Microfluidic biochemical detection disc and detection method thereof

Abstract

The invention discloses a microfluidic biochemical detection disc and a detection method thereof. The microfluidic biochemical detection disc comprises a sample cavity (1), a sample quantification cavity (2), a diluent cavity (3), a diluent quantification cavity (4), a uniform mixing cavity (5) and a first detection cavity (6), a first reagent (R1) is pre-arranged in the uniform mixing cavity (5), and the uniform mixing cavity (5) provides a place for uniform mixing and reaction of the sample solution, the diluent and the first reagent (R1) so as to form a first reaction solution; a second reagent (R2) is preset in the first detection cavity (6), and the first detection cavity (6) provides a place for the reaction of the first reaction liquid and the second reagent (R2) to form a second reaction liquid. The microfluidic biochemical detection disc is provided with the uniform mixing cavity (5) and the first detection cavity (6), the two cavities are distributed in series, and the first reagent (R1) and the second reagent (R2) are distributed and preset in the two cavities, so that a sample solution can react with the first reagent (R1) and the second reagent (R2) in different cavities in sequence, an anti-interference reaction can be firstly performed on a sample, and then a detection reaction is performed; the accuracy of sample detection is improved.

Description

A microfluidic biochemical detection disc and detection method thereof Technical Field

[0001] The present invention relates to the technical field of biochemical detection, and in particular to a microfluidic biochemical detection disc and a detection method thereof. Background Art

[0002] Existing centrifugal microfluidic biochemical analyzers are equipped with centrifugal reagent disks, which are equipped with reaction wells for mixing reagents and samples to form a reaction solution for testing. Multiple reaction wells are arranged around the outermost circle of the reagent disk, and the reagents are placed in freeze-dried pellets in the reaction wells. Typically, each assay has only one reaction well, and the reaction occurs as soon as the sample enters the well. This prevents the interference-resistant properties of liquid biochemical reagents, which are typically divided into two reagents.

[0003] Summary of the Invention

[0004] In one embodiment, a microfluidic biochemical detection disc is provided, comprising:

[0005] A sample chamber, used for accommodating a sample liquid;

[0006] a sample quantification chamber, communicated with the sample chamber, and used for quantifying the sample liquid;

[0007] a diluent chamber, for containing a diluent;

[0008] a diluent quantitative chamber, communicated with the diluent chamber, for quantitatively measuring the diluent;

[0009] a mixing chamber connected to the sample quantification chamber via a first microchannel and connected to the diluent quantification chamber via a second microchannel; a first reagent is pre-placed in the mixing chamber, and the mixing chamber provides a place for mixing and reacting the sample liquid, the diluent, and the first reagent to form a first reaction solution;

[0010] The first detection chamber is connected to the mixing chamber through the third microchannel; the second reagent is pre-placed in the first detection chamber, and the first detection chamber provides a place for the first reaction solution and the second reagent to react to form the second reaction solution.

[0011] In one embodiment, a microfluidic biochemical detection disc is provided, comprising:

[0012] A sample chamber, used for accommodating a sample liquid;

[0013] a sample quantification chamber, communicated with the sample chamber, and used for quantifying the sample liquid;

[0014] a diluent chamber, for containing a diluent;

[0015] a diluent quantitative chamber, communicated with the diluent chamber, for quantitatively measuring the diluent;

[0016] a mixing chamber, connected to the sample chamber via a first microchannel and connected to the diluent chamber via a second microchannel; the mixing chamber is used to provide a place for mixing the sample liquid and the diluent to form a diluted sample;

[0017] a first reaction liquid accommodating chamber, connected to the mixing chamber; a first reagent is pre-placed in the first reaction liquid accommodating chamber, the first reaction liquid accommodating chamber is used to receive the diluted sample output from the mixing chamber, and the first reaction liquid accommodating chamber provides a place for the diluted sample to react with the first reagent to form a first reaction liquid;

[0018] The first detection chamber is connected to the first reaction liquid holding chamber through the third microchannel, and is used to receive the first reaction liquid output from the first reaction liquid holding chamber; the first detection chamber is pre-installed with a second reagent, and the first detection chamber provides a place for the reaction between the first reaction liquid and the second reagent to form the second reaction liquid required for project detection.

[0019] In one embodiment, a microfluidic biochemical detection disc is provided for performing at least one test on a sample to be tested. The microfluidic biochemical detection disc includes at least one detection unit, and the detection unit includes:

[0020] A sample chamber, used for accommodating a sample liquid;

[0021] a sample quantification chamber, communicated with the sample chamber, and used for quantifying the sample liquid;

[0022] a diluent chamber, for containing a diluent;

[0023] a diluent quantitative chamber, communicated with the diluent chamber, for quantitatively measuring the diluent;

[0024] a mixing chamber, connected to the sample chamber via a first microchannel and connected to the diluent chamber via a second microchannel; the mixing chamber is used to provide a place for mixing the sample liquid and the diluent to form a diluted sample;

[0025] a first reaction liquid accommodating chamber, connected to the mixing chamber; a first reagent is pre-placed in the first reaction liquid accommodating chamber, the first reaction liquid accommodating chamber is used to receive the diluted sample output from the mixing chamber, and the first reaction liquid accommodating chamber provides a place for the diluted sample to react with the first reagent to form a first reaction liquid;

[0026] The first detection chamber is connected to the first reaction liquid holding chamber through the third microchannel, and is used to receive the first reaction liquid output from the first reaction liquid holding chamber; the first detection chamber is pre-installed with a second reagent, and the first detection chamber provides a place for the reaction between the first reaction liquid and the second reagent to form the second reaction liquid required for project detection.

[0027] In one embodiment, a microfluidic biochemical detection disc is provided for performing at least one test on a sample to be tested. The microfluidic biochemical detection disc includes at least one detection unit, and the detection unit includes:

[0028] A sample chamber, used for accommodating a sample liquid;

[0029] a sample quantification chamber, communicated with the sample chamber, and used for quantifying the sample liquid;

[0030] a diluent chamber, for containing a diluent;

[0031] a diluent quantitative chamber, communicated with the diluent chamber, for quantitatively measuring the diluent;

[0032] a mixing chamber, connected to the sample quantification chamber via a first microchannel, and connected to the diluent quantification chamber via a second microchannel; a first reagent is pre-placed in the mixing chamber, and the mixing chamber provides a place for mixing and reacting the sample liquid, the diluent, and the first reagent to form a first reaction solution;

[0033] The first detection chamber is connected to the mixing chamber through the third microchannel; the second reagent is pre-placed in the first detection chamber, and the first detection chamber provides a place for the first reaction solution and the second reagent to react to form the second reaction solution.

[0034] In one embodiment, one detection unit corresponds to one detection item.

[0035] In one embodiment, the first reagent is used to react with the sample solution and the diluent to at least remove interfering substances in the sample solution.

[0036] In one embodiment, the interfering substance includes at least one of blood lipids, bilirubin, ascorbic acid, and hemoglobin.

[0037] In one embodiment, the detection unit includes at least two, and the volume of the sample quantitative chamber and / or the volume of the diluent quantitative chamber of any detection unit is different from the volume of the sample quantitative chamber and / or the volume of the diluent quantitative chamber of other detection units.

[0038] In one embodiment, the first reagent is a first reagent freeze-dried pellet made from a first finished liquid biochemical reagent, and / or the second reagent is a second reagent freeze-dried pellet made from a second finished liquid biochemical reagent.

[0039] In one embodiment, the first reagent freeze-dried ball and the second reagent freeze-dried ball respectively have a preset volume, the first detection chamber has a preset volume, the concentration of the first reagent freeze-dried ball and the concentration of the second reagent freeze-dried ball are respectively consistent with the concentration of the liquid biochemical reagent in the fully automatic biochemical analyzer, and the concentration of the second reaction liquid is consistent with the concentration of the working liquid in the fully automatic biochemical analyzer.

[0040] In one embodiment, the second reagent reacts with the first reaction liquid to obtain a sample, and the sample is optically detected to obtain a corresponding detection result.

[0041] In one embodiment, the mixing chamber or the first reaction liquid accommodating chamber is connected to at least two of the first detection chambers through the third microchannel, and the at least two first detection chambers respectively provide places for the reaction of the first reaction liquid and the second reagent to form at least two second reaction liquids.

[0042] In one embodiment, a second detection chamber is further included, which is connected to the mixing chamber or the first reaction liquid storage chamber through the third microchannel and is used to receive the first reaction liquid output from the mixing chamber or the first reaction liquid storage chamber.

[0043] In one embodiment, it includes a fan-shaped base, and the sample chamber, the diluent chamber, the mixing chamber, the first detection chamber, the first microfluidic channel, the second microfluidic channel and the third microfluidic channel are located in the fan-shaped base.

[0044] In one embodiment, a microfluidic biochemical detection method is provided, which uses a microfluidic biochemical detection disc for detection, including the following steps:

[0045] The centrifugal device drives the sample liquid in the sample quantitative chamber and the diluent in the dilution quantitative chamber into the mixing chamber;

[0046] The first reagent pre-placed in the mixing chamber is mixed and reacted with the sample liquid and the diluent to form a first reaction liquid;

[0047] The centrifugal device drives the first reaction liquid in the mixing chamber into the first detection chamber;

[0048] The second reagent pre-placed in the first detection chamber mixes and reacts with the first reaction liquid to form a second reaction liquid;

[0049] The detection device measures the second reaction liquid in the first detection chamber to obtain a detection result.

[0050] In one embodiment, after the mixing chamber obtains the first reaction liquid, the centrifugal device drives the first reaction liquid into the first detection chamber after a preset time interval.

[0051] In one embodiment, the centrifugal device drives the first reaction liquid into at least two of the first detection chambers to form at least two portions of the second reaction liquid; the detection mechanism measures the at least two portions of the second reaction liquid respectively to obtain at least two detection results.

[0052] In one embodiment, an average value of the at least two detection results is taken, and the average value is used as the final detection result.

[0053] In one embodiment, the first reagent is used to react with the sample solution and the diluent to at least remove interfering substances in the sample solution.

[0054] In one embodiment, the interfering substance includes at least one of blood lipids, bilirubin, ascorbic acid, and hemoglobin.

[0055] According to the microfluidic biochemical detection disc and the detection method thereof in the above-mentioned embodiment, since the microfluidic biochemical detection disc has a mixing chamber and a first detection chamber, the two chambers are distributed in series, and the two chambers are preset with a first reagent and a second reagent. The sample liquid can react with the first reagent and the second reagent in different chambers successively, so that the sample can first undergo an anti-interference reaction and then a detection reaction, thereby improving the accuracy of sample detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] FIG1 is a schematic structural diagram of a microfluidic biochemical detection disc in one embodiment;

[0057] FIG2 is a schematic structural diagram of a microfluidic biochemical detection disc in one embodiment;

[0058] FIG3 is a flow chart of a microfluidic biochemical detection method according to an embodiment;

[0059] The accompanying drawings are numerals as follows:

[0060] 10-fan-shaped base, 1-sample chamber, 2-sample quantitative chamber, 3-dilution chamber, 4-dilution quantitative chamber, 5-mixing chamber, 6-first detection chamber, 7-second detection chamber, L1-first microfluidic channel, L2-second microfluidic channel, L3-third microfluidic channel, R1-first reagent, R2-second reagent. DETAILED DESCRIPTION

[0061] Existing microfluidic biochemical detection discs typically feature only one or more parallel reaction chambers, where sample fluid enters and reacts for detection. This single reaction chamber configuration precludes dual-reagent reactions and prevents the removal of interfering substances prior to testing. Ultimately, these interfering substances can affect the detection structure, compromising the accuracy and precision of the test.

[0062] Based on this, the present application expands the mixing chamber into a reaction chamber, presets a first reagent for anti-interference reaction in the mixing chamber, and sets a second reagent for detection reaction in the detection chamber. This allows the sample to remove interfering substances related to the test items before testing, avoiding the impact of interfering substances on the test items, thereby improving the accuracy and precision of the test. Among them, the anti-interference ability of the dual reagent can remove endogenous interfering substances including substrates that can react with the reagents and interference from common blood lipids, bilirubin, ascorbic acid, and hemoglobin.

[0063] The present invention will be further described in detail below by means of specific embodiments in conjunction with the accompanying drawings. Similar elements in different embodiments are numbered with associated similar elements. In the following embodiments, many detailed descriptions are provided to enable the present application to be better understood. However, those skilled in the art will readily appreciate that some of the features may be omitted in different circumstances, or may be replaced by other elements, materials, or methods. In some cases, some operations related to the present application are not shown or described in the specification. This is to avoid the core portion of the present application being overwhelmed by excessive descriptions, and for those skilled in the art, it is not necessary to describe these related operations in detail. They will fully understand the related operations based on the description in the specification and the general technical knowledge in the art.

[0064] In addition, the features, operations, or characteristics described in the specification may be combined in any appropriate manner to form various embodiments. Furthermore, the steps or actions in the method description may be reordered or adjusted in a manner readily apparent to those skilled in the art. Therefore, the various sequences in the specification and drawings are provided solely for the purpose of clearly describing a particular embodiment and are not intended to be mandatory, unless otherwise specified.

[0065] The serial numbers assigned to components herein, such as "first," "second," etc., are used solely to distinguish the objects being described and do not convey any sequential or technical meaning. References to "connection" and "coupling" herein, unless otherwise specified, include both direct and indirect connections (couplings).

[0066] In one embodiment, a microfluidic biochemical detection disk is provided. This microfluidic biochemical detection disk is a centrifugal disk that can transfer liquids such as sample liquid, diluent, and reaction solution to other chambers under the action of centrifugal force. This microfluidic biochemical detection disk is pre-installed with dual reagents, which can react with the sample in different chambers one after another, realizing the dual reagents' anti-interference ability.

[0067] Referring to Figure 1 , the microfluidic biochemical detection disc can be a fan-shaped base 10, which can be installed as a module in a centrifugal device. The fan-shaped base 10 can include a bottom shell and a top cover. The bottom shell has interconnected grooves, and the top cover covers the grooves. The bottom shell and the top cover enclose a corresponding cavity and flow.

[0068] The present microfluidic biochemical detection disc includes a sample chamber 1, a sample quantification chamber 2, a diluent chamber 3, a diluent quantification chamber 4, a mixing chamber 5 and a first detection chamber 6. The sample chamber 1, the sample quantification chamber 2, the diluent chamber 3, the diluent quantification chamber 4, the mixing chamber 5 and the first detection chamber 6 are all located in a fan-shaped base 10. Among them, the sample chamber 1 and the diluent chamber 3 are closest to the centrifugal center, the sample quantification chamber 2 and the diluent quantification chamber 4 are next, the mixing chamber 5 is relatively far away from the centrifugal center, and the first detection chamber 6 is located at the position farthest from the centrifugal center. So that under the action of centrifugal force, the sample liquid can flow in sequence along the sample chamber 1, the sample quantification chamber 2, the mixing chamber 5 and the first detection chamber 6, and the diluent can flow in sequence along the diluent chamber 3, the diluent quantification chamber 4, the mixing chamber 5 and the first detection chamber 6.

[0069] The sample chamber 1 is used to accommodate the sample liquid. The user can manually add the sample liquid into the sample chamber 1 or transfer the sample liquid into the sample chamber 1 through automated equipment such as a sampling needle.

[0070] Sample quantification chamber 2 is connected to sample chamber 1, optionally via a microfluidic channel. Centrifugal force forces the sample fluid in sample chamber 1 into sample quantification chamber 2. Sample quantification chamber 2 has a preset volume, and different sample quantification chambers 2 can be configured to meet specific testing requirements.

[0071] The diluent chamber 3 is used to accommodate the diluent. Similarly, the user can manually add the diluent into the diluent chamber 3, or transfer the diluent into the diluent chamber 3 through automated equipment such as a sampling needle.

[0072] The diluent dosing chamber 4 is connected to the diluent chamber 3, and the two chambers may be connected via a microfluidic channel. Under the action of centrifugal force, the diluent in the diluent chamber 3 can enter the diluent dosing chamber 4. The diluent dosing chamber 4 has a preset volume, and the volume ratio between the diluent dosing chamber 4 and the sample dosing chamber 2 is preset.

[0073] The mixing chamber 5 can be connected to the sample quantification chamber 2 through the first microfluidic channel L1, and the mixing chamber 5 can also be connected to the diluent quantification chamber 4 through the second microfluidic channel L2. Under the action of centrifugal force, the quantitative sample liquid in the sample quantification chamber 2 and the quantitative diluent in the diluent quantification chamber 4 will enter the mixing chamber 5.

[0074] A first reagent R1 is pre-placed in the mixing chamber 5. The mixing chamber 5 provides a place for mixing and reacting the sample liquid, diluent, and first reagent R1 to form a first reaction liquid. The first reagent R1 is used to mix and react with the sample liquid and diluent to at least achieve the purpose of removing interfering substances in the sample liquid. Among them, the interfering substance includes at least one of blood lipids, bilirubin, ascorbic acid, and hemoglobin. Different first reagents R1 can be set for different test items to remove interfering substances related to the test items. The following text will list in detail a number of specific test items, and anti-interference treatment is performed using specific first reagents R1.

[0075] The first detection chamber 6 is connected to the mixing chamber 5 via the third microfluidic channel L3. A second reagent R2 is pre-placed in the first detection chamber 6. The first detection chamber 6 provides a venue for the reaction between the first reaction liquid and the second reagent to form the second reaction liquid. Under the action of centrifugal force, the first reaction liquid formed in the mixing chamber 5 will enter the first detection chamber 6 and react with the second reagent R2. The first detection chamber 6 is also equipped with a corresponding detection mechanism, which can perform optical detection on the second reaction liquid (sample) formed in the first detection chamber 6 to obtain the corresponding detection results.

[0076] The working principle and steps of the microfluidic biochemical detection disc in this embodiment are as follows:

[0077] In the first step, centrifugation is performed, and the diluent and sample solution enter the sample quantification chamber 2 and diluent quantification chamber 4, respectively, for precise quantification. Once the quantification chambers are full, the disk stops rotating, and the diluent and sample solution in the quantification chambers are drawn into the disk mixing chamber 5 by the capillary forces of the first microfluidic channel L1 and the second microfluidic channel L2, respectively.

[0078] The second step involves centrifugation, completely centrifuging the diluent and sample solution in the quantitative chamber into the mixing chamber 5. The disk's rotational speed is adjusted to achieve thorough mixing of the liquids. The first reagent R1 is dissolved in this step, reacting with the sample solution, and the incubation period provides anti-interference protection. After the incubation is complete, the disk stops rotating, and the liquid in the mixing chamber is drawn by capillary force from the third microfluidic channel L3 into the disk's first detection chamber 6.

[0079] The third step is to perform centrifugation to completely throw the mixed liquid in the mixing chamber 5 into the first detection chamber 6 of the disc. The liquid is fully mixed by adjusting the rotation direction of the disc. The second reagent R2 is dissolved in this step and reacts with the mixed liquid, causing the absorbance to change.

[0080] In this embodiment, since the microfluidic biochemical detection disc has a mixing chamber 5 and a first detection chamber 6, the two chambers are distributed in series, and the two chambers are preset with a first reagent R1 and a second reagent R2. The sample liquid can react with the first reagent R1 and the second reagent R2 in different chambers in sequence, so that the sample can first be subjected to an anti-interference reaction and then a detection reaction, thereby improving the accuracy of sample detection.

[0081] In one embodiment, the first detection chamber 6 may include at least two, at least two first detection chambers 6 arranged in parallel, the at least two first detection chambers 6 connected to the mixing chamber 5 via a third microchannel L3, and the second reagent R2 is pre-installed in each of the at least two first detection chambers 6. Under the action of centrifugal force, the first reaction liquid formed in the mixing chamber 5 will enter the at least two first detection chambers 6 and react with the second reagent R2, ultimately forming at least two samples. The detection mechanism can perform optical detection on the at least two samples separately to obtain at least two detection results. The processing unit can also average the at least two detection results, and use the average value as the final detection result, which can reduce errors and improve the accuracy and precision of the detection.

[0082] The number of first detection cavities 6 provided can be considered according to factors such as the size of the microfluidic biochemical detection disc. Providing more first detection cavities 6 within a sufficient area is beneficial to improving the accuracy and precision of the detection.

[0083] In one embodiment, the first reagent R1 may be a first reagent freeze-dried pellet made from a first finished liquid biochemical reagent, and the second reagent R2 may be a second reagent freeze-dried pellet made from a second finished liquid biochemical reagent.

[0084] Among them, the first reagent freeze-dried ball and the second reagent freeze-dried ball respectively have a preset volume, the first detection chamber has a preset volume, the concentration of the first reagent freeze-dried ball and the concentration of the second reagent freeze-dried ball are respectively consistent with the concentration of the liquid biochemical reagent in the full-automatic biochemical analyzer, and the concentration of the second reaction liquid is consistent with the concentration of the working liquid in the full-automatic biochemical analyzer.

[0085] Lyophilized reagent pellets can be directly lyophilized using finished liquid biochemical reagents without adjusting reagent concentrations. By controlling the number and volume of lyophilized pellets, the concentration of the first reagent, R1, is consistent with that in the liquid biochemical working solution. The volume of the reaction wells and the volume of the second reagent, R2, lyophilized pellets are adjusted to ensure the concentration of the second reagent, R2, is consistent with that in the liquid biochemical working solution. For example, if the R1:R2 ratio in the ALT liquid reagent working solution is 4:1, the first reagent, R1, will be diluted 4 / 5 and the second reagent, R2, will be diluted 1 / 5 in the working solution. By ensuring that the volume of the first reagent, R1, lyophilized pellets, accounts for 4 / 5 of the mixed solution and 1 / 5 of the reaction well volume, the reaction system and concentration of the liquid biochemical reagent are perfectly matched, ensuring that the reagent performance in the retest disk is completely consistent with that of the liquid biochemical reagent, reducing the workload and unknown risks of reagent development.

[0086] In one embodiment, only one of the first reagent R1 and the second reagent R2 can be configured as a lyophilized pellet made from a finished liquid biochemical reagent, while the other can be prepared by other methods. This can reduce the development of at least one reagent and has certain advantages and feasibility.

[0087] In one embodiment, the microfluidic biochemical detection disc may include at least one detection unit, and each detection unit includes one of the above-mentioned sector-shaped substrates 10 .

[0088] A detection unit on a microfluidic biochemical detection disc is generally configured to correspond to a detection item, and the volume ratio of the sample quantitative cavity 2 and the diluent quantitative cavity 4 on the detection unit is set to be adapted to the corresponding detection item. The volume of the sample quantitative cavity 2 and the volume of the diluent quantitative cavity 4 in a specific ratio can improve the accuracy of the detection of the item and reduce the opening of the reagent. For different detection items, the volume of the sample quantitative cavity 2 and the volume of the diluent quantitative cavity 4 in different ratios are generally required. Therefore, a plurality of detection units designed for different detection items have different volume ratios of the sample quantitative cavity 2 and the diluent quantitative cavity 4. Among them, the volume of the sample quantitative cavity 2 and the volume of the diluent quantitative cavity 4 can be different, or one of the volume of the sample quantitative cavity 2 and the volume of the diluent quantitative cavity 4 can be different or the same.

[0089] For example, a microfluidic biochemical detection disk can be equipped with six detection units, distributed along the centrifugal center and sharing the same centrifugal center. Each of the six detection units corresponds to a specific test item, allowing the microfluidic biochemical detection disk to perform a total of six different test items. Designing each detection unit to correspond to only one test item facilitates the targeted setting of the ratio between the sample quantification chamber 2 and the diluent quantification chamber 4, as well as the preset volumes and ratios of the first reagent R1 and the second reagent R2, to achieve optimal reaction conditions for the sample solution, thereby improving test accuracy.

[0090] In one embodiment, a first reaction liquid accommodating chamber is provided between the mixing chamber 5 and the first detection chamber 6. The mixing chamber 5 is connected to the first reaction liquid accommodating chamber via a microchannel, and the first reaction liquid accommodating chamber is connected to the first detection chamber 6 via a microchannel. The mixing chamber 5 merely provides a mixing function, providing a place for the sample liquid and the diluent to mix to form a diluted sample liquid. The first reagent R1 is pre-placed in the first reaction liquid accommodating chamber, which provides a place for the first reagent R1 to react with the diluted sample liquid to form the first reaction liquid. Under the action of centrifugal force, the first reaction liquid in the first reaction liquid accommodating chamber can enter the first detection chamber 6.

[0091] The dilution and mixing of the sample liquid and the reaction of the first reagent R1 are separated in two different chambers, so that the dilution and anti-interference treatment of the sample liquid can also be achieved.

[0092] Referring to Figure 2 , in one embodiment, the microfluidic biochemical detection disc further includes a second detection chamber 7, which is arranged in parallel with the first detection chamber 6. The second detection chamber 7 is also connected to the mixing chamber 5 or the first reaction liquid accommodating chamber via the third microchannel L3. The second detection chamber 7 is configured to receive the first reaction liquid output from the mixing chamber 5 or the first reaction liquid accommodating chamber. The second detection chamber 7 is empty and does not contain any reagents.

[0093] The second detection chamber 7 is also used for optical detection by the detection mechanism. The optical detection mechanism is used to detect the first reaction liquid. The absorbance of the first reaction liquid can be read as the starting point of the reaction and participate in concentration calculation.

[0094] The second detection chamber 7 can be closer to the mixing chamber 5 or the first reaction liquid holding chamber relative to the first detection chamber 6, so that the first reaction liquid output from the mixing chamber 5 or the first reaction liquid holding chamber can first enter the second detection chamber 7, ensuring that when the second reaction liquid is formed in the first detection chamber 6 for detection, the second detection chamber 7 has the first reaction liquid for detection together.

[0095] The provision of the second detection chamber 7 can assist in the detection and calculation of the sample, and is beneficial to improving the accuracy of the detection and calculation.

[0096] In one embodiment, the microfluidic biochemical detection disc is further provided with a plurality of waste liquid chambers. For example, the third microchannel L3 is connected to a waste liquid chamber, and the waste liquid chamber is used to collect excess first reaction liquid.

[0097] The waste liquid chamber can also be communicated with the sample chamber and the diluent chamber, and the waste liquid chamber is used to collect excess sample liquid and diluent.

[0098] Multiple waste liquid chambers can be connected to form one chamber, which is convenient for reducing the space occupied by the waste liquid chamber.

[0099] In one embodiment, a microfluidic life detection method is provided, and the microfluidic life detection method uses the above-mentioned microfluidic life detection disc for detection.

[0100] Referring to Figure 3, the present microfluidic life detection method includes the following steps:

[0101] S01: Sample quantification and dilution quantification;

[0102] The centrifugal device drives the sample liquid in the sample chamber 1 into the sample quantitative chamber 2, and at the same time drives the diluent in the diluent chamber 3 into the diluent quantitative chamber 4, and quantitatively prepares the sample liquid and diluent in a certain ratio respectively;

[0103] S02: first reaction of sample;

[0104] The centrifugal device drives the sample liquid in the sample quantitative chamber 2 and the diluent in the diluent quantitative chamber 4 into the mixing chamber 5;

[0105] The first reagent R1 preset in the mixing chamber is mixed with the sample solution and the diluent and reacts to form a first reaction solution;

[0106] The first reagent R1 is used to react with the sample solution and diluent to at least remove interfering substances from the sample solution. Interfering substances include at least one of blood lipids, bilirubin, ascorbic acid, and hemoglobin. Different first reagents R1 can be used for different test items to remove interfering substances related to the test item.

[0107] S03: second sample reaction;

[0108] The centrifugal device drives the first reaction liquid in the mixing chamber 5 into the first detection chamber 6. After the mixing chamber 5 receives the first reaction liquid, the centrifugal device drives the first reaction liquid into the first detection chamber 6 again after a preset interval. This interval operation allows sufficient reaction time between the sample and the first reagent R1 to form a complete reaction.

[0109] The second reagent R2 pre-placed in the first detection chamber 6 mixes and reacts with the first reaction liquid to form a second reaction liquid;

[0110] S04: Optical inspection.

[0111] The detection device measures the second reaction liquid in the first detection chamber 6 to obtain a detection result.

[0112] In one embodiment, the centrifugal device drives the first reaction liquid in the mixing chamber 5 into at least two first detection chambers 6 , forming at least two second reaction liquids in the at least two first detection chambers 6 .

[0113] The detection mechanism performs optical detection on at least two reaction solutions, obtaining at least two detection results. The at least two detection results are averaged, and the average of the multiple detection results is used as the final detection result. Calculating the final detection result by averaging can eliminate false triggers in individual detections, thereby improving detection accuracy.

[0114] In one embodiment, the centrifugal device drives the first reaction liquid in the mixing chamber 5 into the first detection chamber 6 , and at the same time, the centrifugal device drives the first reaction liquid in the mixing chamber 5 into the second detection chamber 7 .

[0115] After or before testing the second reaction liquid in the first detection chamber 6, the detection mechanism tests the first reaction liquid in the second detection chamber 7. The first reaction liquid is in an initial state before the second reaction of the sample liquid, and can provide initial data for the test item, which is helpful for assisting in calculating the final result of the test item and improving the accuracy of the test.

[0116] In one embodiment, the sample liquid may be directly injected into the sample quantitative chamber 2 and the diluent may be directly injected into the diluent quantitative chamber 4, that is, the first step of the centrifugal quantitative operation may be omitted.

[0117] In one embodiment, the microfluidic biochemical detection disc is applied to a specific biochemical detection project, taking the reagent ALT as an example.

[0118] Serum ALT catalyzes the transamination reaction of alanine and α-ketoglutarate to generate pyruvate and L-glutamate. The generated pyruvate is oxidized to the reduced form of nicotinamide adenine dinucleotide (NADH) by lactate dehydrogenase (LDH) to oxidized form of nicotinamide adenine dinucleotide (NAD+). NADH has strong light absorption at a wavelength of 340nm, while NAD+ has no absorption. In the case of excess substrate, the rate of pyruvate generation is proportional to the serum ALT concentration, and the rate of NADH decrease is proportional to the rate of pyruvate generation. Therefore, the serum ALT activity concentration can be measured by monitoring the rate of NADH decrease. The enzyme-coupled reaction formula for ALT determination is as follows:

[0119]

[0120]

[0121] The first reagent R1 and the second reagent R2 of the liquid alanine aminotransferase detection reagent (ALT) were made into freeze-dried pellets, respectively, and used with an improved centrifugal microfluidic disk. NADH and LDH were added to the first reagent R1 to consume endogenous α-keto acids, and a surfactant such as Triton-X100 was added to enhance the anti-interference ability against lactones.

[0122] After the sample and diluent are mixed, the first reagent R1 is dissolved. After incubation for a period of time, the first reagent R1 consumes endogenous α-keto acids and other interfering substances. The mixture is then centrifuged and directed into the first detection chamber 6, where the second reagent R2 is dissolved. The decrease in NADH in the first detection chamber 6, measured by the rate of decrease in absorbance at 340 nm, is monitored to determine the serum ALT activity concentration. Adding the same second reagent R2 to two first detection chambers 6 allows for the simultaneous averaging of two results. This allows for stronger interference resistance during retesting, resulting in more accurate results.

[0123] Among them, the ALT reagent formula is as follows:

[0124] First reagent R1:

[0125] Tris (hydroxymethylaminomethane) 10-100 mmol / L

[0126] pH(37~)7-8

[0127] EDTA-2NA 1-10g / L

[0128] NADH 0.1-0.3mmol / L

[0129] LDH 1000-2000U / L

[0130] Triton X-100 1-10g / L

[0131] Second reagent R2:

[0132] Tris (hydroxymethylaminomethane) 10-100 mmol / L

[0133] EDTA-2NA 1-10g / L

[0134] α-ketoglutaric acid 10-30mmol / L

[0135] L-alanine 100-1000mmol / L

[0136] Pyridoxal phosphate 0.1-0.5mmol / L

[0137] Excipient component (10-100 g / L): can be mannitol, inositol, a polyol; can also be a high molecular polymer, such as dextran, polyethylene glycol, water-soluble starch.

[0138] Protective agent component (10-100 g / L): can be a non-crystalline sugar, such as trehalose, sucrose; can also be a macromolecular protein, such as bovine serum albumin.

[0139] Anti-interference method: NADH and LDH are added to the first reagent R1 to consume endogenous α-keto acids, and a surfactant such as Triton-X100 is added to enhance the anti-interference ability against lactones.

[0140] Testing method: A centrifugal microfluidic disk is designed in a fan shape. A lyophilized bead of the first reagent R1 is placed in the mixing chamber 5. Three reaction wells are stored: one blank well and two reaction wells containing the lyophilized bead of the second reagent R2 (i.e., two first detection chambers 6 and one second detection chamber 7). Two sets of data are obtained and averaged. This solution is highly feasible and well-suited for retesting individual clinical items. It offers higher retest accuracy and stronger anti-interference capabilities than a large disk.

[0141] The test data is as follows:

[0142] (1) Precision

[0143]

[0144]

[0145] (2) Linear range

[0146]

[0147]

[0148] (3) Concentration

[0149]

[0150] (4) Anti-interference ability, including four different detection items

[0151]

[0152]

[0153]

[0154]

[0155]

[0156]

[0157] From the above test results, it can be seen that the use of the present microfluidic biochemical detection disc for dual-reagent anti-interference detection can eliminate specific interfering substances and improve the accuracy of the detection results.

[0158] In one embodiment, other liquid bioreagents can be used in projects with dual reagents, such as Na, Urea, Crea, etc. The following are some examples of anti-interference:

[0159] By adding the masking agent cryptand to the Na reagent (R1), the sodium ion concentration in the serum can be reduced to 55mmol / L during the incubation process, and the K+ / Na+ selectivity can be increased to 600:1, thereby eliminating the interference of sodium ions.

[0160] Incubation of the sample with Urea reagent (R1) removes interference from NH4+, which can cause the results to be biased high.

[0161] Incubation of the sample with Crea reagent (R1) removes interference from endogenous creatine, which can cause inflated results.

[0162] All reagents based on the Trinder reaction principle, such as TC, TG, LDL-C, UA, and GLU-O, will be interfered with by bilirubin and vitamin C. Adding potassium ferrocyanide and ascorbic acid oxidase to the first reagent R1 can eliminate the interference.

[0163] The above examples are used to illustrate the present invention, which are only used to help understand the present invention and are not intended to limit the present invention. Those skilled in the art can make several simple deductions, modifications or substitutions based on the concept of the present invention.

Claims

1. A microfluidic biochemical detection disc, characterized in that: include: A sample chamber, used for containing a sample liquid; A sample quantification cavity, connected to the sample cavity, for quantifying the sample liquid; A diluent chamber, used for containing a diluent; a diluent quantitative chamber, connected to the diluent chamber, for quantitatively measuring the diluent; A mixing chamber is connected to the sample quantitative chamber through a first microchannel and is connected to the diluent quantitative chamber through a second microchannel; a first reagent is pre-placed in the mixing chamber, and the mixing chamber provides a place for mixing and reacting the sample liquid, the diluent and the first reagent to form a first reaction liquid; The first detection chamber is connected to the mixing chamber through the third microchannel; the second reagent is preset in the first detection chamber, and the first detection chamber provides a place for the first reaction solution and the second reagent to react to form the second reaction solution.

2. A microfluidic biochemical detection disc, characterized in that: include: A sample chamber, used for containing a sample liquid; A sample quantification cavity, connected to the sample cavity, for quantifying the sample liquid; A diluent chamber, used for containing a diluent; a diluent quantitative chamber, connected to the diluent chamber, for quantitatively measuring the diluent; A mixing chamber, which is connected to the sample chamber through a first microchannel and is connected to the diluent chamber through a second microchannel; the mixing chamber is used to provide a place for mixing the sample liquid and the diluent to form a diluted sample; a first reaction liquid containing chamber, connected to the mixing chamber; a first reagent is pre-installed in the first reaction liquid containing chamber, the first reaction liquid containing chamber is used to receive the diluted sample output from the mixing chamber, and the first reaction liquid containing chamber provides a place for the reaction between the diluted sample and the first reagent to form a first reaction liquid; The first detection chamber is connected to the first reaction liquid containing chamber through the third microchannel, and is used to receive the first reaction liquid output from the first reaction liquid containing chamber; the first detection chamber is pre-installed with a second reagent, and the first detection chamber provides a place for the reaction between the first reaction liquid and the second reagent to form a second reaction liquid required for project detection.

3. A microfluidic biochemical detection disc, used to perform at least one test on a sample to be tested, characterized in that: The microfluidic biochemical detection disc comprises at least one detection unit, and the detection unit comprises: A sample chamber, used for containing a sample liquid; A sample quantification cavity, connected to the sample cavity, for quantifying the sample liquid; A diluent chamber, used for containing a diluent; a diluent quantitative chamber, connected to the diluent chamber, for quantitatively measuring the diluent; A mixing chamber, which is connected to the sample chamber through a first microchannel and is connected to the diluent chamber through a second microchannel; the mixing chamber is used to provide a place for mixing the sample liquid and the diluent to form a diluted sample; a first reaction liquid containing chamber, connected to the mixing chamber; a first reagent is pre-installed in the first reaction liquid containing chamber, the first reaction liquid containing chamber is used to receive the diluted sample output from the mixing chamber, and the first reaction liquid containing chamber provides a place for the reaction between the diluted sample and the first reagent to form a first reaction liquid; The first detection chamber is connected to the first reaction liquid containing chamber through the third microchannel, and is used to receive the first reaction liquid output from the first reaction liquid containing chamber; the first detection chamber is pre-installed with a second reagent, and the first detection chamber provides a place for the reaction between the first reaction liquid and the second reagent to form a second reaction liquid required for project detection.

4. A microfluidic biochemical detection disc, used to perform at least one item detection on a sample to be tested, characterized in that: The microfluidic biochemical detection disc comprises at least one detection unit, and the detection unit comprises: A sample chamber, used for containing a sample liquid; A sample quantification cavity, connected to the sample cavity, for quantifying the sample liquid; A diluent chamber, used for containing a diluent; a diluent quantitative chamber, connected to the diluent chamber, for quantitatively measuring the diluent; A mixing chamber is connected to the sample quantitative chamber through a first microchannel, and is connected to the diluent quantitative chamber through a second microchannel; a first reagent is pre-placed in the mixing chamber, and the mixing chamber provides a place for mixing and reacting the sample liquid, the diluent and the first reagent to form a first reaction liquid; The first detection chamber is connected to the mixing chamber through the third microchannel; the second reagent is preset in the first detection chamber, and the first detection chamber provides a place for the first reaction solution and the second reagent to react to form the second reaction solution.

5. The microfluidic biochemical detection disc according to claim 3 or 4, characterized in that: One detection unit corresponds to one detection item.

6. The microfluidic biochemical detection disc according to any one of claims 1 to 5, characterized in that: The first reagent is used to react with the sample solution and the diluent to at least achieve the purpose of removing interfering substances in the sample solution.

7. The microfluidic biochemical detection disc according to claim 6, characterized in that: The interfering substance includes at least one of blood lipids, bilirubin, ascorbic acid and hemoglobin.

8. The microfluidic biochemical detection disc according to claim 3 or 4, characterized in that: The detection units include at least two, and the volume of the sample quantitative chamber and / or the volume of the diluent quantitative chamber of any detection unit is different from the volume of the sample quantitative chamber and / or the volume of the diluent quantitative chamber of other detection units.

9. The microfluidic biochemical detection disc according to any one of claims 1 to 8, characterized in that: The first reagent is a first reagent freeze-dried pellet made from a first finished liquid biochemical reagent, and / or the second reagent is a second reagent freeze-dried pellet made from a second finished liquid biochemical reagent.

10. The microfluidic biochemical detection disc according to claim 9, characterized in that: The first reagent freeze-dried ball and the second reagent freeze-dried ball have preset volumes respectively, the first detection chamber has a preset volume, the concentration of the first reagent freeze-dried ball and the concentration of the second reagent freeze-dried ball are respectively consistent with the concentration of the liquid biochemical reagent in the full-automatic biochemical analyzer, and the concentration of the second reaction solution is consistent with the concentration of the working solution in the full-automatic biochemical analyzer.

11. The microfluidic biochemical detection disc according to any one of claims 1 to 10, characterized in that: The second reagent reacts with the first reaction liquid to obtain a sample, and the sample is optically detected to obtain a corresponding detection result.

12. The microfluidic biochemical detection disc according to any one of claims 1 to 11, characterized in that: The mixing chamber or the first reaction liquid containing chamber is connected to at least two first detection chambers through the third microchannel, and the at least two first detection chambers respectively provide places for the reaction of the first reaction liquid and the second reagent to form at least two second reaction liquids.

13. The microfluidic biochemical detection disc according to any one of claims 1 to 4, characterized in that: It also includes a second detection chamber, which is connected to the mixing chamber or the first reaction liquid containing chamber through the third microchannel and is used to receive the first reaction liquid output from the mixing chamber or the first reaction liquid containing chamber.

14. The microfluidic biochemical detection disc according to any one of claims 1 to 13, characterized in that: It comprises a fan-shaped matrix, wherein the sample chamber, the diluent chamber, the mixing chamber, the first detection chamber, the first microfluidic channel, the second microfluidic channel and the third microfluidic channel are located in the fan-shaped matrix.

15. A microfluidic biochemical detection method, using a microfluidic biochemical detection disc for detection, characterized in that: The steps include: The centrifugal device drives the sample liquid in the sample quantitative chamber and the diluent in the dilution quantitative chamber into the mixing chamber; The first reagent pre-placed in the mixing chamber is mixed and reacted with the sample solution and the diluent to form a first reaction solution; The centrifugal device drives the first reaction liquid in the mixing chamber to enter the first detection chamber; The second reagent preset in the first detection chamber is mixed and reacted with the first reaction liquid to form a second reaction liquid; The detection device measures the second reaction liquid in the first detection chamber to obtain a detection result.

16. The microfluidic biochemical detection method according to claim 15, characterized in that: After the mixing chamber obtains the first reaction liquid, the centrifugal device drives the first reaction liquid into the first detection chamber again after a preset time interval.

17. The microfluidic biochemical detection method according to claim 15 or 16, characterized in that: The centrifugal device drives the first reaction liquid into at least two of the first detection chambers to form at least two portions of the second reaction liquid; the detection mechanism measures the at least two portions of the second reaction liquid respectively to obtain at least two detection results.

18. The microfluidic biochemical detection method according to claim 17, characterized in that: An average value is taken for the at least two detection results, and the average value is used as the final detection result.

19. The microfluidic biochemical detection method according to any one of claims 15 to 18, characterized in that: The first reagent is used to react with the sample solution and the diluent to at least achieve the purpose of removing interfering substances in the sample solution.

20. The microfluidic biochemical detection method according to claim 19, characterized in that: The interfering substance includes at least one of blood lipids, bilirubin, ascorbic acid and hemoglobin.

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