -C ADP ) / (C MAX -C Blood )]+b;

[0149] C MAX = C Blood+Mic or C ADP.MAX ;

[0150] Platelet aggregation response unit PU BASE :

[0151] PU BASE =a*[(C MAX -C TRAP ) / (C MAX -C Blood )]+b;

[0152] C MAX = C Blood+Mic or C TRAP.MAX ;

[0153] Platelet aggregation response unit AU of cyclooxygenase-1 activation pathway AA :

[0154] AU AA =a*[(C MAX -C AA ) / (C MAX -C Blood )]+b;

[0155] C MAX = C Blood+Mic or C AA.MAX ;

[0156] Platelet aggregation response unit CU COL :

[0157] CU COL =a*[(C MAX -C COL ) / (C MAX -C Blood )]+b;

[0158] C MAX = C Blood+Mic or C COL.MAX ;

[0159] Platelet aggregation response unit PU of phospholipase A and phospholipase C activation pathway 2P :

[0160] PU 2P =a*[(C MAX -C 2P ) / (C MAX -C Blood )]+b;

[0161] C MAX = C Blood+Mic or C 2P.MAX ;

[0162] In the above formula, a and b are constant terms; C MAX It is a variable, which indicates the maximum concentration reference parameter used when calculating platelet aggregation reaction units, as described below:

[0163] 1) One of the test chambers of the reagent card is only equipped with a test result amplifier, and its test corresponding concentration is C Blood+Mic ; When calculating the platelet aggregation units of other test chambers, this is used as the maximum concentration reference parameter C MAX ;

[0164] 2) All test chambers of the reagent card are equipped with platelet activators and test result amplifiers at the same time; at this time, the platelet aggregation unit of the test chamber is calculated, and the maximum test concentration of each test chamber is used as the reference parameter C MAX .

[0165] Example 2

[0166] This embodiment discloses a five-channel reagent card 1 , which comprises five independent detection chambers Ⅰ to Ⅴ arranged in sequence and connected to a sample temporary storage chamber 2 .

[0167] 1) Method 1: Use detection chamber III as background channel

[0168] The detection chamber III does not contain the detection reagent, which serves as a blank control; the detection reagents contained in the remaining four detection chambers I, II, IV and V are exactly the same as the corresponding detection chambers in the following "Method 2". The sample to be tested enters the sample flow channel 9 of the reagent card 1 through the sample inlet, flows along the sample flow channel 9 to the five detection chambers 4, and the detection reagents in the detection chambers I, II, IV and V are dissolved in the sample, and then react with the detection components in the sample, generating a signal change relative to the blank control in the sample detection chamber III, and the sample is tested by the size of the signal change.

[0169] 2) Method 2: Use temporary storage 2 as background channel

[0170] The temporary storage chamber 2 does not contain any detection reagent and serves as a blank control; each detection chamber 4 contains a detection result amplifier and a platelet activator; the platelet activators used in each detection chamber are different; the detection result amplifier is selected from dyed latex microspheres, the surface of which is coated with fibrinogen.

[0171] The detection chamber I contains a channel I detection reagent, and the channel I detection reagent contains a freeze-dried agent made of a detection result amplifier; the detection chamber II contains a channel II detection reagent, and the channel II detection reagent contains a freeze-dried agent made of a platelet activator and a detection result amplifier; the detection chamber III contains a channel III detection reagent, and the channel III detection reagent contains a freeze-dried agent made of a platelet activator and a detection result amplifier; the detection chamber IV contains a channel IV detection reagent, and the channel IV detection reagent contains a freeze-dried agent made of a platelet activator and a detection result amplifier; the detection chamber V contains a channel V detection reagent, and the channel V detection reagent contains a freeze-dried agent made of a platelet activator and a detection result amplifier. The sample to be tested enters the sample flow channel 9 of the reagent card 1 through the sample inlet, and the sample flows along the sample flow channel 9 to the five detection chambers 4, and the detection reagents in the detection chambers I, II, III, IV and V are dissolved in the sample, and the detection reagent reacts with the detection component in the sample, and a signal change is generated relative to the blank control in the sample temporary storage chamber 2, and the sample is detected through the signal change.

[0172] Preferably, the platelet activator is selected from any one of arachidonic acid, adenosine diphosphate, collagen, adrenaline, 5-hydroxytryptamine, prostaglandin, thromboxane A2, ristocetin, phospholipase A, phospholipase C, thrombin, isothrombin, and isothrombin activating peptide;

[0173] Preferably, the detection chamber I contains a detection result amplifier;

[0174] Preferably, the test chamber II contains one of the platelet activators, isothrombin activating peptide;

[0175] Preferably, the detection chamber III refers to the following two methods:

[0176] Method 1 uses detection chamber III as the background channel: detection chamber III does not contain detection reagents, serving as a blank control;

[0177] Method 2 uses temporary storage chamber 2 as the background channel: detection chamber III contains one of the platelet activators, a mixture of phospholipase A and phospholipase C or arachidonic acid;

[0178] Preferably, the detection chamber IV contains adenosine diphosphate, one of the platelet activators;

[0179] Preferably, the detection chamber V contains one of the platelet activators, ristocetin.

[0180] Finally, the device will give a quantitative value based on the changes in the turbidity of the entire sample in different channels:

[0181] The platelet aggregation rate AggI in channel I was calculated using the supporting software;

[0182] The platelet aggregation rate AggⅡ in channel Ⅱ was calculated using the supporting software;

[0183] The platelet aggregation rate AggⅢ in channel III was calculated using the supporting software;

[0184] The platelet aggregation rate AggⅣ in channel Ⅳ was calculated using the supporting software;

[0185] The platelet aggregation rate AggV in channel V was calculated using the supporting software;

[0186] The software algorithm is the same as that in Example 1 and will not be described again here.

[0187] Example 3

[0188] Based on Example 1 and Example 2, six-channel, seven-channel or more-channel reagent cards are prepared according to clinical needs; the reagent card 1 includes six or seven or more independent detection chambers 4. The reagents of the first five channels of the six-channel reagent card 1 are consistent with those of the five-channel reagent card 1, that is, based on the detection items of the five-channel card, a platelet activator different from other detection chambers 4 is added inside the VI detection chamber 4 to meet more detection requirements;

[0189] The function of the seven-channel reagent card 1 is similar to the above description. The first six detection chambers 4 are consistent with the six-channel reagent card 1. By adding a platelet activator different from that in other detection chambers inside the VII detection chamber 4, the detection requirements are further expanded.

[0190] The same goes for more channel reagent cards 1, which will not be described in detail here.

[0191] In summary, a small amount of blood can be drawn at one time to achieve the results required by the clinic, and a more accurate, faster and more convenient guidance plan can be provided for clinical diagnosis and treatment.

[0192] In order to further understand the working principle of the reagent card, the detailed structure of the platelet capacity tester is introduced as follows:

[0193] The methodological basis of this platelet capacity detector is optical turbidimetry, also known as turbidity determination, which is a method for determining the concentration of suspended matter by measuring the light intensity passing through a suspended particle medium. It is a world-recognized methodology for detecting platelet function. Optical turbidimetry uses the fact that changes in the concentration of relevant absorbing substances in a solution will affect the absorbance of the solution to light of a specific wavelength, that is: the higher the concentration of the absorbing substance, the higher the absorbance of the solution to light of a specific wavelength, and the lower the light transmittance; conversely, the lower the concentration of the absorbing substance, the lower the absorbance of the solution to light of a specific wavelength, and the higher the light transmittance. The concentration of the specific absorbing substance can be determined and then related analysis can be performed. The stability of the light source of the platelet capacity detector and the measurement accuracy of its luminous flux output ultimately affect the accuracy of the results. Only by ensuring the stability of the light source drive and the accuracy of the light signal collection can the risk be minimized. In response to the above risks, the present invention makes the following innovations to the platelet capacity detector:

[0194] The detector includes a reagent card storage compartment, a control module, an automatic sampling module, a motor stirring module, a temperature control module, an optical path detection module, a calibration module, and a data display and printing module;

[0195] Specific:

[0196] The control module includes an electrically connected MCU slave processing module and a control circuit;

[0197] The automatic sampling module includes a reagent card recognition module, a vacuum pump, a first solenoid valve, a second solenoid valve, a one-way valve, a first infrared sensor receiving module, a second infrared sensor receiving module, and a pressure sensor;

[0198] like Figure 4 As shown, the reagent card identification module includes an information code label 6 attached to the reagent card 1 and a barcode scanner provided on the detector; the barcode scanner is used to scan the information code label 6 on the reagent card 1 and output the information to the control circuit, which then transmits the data to the MCU slave processing module.

[0199] like Figure 5As shown, the detection chamber 4 is connected to the one-way valve through the first vent 2b of the microfluidic channel; the temporary storage chamber 2 is respectively connected to the No. 1 solenoid valve and the No. 2 solenoid valve through the second vent 2c of the microfluidic channel and the air circuit, the exhaust port of the vacuum pump is connected to the No. 2 solenoid valve for inflating the temporary storage chamber 2; the air inlet of the vacuum pump is connected to the No. 1 solenoid valve for exhausting the temporary storage chamber 2; the control circuit can control the operation of the vacuum pump; the pressure sensor is used to detect the air pressure on the No. 1 solenoid valve air circuit to determine whether there is an abnormality in the vacuum pumping process; when the temporary storage chamber 2 is in a state of being exhausted, ... A negative pressure environment is required in the warehouse 2 to extract the sample in the blood collection tube 5, the one-way valve is closed, and no air flows into the first vent 2b; the No. 2 solenoid valve is closed, the No. 1 solenoid valve is turned on, and the vacuum pump controls the No. 1 solenoid valve to extract the air in the temporary storage warehouse 2 from the second vent 2c; when the sample in the temporary storage warehouse 2 reaches the set height, the No. 1 solenoid valve is closed, the No. 2 solenoid valve is turned on, and the air enters the temporary storage warehouse 2 through the second vent 2c; the one-way valve is opened, and as the sample is injected into each detection warehouse 4 through the microfluidic channel 3, the air is discharged through the first vent 2b.

[0200] like Figure 6 As shown, the first infrared sensor receiving module detects the color change of the blood-absorbing cotton 2a on the temporary storage bin 2 through an infrared sensor, and feeds back a signal to the MCU slave processing module. The MCU slave processing module sends an instruction to the control circuit to determine the working state of the vacuum pump based on the signal.

[0201] like Figure 7 As shown, the MCU slave processing module sends an air pumping command to the control circuit, and the vacuum pump pumps air into the temporary storage bin 2 through the second solenoid valve, and the temporary storage bin 2 changes from negative pressure to positive pressure. At this time, since each detection bin channel of the reagent card 1 is under negative pressure, the sample liquid (whole blood or quality control liquid) flows to each detection bin 4. A detection cotton 4a is provided at the same position in each detection bin 4; when the sample height reaches the detection cotton 4a, the color of the detection cotton 4a will change; the second infrared sensor receiving module monitors the color change of the detection cotton 4a through multiple infrared sensors corresponding to the detection bin 4 and outputs a signal to the MCU slave processing module, and sends a closing air pumping command to the control circuit, thereby stopping the injection of samples into the detection bin 4.

[0202] According to the above structure, the working steps of the automatic injection module are briefly described as follows:

[0203] Step 1) Reagent Card 1 determination:

[0204] After attaching the information code label 6 to the specified area of ​​the reagent card 1, put it into the reagent card storage compartment of the platform; after the test starts, the MCU slave processing module controls the power-on of the device's barcode scanner, reads the reagent card 1 label information, and outputs it to the MCU slave processing module; the MCU slave processing module receives the reagent card 1 information, performs data analysis and judgment, and then executes the corresponding detection program.

[0205] Step 2) Sample injection into temporary storage compartment 2 of reagent card 1:

[0206] The MCU slave processing module outputs a vacuuming instruction, and uses the pressure sensor to detect whether the pressure of the gas circuit meets the normal level to determine whether there is any abnormality in the gas circuit; if the gas circuit is normal, the control circuit powers on the vacuum pump to start vacuuming the temporary storage bin 2, and the No. 1 solenoid valve changes from a closed state to a conducting state after powering on; the pressure sensor converts the detected air pressure into an electrical signal, and outputs it to the MCU slave processing module through the control circuit; after vacuuming, the negative pressure in the temporary storage bin 2 of the reagent card 1 is much lower than the air pressure in the blood collection tube 5 inserted into the reagent card 1, and the sample (whole blood or quality control liquid) in the blood collection tube 5 begins to flow into the temporary storage bin 2 of the reagent card 1.

[0207] Step 3) Stop injecting reagent card 1 into temporary storage compartment 2

[0208] When the liquid intake reaches the specified position of the temporary storage bin 2 of the reagent card 1, the blood-absorbing cotton 2a will change color. The first infrared sensor receiving module will feed back the color change signal of the blood-absorbing cotton 2a to the MCU slave processing module through the corresponding infrared sensor. The MCU slave processing module will send an instruction to the control circuit based on the signal to stop the power supply of the vacuum pump and the solenoid valve 1, and complete the sampling of the temporary storage bin 2.

[0209] Step 4) Sample injection into the detection chamber 4 of the reagent card 1:

[0210] The MCU slave processing module outputs an air-inflating instruction, and uses the pressure sensor to detect whether the pressure of the gas circuit meets the normal level to determine whether there is any abnormality in the gas circuit; if the gas circuit is normal, the control circuit starts to inflate after the vacuum pump is powered on, the No. 1 solenoid valve is powered off and closed, and the No. 2 solenoid valve is powered on and turned on, and the vacuum pump inflates air into the temporary storage bin 2 through the No. 2 solenoid valve, and the negative pressure in the temporary storage bin 2 is changed to positive pressure. At this time, since each detection bin channel of the reagent card 1 is at negative pressure, the sample liquid (whole blood or quality control liquid) flows to each detection bin 4.

[0211] Step 5) Stop sampling in the detection chamber 4 of the reagent card 1:

[0212] After all the detection chambers 4 are filled with samples, the corresponding detection cotton 4a inside them will change color; the output signals of the corresponding infrared sensors will also change accordingly and be sent to the MCU slave processing module; after receiving the infrared sensor signal, the MCU slave processing module sends an instruction to the control circuit to stop the power supply of the vacuum pump and the solenoid valve, and complete the sampling of the detection chamber 4.

[0213] like Figure 8 As shown, the temperature control module includes a heating resistor and a thermistor detection element; the reagent card storage compartment contains a heat-conducting aluminum block, which is attached to one side of the reagent card 1 and covers the temporary storage compartment 2 and each detection compartment 4 at the same time; the heating resistor and the thermistor detection element are fixed on the heat-conducting aluminum block, and heat is conducted through the aluminum block to control the temperature of the samples in the temporary storage compartment 2 and each detection compartment 4; after the instrument is turned on, the MCU slave processing module of the control module outputs the control signal of the heating circuit according to the preset PWM frequency, supplies power to the heating resistor, and feeds back the receiving signal of the thermistor detection element to the MCU slave processing module to perform temperature closed-loop feedback control processing; so that the detection sample of the reagent card 1 is maintained at a specified temperature;

[0214] In order to more accurately replicate the "gold standard" turbidimetry, the following technical basic work and improvements were made to the equipment used with reagent card 1. First, the accurate emission of photons in the device is achieved using precision optical devices and precise control systems, including a specific LED light source, so that the light source in the device can achieve a stable output power and wavelength to ensure that the emission of photons is stable and controllable. Secondly, the device captures photons. The photodiode detection device used in the device has high sensitivity and low noise to ensure that the capture of photons is efficient and reliable. In addition, precise optical lenses and filter devices are used at the same time to ensure the controllability of the capture direction and wavelength range of photons.

[0215] like Figure 8As shown, the optical path detection module includes a light source transmitting module and a light source receiving module; the reagent card 1 is fixed to the reagent card storage compartment and corresponds to the light source transmitting module and the light source receiving module, and the reagent card 1 is perpendicular to the detection light; the MCU slave processing module starts the various light-emitting LEDs (corresponding to the light detection channel 8 one by one) of the light source transmitting module to power on, and the light-emitting LED sends a light signal to each detection compartment 4, and the various photodiodes (corresponding to the light detection channel 8 one by one) of the light source receiving module will filter, amplify and photoelectrically convert the light signals passing through each light detection channel 8 and transmit them to the MCU slave processing module, and the detection results AU or PU are obtained through processing, and displayed on the device screen interface. Specifically, the MCU slave processing module of the control module outputs a pre-sampling instruction, first gradually increases the power supply voltage of the predetermined channel circuit of the light source board, so that the corresponding channel receiving value of the light source receiving module reaches the preset standard, and then according to the stored calibration compensation data, the remaining light source transmitting circuits of the light source transmitting module are powered respectively, and the optical path detection work is started. The lower computer software of the present invention ensures the stable driving of the light source through a 12-bit DAC current source, and simultaneously collects and analyzes the spectral signal through a 12-bit ADC module, a photodiode, and a 3.3V reference source, thereby ensuring the accurate collection of the optical signal and minimizing the risk.

[0216] like Fig. 9 As shown, the motor stirring module includes a stirring device placed in each detection chamber 4 and a detection motor arranged below the reagent card 1; the stirring device includes a limit baffle 4b horizontally fixed to the inner wall of the detection chamber and a steel ball 4c at the bottom thereof; a magnet is arranged on the transmission shaft of the detection motor, so that when the motor rotates, it can drive the steel ball in the detection chamber 4 to produce a certain trajectory of movement to stir the tested sample evenly; there is a gap between the limit baffle 4b and the inner wall of the detection chamber for sample circulation; the width of the gap is smaller than the diameter of the steel ball 4c, so that the steel ball is restricted to the bottom of the detection chamber and can only move in the horizontal direction. At the same time, the speed data of the stirring motor is fed back to the MCU slave processing module through the sensor corresponding to the detection motor transmission shaft handle on the light source board, and the stirring frequency closed-loop feedback control processing is performed.

[0217] like Fig.10As shown, the calibration module includes an optical attenuation sheet, an electronic quality control card, a data storage module, and a PC host computer; the optical attenuation sheet is fixed to the electronic quality control card and is fixed as a whole between the light source transmitting module and the light source receiving module through the reagent card accommodating compartment, and the optical attenuation sheet vertically corresponds to each light-emitting LED and the photosensitive receiving diode; the working principle of the module is: the PC host computer software sends a calibration instruction, the MCU slave processing module starts the calibration processing software, first gradually increases the power supply voltage of the pre-sampling channel circuit of the light source board, so that the corresponding channel receiving value of the light source receiving module reaches the preset standard; then adjusts the power supply of the other light sources respectively, so that the corresponding receiving channels also reach the preset standard; finally, the corresponding relationship between the power supply of the other several power supplies of the light source board and the pre-sampling channel power supply is calculated by the existing technology, and the calibration compensation data is obtained and transmitted to the data storage module.

[0218] The fully automatic platelet aggregation instrument uses the principle of optical turbidimetry. During detection, the working principle of the automatic sampling module is as follows: the information code on the reagent card 1 is scanned and identified through the reagent card identification module; then the blood sample is automatically extracted and added according to the information obtained, and at the same time, it is combined with the gas circuit, vacuum pump and corresponding solenoid valve of the platform and the liquid circuit structure of the reagent card 1 to ensure that the samples entering the detection chamber are all unactivated samples; the motor stirring module is according to the stirring frequency preset by the software, and the control module outputs the command to control the detection motor, and stirs the reaction in the detection chamber 4 by the rotation of the detection motor; the temperature control module simulates the in vivo reaction environment to ensure the stability of the surrounding temperature during in vitro detection, and the different reaction chambers of the reagent card 1 undergo corresponding polymerization reactions during the reaction process; the optical path detection module detects the change in the photometric value caused by the polymerization reaction and calculates it to obtain the experimental results; the calibration module ensures that the power supply voltage of the optical path detection module can reach the preset standard through calibration compensation; the data display and printing module realizes the display and printing functions of the final experimental results.

[0219] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A whole blood quantitative platelet aggregation ability detection reagent card, characterized in that: The reagent card comprises a sample inlet, a temporary storage chamber, a microfluidic channel, a main liquid separation guide groove, a sample flow channel and N detection chambers with the same internal volume, N is a positive integer, and N≥2; The sample inlet is used to fix the blood collection tube, the blood collection needle passes through the blood collection tube from the bottom, and guides the sample to the temporary storage bin through the connecting tube at the tail of the blood collection needle; each detection bin is provided with a sample flow channel; a blood-absorbing cotton is provided at a specified height on the inner wall of the temporary storage bin; The microfluidic channel is provided with a first vent and a second vent, the first vent is connected to a one-way valve; the second vent is respectively connected to a No. 1 solenoid valve and a No. 2 solenoid valve through an air path, the No. 1 solenoid valve and the No. 2 solenoid valve are connected in parallel, and the No. 1 solenoid valve is connected to an air inlet of a vacuum pump, and the No. 2 solenoid valve is connected to an air outlet of the vacuum pump; A ventilation groove is provided on the top of all the detection chambers, and a main exhaust hole connected to the outside is provided at the center of the ventilation groove; each detection chamber is connected to the ventilation groove through an exhaust groove, and an exhaust hole and detection cotton facing the exhaust hole are provided at the connection between the exhaust groove and the ventilation groove; there is a gap for air circulation between the detection cotton and the top and bottom of the exhaust groove; the bottom of the detection cotton expands rapidly at the moment of contact with the sample, closing the exhaust hole, and the detection chamber cannot exhaust gas and cannot continue to inject samples, and there is a gap between the top of the detection cotton and the exhaust groove, which does not affect the gas circulation of other detection chambers; when all the detection cottons expand and change color, the infrared sensor detected by the identification module will send the color change signal of the detection cotton to the MCU slave processing module, stop the power supply of the vacuum pump, the No. 1 solenoid valve and the No. 2 solenoid valve, and complete the injection of all detection chambers; One of the temporary storage chamber or the N detection chambers is a blank control, and no lyophilized agent is placed therein; one or more of the remaining detection chambers are placed with a designated lyophilized agent, and the lyophilized agent includes a platelet activator and a detection result amplifier; The samples to be tested in the temporary storage chamber enter into each detection chamber in turn through the microfluidic channel, the main liquid separation guide groove and the sample flow channel, and the detection reagent in the detection chamber dissolves in the samples to be tested and then reacts.

2. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 1, characterized in that: Optical detection channels are provided on opposite sides of the temporary storage bin and each detection bin.

3. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 1, characterized in that: The detection method of the platelet aggregation ability by the reagent card is: 1) Calculate the platelet aggregation rate in each test chamber containing platelet activator and test result amplifier using the supporting software; 2) The blank control is used as a transmittance reference, which is used for pre-sampling of the instrument optical path and as a reference indicator for calculation in step 1); The supporting software algorithm adopts the mathematical model fitting method: first, the platelet aggregation reaction process is established with time t as the independent variable and light transmittance as the dependent variable. According to the Lambert-Beer law and the change of microsphere concentration, the algorithm formula is fitted.

4. The whole blood quantitative platelet aggregation ability detection reagent card as claimed in claim 3, characterized in that: The concentration of the ADP curve is recorded as C ADP ,C ADP =a*lg(I Blood / I ADP ); Its maximum concentration is recorded as C ADP.MAX ,C ADP.MAX =a*lg(I Blood / I ADP.MIN ); The concentration of the TRAP curve is recorded as C TRAP ,C TRAP =a*lg(I Blood / I TRAP ); Its maximum concentration is recorded as C TRAP.MAX ,C TRAP.MAX =a*lg(I Blood / I TRAP.MIN ); The concentration of PLA+PLC curve is recorded as C 2P , C 2P =a*lg(I Blood / I 2P ); Its maximum concentration is recorded as C 2P.MAX ,C 2P.MAX =a*lg(I Blood / I 2P.MIN ); The concentration of the COL curve is recorded as C COL ,C COL =a*lg(I Blood / I COL ); Its maximum concentration is recorded as C COL.MAX ,C COL.MAX =a*lg(I Blood / I COL.MIN ); The concentration of COX-1 curve is recorded as C AA ,C AA =a*lg(I Blood / I AA ); Its maximum concentration is recorded as C AA.MAX ,C AA.MAX =a*lg(I Blood / I AA.MIN ); The concentration of blood is recorded as C Blood =a*lg(I Blood / I Blood )=0; The concentration of Blood+Mic is recorded as C Blood+Mic =a*lg(I Blood / I Blood+Mic ); The temporary storage or testing chamber where the blood is located corresponds to the maximum transmittance I Blood , which has the lowest concentration; The transmittance of the detection chambers where TRAP, ADP, PLA+PLA, COL and COX-1 are located is I TRAP ,I ADP ,I 2P ,I COL and I AA , the minimum transmittance of each detection chamber is I TRAP.MIN ,I ADP.MIN ,I 2P.MIN ,I COL.MIN and I AA.MIN ; The minimum light transmittance of the detection chamber where Blood+Mic is located is I Blood+Mic , which has the highest concentration; Based on the above, the reagents are tested according to the configuration of the reagent card test chamber: Platelet aggregation response unit PU of P2Y12 receptor activation pathway P2Y12 : PU P2Y12 = a*[(C MAX -C ADP ) / (C MAX -C Blood )]+b; C MAX = C Blood+Mic or C ADP.MAX ; Platelet aggregation response unit PU BASE : PU BASE =a*[(C MAX -C TRAP ) / (C MAX -C Blood )]+b; C MAX = C Blood+Mic or C TRAP.MAX ; Platelet aggregation response unit AU of cyclooxygenase-1 activation pathway AA : AT AA =a*[(C MAX -C AA ) / (C MAX -C Blood )]+b; C MAX = C Blood+Mic or C AA.MAX ; Platelet aggregation response unit CU COL : WITH COL =a*[(C MAX -C COL ) / (C MAX -C Blood )]+b; C MAX = C Blood+Mic or C COL.MAX ; Platelet aggregation response unit PU of phospholipase A and phospholipase C activation pathway 2P : PU 2P =a*[(C MAX -C 2P ) / (C MAX -C Blood )]+b; C MAX = C Blood+Mic or C 2P.MAX ; In the above formula, a and b are constant terms; C MAX It is a variable, which indicates the maximum concentration reference parameter used when calculating platelet aggregation reaction units, as described below: 1) One of the test chambers of the reagent card is only equipped with a test result amplifier, and its test corresponding concentration is C Blood+Mic ; When calculating the platelet aggregation units of other test chambers, this is used as the maximum concentration reference parameter C MAX ; 2) All test chambers of the reagent card are equipped with platelet activators and test result amplifiers at the same time; at this time, the platelet aggregation unit of the test chamber is calculated, and the maximum test concentration of each test chamber is used as the reference parameter C MAX .

5. The whole blood quantitative platelet aggregation ability detection reagent card as claimed in claim 1, characterized in that: A stirring device for stirring samples is placed in each testing chamber.

6. The whole blood quantitative platelet aggregation ability detection reagent card as claimed in claim 5, characterized in that: The stirring device comprises a limit baffle plate horizontally fixed on the inner wall of the detection bin and a steel ball at the bottom thereof.

7. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 6, characterized in that: There is a gap between the limit baffle and the inner wall of the detection chamber to facilitate the flow of samples; the width of the gap is smaller than the diameter of the steel ball.

8. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 1, characterized in that: The platelet activator is selected from arachidonic acid, adenosine diphosphate, collagen, adrenaline, 5-hydroxytryptamine, prostaglandin, thromboxane A 2、 Any one of ristocetin, phospholipase A, phospholipase C, thrombin, isothrombin, and isothrombin activating peptide; The detection result amplifier is selected from dyed latex microspheres, the surface of which is coated with fibrinogen.

9. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 1, characterized in that: The reagent card is a 4-channel reagent card, that is, N=4; any one of the detection chambers is a blank control, and no lyophilized agent is placed; and the remaining 1 to 3 detection chambers are placed with lyophilized agents.

10. The whole blood quantitative platelet aggregation ability detection reagent card according to claim 9, characterized in that: The platelet activator in the lyophilized agent is selected from any one of arachidonic acid, adenosine diphosphate and isothrombin activating peptide.

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