High-efficiency microfluidic detection chip with one-time injection

Abstract

The present invention discloses a one-time injection high-efficiency microfluidic detection chip, the detection chip has a rotation center, and the detection chip includes at least one detection unit that can independently perform detection, the detection unit includes a mixed cultivation tank for providing a reaction space, and a solid reagent storage tank storing N different solid reagents is arranged in the mixed cultivation tank; the solid reagent includes a first, a second, ..., and an Nth solid reagent, all of which are coated with a protective film on the outside, and the outside of the Nth solid reagent is sequentially coated with a first protective film, a second protective film, ..., and an Nth protective film from the outside to the inside. The present invention can directly use whole blood, peripheral blood, serum, and plasma samples without sample pretreatment; it can process samples of high hematocrit / polycythemia; only one-time injection of the sample is required to automatically complete complex sample addition and reaction steps; it can reduce the overall reaction time without affecting the detection efficiency, and quickly complete blood testing.

Description

Technical Field

[0001] The invention relates to the technical field of microfluidic chips, and in particular to a one-time injection high-efficiency microfluidic detection chip. Background Art

[0002] In order to simplify the defects of traditional detection steps that are lengthy and complicated, microfluidic chip technology is often used for detection (such as: biochemical detection, immune detection, molecular detection, etc.). However, there are still many difficulties that cannot be overcome in microfluidic detection chips, such as: (1) It is impossible to handle high blood volume ratio samples. (2) The sample addition steps are complicated (the detection process often requires multiple sample addition steps, such as: samples, quality control products / calibrators, diluents, cleaning solutions, various reagents, etc.). For this reason, how to design the structure of the microfluidic chip to integrate the traditional lengthy and complicated detection scheme into a simple, fast, efficient, highly integrated, and suitable for industrial mass production detection scheme is an important topic. Summary of the invention

[0003] In order to solve at least one of the above technical problems, the present invention provides a high-efficiency microfluidic detection chip with one-time injection.

[0004] The technical solution adopted by the present invention is: a high-efficiency microfluidic detection chip for one-time injection, the detection chip has a rotation center, and the detection chip includes at least one detection unit that can independently perform detection, the detection unit includes a mixed cultivation tank for providing a reaction space, and a solid reagent storage tank storing N different solid reagents is arranged in the mixed cultivation tank;

[0005] The solid reagent includes first, second, ..., and Nth solid reagents, each of which is coated with a protective film on the outside. The first solid reagent is coated with a first protective film on the outside, and the second solid reagent is coated with a first protective film and a second protective film in sequence from the outside to the inside. By analogy, the Nth solid reagent is coated with a first protective film, a second protective film, ..., and an Nth protective film in sequence from the outside to the inside.

[0006] The first to Nth protective films can all be dissolved in the mixed incubation tank under the action of the dissolving reagent provided by the detection unit, so that the protective films coated on the outside of the first to Nth solid reagents have different dissolution times.

[0007] Preferably, it further comprises at least one dissolving reagent storage tank and at least one reaction reagent storage tank communicated with the mixed incubation tank;

[0008] The first to Nth protection films are formed of the same material that can be dissolved by the dissolving reagent stored in the dissolving reagent storage tank.

[0009] Preferably, the liquid reagents in the dissolving reagent storage tank and the reaction reagent storage tank are sealed and packaged in pre-sealed bags, and the pre-sealed bags can be partially ruptured under the action of external force to release the liquid reagents therein and provide them to the mixed incubation tank.

[0010] Preferably, the body of the detection chip comprises a bottom plate, an upper cover and a microfluidic channel opened on the bottom plate, and the upper cover is sealed and buckled on the bottom plate, so that the microfluidic channel between the bottom plate and the upper cover forms the detection unit;

[0011] The upper cover above the liquid reagent storage tank is provided with a puncture hole, and the pre-sealed bag can be punctured through the puncture hole by an external puncturing device to release the liquid reagent therein.

[0012] Preferably, it is characterized in that the detection unit further comprises a whole blood storage tank and a whole blood injection hole connected to the whole blood storage tank for injecting the blood sample to be tested, and a blood filter membrane is arranged in the whole blood storage tank.

[0013] Preferably, the detection unit also includes a plasma serum quantitative tank connected to the whole blood storage tank through a whole blood transmission channel, a plasma serum transmission channel connecting the plasma serum quantitative tank to the mixed incubation tank, a waste liquid tank connected to the mixed incubation tank through a waste liquid transmission channel, and a vent connected to the waste liquid tank.

[0014] Preferably, the whole blood transmission channel is connected to a residual blood cell storage tank at a position upstream of the quantitative tank;

[0015] An overflow port is arranged on one side of the plasma serum quantitative tank close to the rotation center, and the overflow port is connected to a plasma serum waste liquid tank.

[0016] Preferably, the whole blood transmission channel comprises a radial segment channel connected to the outlet end of the whole blood storage tank and a circumferential segment channel connected to the end of the radial segment channel, the end of the circumferential segment channel is connected to the plasma serum quantitative tank, and the plasma serum quantitative tank is located on a side of the circumferential segment channel close to the rotation center;

[0017] The remaining blood cell storage groove is communicated with the middle part of the circumferential section channel, and the remaining blood cell storage groove is located on a side of the circumferential section channel away from the rotation center.

[0018] Preferably, a microfluidic valve is provided on the waste liquid transmission channel connected to the outlet end of the mixed cultivation tank.

[0019] Preferably, the plasma serum transmission channel has a first inverted U-shaped section, and the waste liquid transmission channel has a second inverted U-shaped section.

[0020] The present invention at least includes the following beneficial effects:

[0021] 1. The one-time injection high-efficiency microfluidic detection chip of the present invention can directly use whole blood, peripheral blood, serum, and plasma samples without sample pretreatment;

[0022] 2. The present invention is provided with a blood filter membrane, which can process samples with high hematocrit / polycythemia;

[0023] 3. The present invention pre-seales solid reagents and liquid reagents in the chip, and only requires one injection of the sample to automatically complete the complex sample addition and reaction steps;

[0024] 4. The present invention only requires a small amount of sample (tens of microliters) for detection;

[0025] 5. The present invention can also shorten the overall reaction time without affecting the detection performance, and quickly complete the blood test. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is a schematic diagram of the structure of the high-efficiency microfluidic detection chip for one-time injection of the present invention;

[0027] Figure 2 It is a structural schematic diagram of the detection unit of the present invention;

[0028] Figure 3 This is a schematic diagram of the principle of embodiment 1 of the present invention;

[0029] Figure 4 This is a schematic diagram of the principle of Example 2 of the present invention.

[0030] Description of reference numerals:

[0031] 1—whole blood injection hole; 2—whole blood storage tank; 3—blood filter membrane; 4—whole blood transmission channel; 5—residual blood cell storage tank; 6—plasma serum waste liquid tank; 7—plasma serum quantitative tank; 8—plasma serum transmission channel; 9—dissolving reagent storage tank; 10—first reaction reagent storage tank; 11—second reaction reagent storage tank; 12—mixing incubation tank; 13—solid reagent storage tank; 14—microfluidic valve; 15—waste liquid transmission channel; 16—waste liquid tank; 17—vent; 80—first inverted U-shaped section; 100—detection unit; 101—body of the detection chip; 102—rotation center; 150—second inverted U-shaped section. DETAILED DESCRIPTION

[0032] The present invention is further described in detail below in conjunction with embodiments so that those skilled in the art can implement the invention with reference to the description.

[0033] It should be understood that the terms such as “having”, “including” and “comprising” used herein do not exclude the existence or addition of one or more other elements or combinations thereof.

[0034] like Figure 1-2 As shown, a high-efficiency microfluidic detection chip for one-time injection in this embodiment has a rotation center 102, and the detection chip includes at least one detection unit 100 that can independently perform detection, and the detection unit 100 includes a mixed incubation tank 12 for providing a reaction space, and a solid reagent storage tank 13 storing N different solid reagents is arranged in the mixed incubation tank 12; wherein, the solid reagent storage tank can also store a variety of insoluble microspheres, such as magnetic or non-magnetic immune microspheres.

[0035] The solid reagent includes first, second, ..., and Nth solid reagents, all of which are coated with protective films on the outside. The first solid reagent is coated with a first protective film on the outside, and the second solid reagent is coated with a first protective film and a second protective film in sequence from the outside to the inside. By analogy, the Nth solid reagent is coated with a first protective film, a second protective film, ..., and an Nth protective film in sequence from the outside to the inside.

[0036] The first to Nth protective films can all be dissolved in the mixed incubation tank 12 under the action of the liquid reagent provided by the detection unit 100, so that the protective films coated on the outside of the first to Nth solid reagents have different dissolution times.

[0037] The high-efficiency microfluidic detection chip also includes at least one dissolving reagent storage tank 9 and at least one reaction reagent storage tank connected to the mixed incubation tank 12;

[0038] The first to Nth protection films are made of the same material that can be dissolved by the dissolving reagent stored in the dissolving reagent storage tank 9 .

[0039] In a preferred embodiment, the liquid reagents in the dissolving reagent and reaction reagent storage tanks are sealed and packaged in pre-sealed bags, which can be partially ruptured under the action of external force to release the liquid reagents therein and provide them to the mixed incubation tank 12.

[0040] The chip of the present invention is mainly used for blood sample detection. During the detection process, a variety of solid reagents are usually required to react with samples in sequence (liquid reagents also need to be added during the process). In the traditional scheme, they are added in sequence during the reaction, which is relatively cumbersome. In the present invention, the solid reagent is pre-installed in the solid reagent storage tank 13, and the liquid reagent is pre-packaged in the dissolving reagent storage tank 9 and the reaction reagent storage tank. Only one-time addition is required. During the reaction process, the pre-sealed bag is ruptured by operation to release the liquid reagent in sequence as required; the solid reagent has different dissolution times due to the design of the protective film, so that it can also react in sequence as required. Specific examples will be further explained in the following embodiments.

[0041] Among them, the protective film is designed to be used to coat solid reagents on the one hand to achieve the molding and packaging of solid reagents, and on the other hand, more importantly, to make different solid reagents have the required dissolution time. All protective films are made of the same material and are designed to be able to dissolve the dissolved reagents pre-packaged in the chip in a certain period of time. Obviously, the thickness of the protective film is positively correlated with the dissolution time, so different dissolution times can be obtained by controlling different protective film thicknesses and / or the number of protective film layers. Therefore, for the chip of the present invention, the relevant reaction reagents can be added at one time, and can react in sequence according to the design requirements. The above is the overall concept of the present invention. For a more specific design scheme for the protective film, the present invention also provides the following two specific embodiments.

[0042] First of all, it is important to understand that when the reagents in the reaction are solid reagents, the process in the reaction is: the solid reagent is dissolved first, and then the reaction is carried out after it is completely dissolved. Therefore, the time required for a single reaction includes the time for the solid reagent to dissolve plus the time for the reaction.

[0043] Example 1

[0044] In this embodiment, the design scheme is: after the previous reaction is completed, the next reaction starts (that is, the protective film outside the solid reagent participating in the next reaction is completely dissolved, and the solid reagent can enter the solution system to start the reaction), and several reactions are carried out completely in series.

[0045] For ease of understanding, relevant symbols and formulas are introduced below for description.

[0046] Assume that a total of N reactions need to be carried out in sequence: F1, F2, ..., F n ;

[0047] The solid reagents participating in N reactions are the first, second, ..., Nth solid reagents in sequence; the first solid reagent is coated with a first protective film on the outside, the second solid reagent is coated with a first protective film and a second protective film on the outside, and so on, the Nth solid reagent is coated with a first protective film, a second protective film, ..., an Nth protective film on the outside; all protective films can be dissolved by a dissolving reagent within a certain period of time;

[0048] The reaction process requires the first to Nth solid reagents to dissolve and react in sequence, that is, when the N-1th reaction is completed, all the protective films on the outside of the Nth solid reagent (the first, second, ..., N-1th, Nth protective films) are completely dissolved, so that the Nth solid reagent body can begin to dissolve and enter the reaction system to react;

[0049] The total time T1 required for the first reaction F1 is the dissolution time tm1 of the first protective film plus the reaction time tz1 of the first solid reagent body, that is, T1 = tm1 + tz1;

[0050] The total time T2 required for the second reaction F2 is the dissolution time tm1 of the first protective film and the dissolution time tm2 of the second protective film plus the reaction time tz2 of the first solid reagent body, that is, T2 = tm1 + tm2 + tz2; ...

[0051] Reaction F n-1 The total time required is T N-1 is the sum of the dissolution time of the first to N-1th protective films: (tm1+tm2+...+tm N-1 ) plus the reaction time tz of the N-1 solid reagent body N-1 , that is, T N-1 =(tm1+tm2+...+tm N-1 )+tz N-1 ;

[0052] Nth Reaction F n The total time required is T N is the sum of the dissolution time of the first to Nth protective films: (tm1+tm2+...+tm N-1 +tm N ) plus the reaction time tz of the Nth solid reagent body N , that is, T N =(tm1+tm2+...+tm N-1 +tm N )+tz N ;reference Figure 3 ;

[0053] Satisfies the following relationship: tm1+tm2+...+tm N-1+tm N =T N-1 =(tm1+tm2+...+tm N-1 )+tz N-1 ;

[0054] That is tm N =tz N-1 , that is, the dissolution time of the innermost protective film of the solid reagent in the latter reaction is exactly the same as the reaction time of the solid reagent body in the previous reaction, so by controlling the thickness of each protective film, the required reaction time of the protective film can be achieved. For example, for the first and second reactions, it satisfies: tm1+tm2=T1=tm1+tz1, tm2=tz1.

[0055] Total reaction time T=tm1+tz1+tz2+...+tz N-1 +tz N Compared with the conventional method, it is equivalent to only adding one more tm1 (dissolution time of the first protective film), but the reaction reagents are added at one time, which reduces the operation steps and complexity.

[0056] For example, in one embodiment, there are three reactions A, B, and C that need to be performed in sequence, and the reaction time of each reaction is 5 minutes, 10 minutes, and 15 minutes, respectively. In the three reactions, the solid reagents involved are solid reagent 1, solid reagent 2, and solid reagent 3, respectively;

[0057] The solid reagent 1 has a first protective film on the outside, the dissolution time of the first protective film is 1 minute, and the total reaction time of the solid reagent 1 is 4 minutes;

[0058] The solid reagent 2 has a first protective film and a second protective film from the outside to the inside. The dissolution time of the first protective film is 1 minute, so it is necessary to design the dissolution time of the second protective film to be 4 minutes, that is, the total reaction time of the solid reagent 2 is 5 minutes;

[0059] The solid reagent 3 has a first protective film, a second protective film, and a third protective film from the outside to the inside. The dissolution time of the first protective film is 1 minute, and the dissolution time of the second protective film is 4 minutes. It is necessary to design the dissolution time of the third protective film to be 5 minutes; that is, the total reaction time of the solid reagent 3 is 5 minutes.

[0060] During the reaction, solid reagent 1, solid reagent 2, and solid reagent 3 are pre-installed in the solid reagent storage tank 13 of the chip body. When the dissolved reagent immerses all the solid reagents, the first protective films on the outside of solid reagent 1, solid reagent 2, and solid reagent 3 start to react at the same time, and all the first protective films are completely dissolved after 1 minute; the solid reagent 1 body starts to react with the solution system (if a liquid reagent is needed at this time, the liquid reagent stored in the reaction reagent storage tank can be allowed to enter the solution system), and reaction A is completed after 4 minutes; at this time, the second protective film on the outside of solid reagent 2 and solid reagent 3 is completely dissolved (if a liquid reagent is needed at this time, the liquid reagent stored in the reaction reagent storage tank can be allowed to enter the solution system), the solid reagent 2 body starts to react with the solution system, and after 5 minutes, reaction B is completed; at this time, the third protective film on the outside of solid reagent 3 is completely dissolved (if a liquid reagent is needed at this time, the liquid reagent stored in the reaction reagent storage tank can be allowed to enter the solution system), the solid reagent 3 body starts to react with the solution system, and reaction C is completed after 5 minutes. The total time for the three reactions is 1+5+10+15=31 minutes, which is only 1 minute longer than the original time.

[0061] Furthermore, the time required for conventional chips also includes the time required for reagent injection because reagents need to be injected. For example, in the above embodiment, the time required is: 5+10+15+injection time. Assuming that the injection time for each reagent is 1 minute, there are 8 reaction units in this embodiment, and 3 reagents need to be injected, then the time required is: 8+5+8+10+8+15=54 minutes, while the method of this embodiment only requires 1+5+10+15=31 minutes.

[0062] For example, in an optional scheme, for the above-mentioned protective film, the material is trehalose film, the dissolving agent is deionized water, the thickness of the first protective film is 0.01-0.05 mm, and the dissolution time is 1 minute; the thickness of the second protective film is 4 times that of the first protective film, and the dissolution time is 4 minutes; the thickness of the third protective film is 5 times that of the first protective film, and the dissolution time is 5 minutes.

[0063] Example 2

[0064] Generally speaking, the detection reaction is mostly "tandem", which means first reacting with A, then reacting with B, then reacting with C, and so on. In order to maintain its detection sensitivity, the incubation / reaction time cannot be shortened.

[0065] That is, in the mixed culture tank 12, different reagents and samples need to be sequentially reacted to achieve detection, that is, usually after the previous reaction is completed, the reagents for the next reaction are injected to carry out the next reaction. For the case where solid reagents participate in the reaction, the process in the reaction is: the solid reagent is dissolved first, and then the reaction is carried out after it is completely dissolved. Therefore, the time required for a single reaction includes the time for the solid reagent to dissolve plus the time for the actual reaction. But in fact, only the previous reaction needs to be completed before the next reaction can be started, and the simultaneous dissolution will not affect the entire reaction, that is: the next reaction can be dissolved when the reaction of the previous reaction itself is carried out, and after the previous reaction is completed, the solid reagent of the next reaction is just completely dissolved. The scheme of this embodiment is based on this principle, and is carried out by further optimizing the structure of the protective film (mainly through thickness design, so that it has different dissolution times).

[0066] In the conventional scheme, the next reaction starts after the previous reaction is completed. Several reactions are completely serial, and the time required for the reactions is accumulated successively.

[0067] In this embodiment, not only the advantages of one-time addition as in Example 1 are achieved, but also several reactions can share the dissolution time, thereby realizing a partially parallel reaction, thereby reducing the time required for the reaction. The principle is: the solid reagent in the latter reaction enters the reaction system at the same time as the previous reaction, and when the previous reaction is completed, all protective films on the outside of the solid reagent in the latter reaction and the solid reagent body are completely dissolved, so when the previous reaction is completed, the latter reaction has already completed the dissolution and will react immediately, thereby saving the dissolution time of the solid reagent in the latter reaction.

[0068] Relevant formulas and symbols are introduced below for a more specific explanation.

[0069] Assume that a total of N reactions need to be carried out in sequence: F1, F2, ..., F n ;

[0070] The solid reagents participating in N reactions are the first, second, ..., Nth solid reagents in sequence; the first solid reagent is coated with a first protective film on the outside, the second solid reagent is coated with a first protective film and a second protective film on the outside, and so on, the Nth solid reagent is coated with a first protective film, a second protective film, ..., an Nth protective film on the outside; all protective films can be dissolved by a dissolving reagent within a certain period of time;

[0071] The reaction process requires the first to Nth solid reagents to dissolve and react in sequence, requiring that when the N-1th reaction is completed, all the protective films on the outside of the Nth solid reagent (the first, second, ..., N-1th, Nth protective films) and the Nth solid reagent are completely dissolved, so that the Nth solid reagent body can immediately enter the reaction system to react;

[0072] The total time T1 required for the first reaction F1 is the dissolution time tm1 of the first protective film plus the reaction time tz1 of the first solid reagent body, that is, T1=tm1+tz1; and the reaction time tz1 of the first solid reagent body specifically includes the dissolution time tb1 of the first solid reagent body plus the actual reaction time tf1 of the first solid reagent body and the system reagent, that is, tz1=tb1+tf1, so T1=(tm1+tb1)+tf1;

[0073] The total time T2 required for the second reaction F2 is the sum of the dissolution time tm1 of the first protective film and the dissolution time tm2 of the second protective film plus the reaction time tz2 of the second solid reagent body, that is, T2=tm1+tm2+tz2; and the reaction time tz2 of the second solid reagent body specifically includes the dissolution time tb2 of the second solid reagent body plus the actual reaction time tf2 of the second solid reagent body and the system reagent, that is, tz2=tb2+tf2, so T2=(tm1+tm2+tb2)+tf2; ...

[0074] Reaction F n-1 The total time required is T N-1 is the sum of the dissolution time of the first to N-1th protective films: (tm1+tm2+...+tm N-1 ) plus the reaction time tz of the N-1 solid reagent body N-1 , that is, T N-1 =(tm1+tm2+...+tm N-1 )+tz N-1 ; and the reaction time of the N-1 solid reagent body is tz N-1 Specifically including the dissolution time tb of the N-1 solid reagent body N-1 Add the actual reaction time tf of the N-1 solid reagent body and the system reagent N-1 , that is, tz N-1 =tb N-1 +tf N-1 , so T N-1 =(tm1+tm2+...+tm N-1 +tb N-1 )+tf N-1 ;

[0075] Nth Reaction Fn The total time required is T N is the sum of the dissolution time of the first to Nth protective films: (tm1+tm2+...+tm N-1 +tm N ) plus the reaction time tz of the Nth solid reagent body N , that is, T N =(tm1+tm2+...+tm N-1 +tm N )+tz N ; and the reaction time of the Nth solid reagent body is tz N- Specifically including the dissolution time tb of the Nth solid reagent body N Add the actual reaction time tf of the Nth solid reagent and the system reagent N , that is, tz N =tb N +tf N , so T N =(tm1+tm2+...+tm N-1 +tm N +tb N )+tf N ; can refer to Figure 4 ;

[0076] According to the above requirements, the following relationship is satisfied: the sum of the dissolution time of all protective films and the solid reagent body outside the Nth solid reagent in the Nth reaction (tm1+tm2+...+tm N-1 +tm N +tb N ) is equal to the total time T required for the N-1th reaction N-1 , that is, tm1+tm2+...+tm N-1 +tm N +tb N =T N-1 =(tm1+tm2+...+tm N-1 +tb N-1 )+tf N-1 , thus obtaining tm N +tb N =tb N-1 +tf N-1 , which means the sum of the dissolution time of the innermost protective film of the Nth solid reagent and the dissolution time of the Nth solid reagent body, the dissolution time of the N-1th solid reagent body and the reaction time of the N-1th solid reagent body. For example, for the first and second reactions, tm2+tb2=tb1+tf1 is satisfied.

[0077] Total reaction time T = tm1 + tb1 + tf1 + tf2 + ... + tfN-1 +tf N ; Compared with the normal sequential addition, the time saved = (tz1+tz2+...+tz N-1 +tz N )-(tm1+tb1+tf1+tf2+...+tf N-1 +tf N )=(tm1+tb1+tf1+tb2+tf2+...+tb N-1 +tf N-1 +tb N +tf N )-(tm1+tb1+tf1+tf2+...+tf N-1 +tf N )=tb2+...+tb N-1 +tb N -tm1.

[0078] Among them, it is necessary to understand that due to tm N +tb N ,tb N-1 、tf N-1 can only be positive numbers; so in general, tb N <tb N-1 +tf N-1 , that is to say, the dissolution time of the Nth solid reagent cannot be too long (cannot be greater than the sum of the dissolution time and reaction time of the solid reagent in the previous reaction). Usually, the solid reagents used in blood testing (such as immune microspheres, freeze-dried reagent beads, etc.) can obviously meet the above requirements. Therefore, there is generally no tb N >tb N-1 +tf N-1 (i.e., the Nth solid reagent body is particularly difficult to dissolve, and even exceeds the sum of the time required for the solid reagent body of the previous reaction to dissolve and react), the above scheme of the present invention is generally applicable. In addition, if there is tb in some extreme cases N >tb N-1 +tf N-1 The situation, that is, tb N -(tb N-1 +tf N-1 )=Δt, Δt>0, which means that after the N-1 reaction is completed, the Nth solid reagent body is not completely dissolved; in actual operation, this problem can be solved by conventional means, for example: using an external device to provide oscillation to promote the dissolution of the Nth solid reagent body, shorten the dissolution time, and make tb N <tb N-1 +tf N-1or adding Δt to the Nth reaction, that is, the Nth reaction is extended by Δt. In summary, the solution of this embodiment still has good universality.

[0079] For example, in one embodiment, there are three reactions A, B, and C that need to be performed in sequence, and the reaction time of each reaction is 5 minutes (solid reagent dissolution time + reaction time), 10 minutes, and 15 minutes, respectively. In the three reactions, the solid reagents involved are solid reagent 1, solid reagent 2, and solid reagent 3, respectively;

[0080] The solid reagent 1 has a first protective film on the outside, and the dissolution time of the first protective film is tm1=1 minute; since the reaction time tz1 of the solid reagent 1 body is 5 minutes in total, of which the dissolution time tb1 of the solid reagent 1 body is 2 minutes, the actual reaction time tf1 of the solid reagent 1 body is 3 minutes;

[0081] The outside of the solid reagent 2 has a first protective film and a second protective film in order from the outside to the inside. The dissolution time of the first protective film is tm1=1 minute, and the dissolution time of the solid reagent 2 is tb2=4 minutes. Therefore, it is necessary to design the dissolution time of the second protective film to be tm2=tb1+tf1-tb2=2+3-4=1 minute. Since the reaction time tz2 of the solid reagent 2 body is a total of 10 minutes, of which the dissolution time tb2 of the solid reagent 2 body is 4 minutes, the actual reaction time tf2 of the solid reagent 2 body is 6 minutes.

[0082] The outside of the solid reagent 3 has a first protective film, a second protective film, and a third protective film from the outside to the inside. The dissolution time of the first protective film is tm1=1 minute, the dissolution time of the second protective film is tm2=1 minute, and the dissolution time of the solid reagent 3 is tb3=6 minutes. Therefore, it is necessary to design the dissolution time of the third protective film to be tm3=tb2+tf2-tb3=4+6-6=4 minutes. Since the reaction time tz3 of the solid reagent 3 body is a total of 15 minutes, of which the dissolution time of the solid reagent 3 body is tb3=6 minutes, the actual reaction time of the solid reagent 3 body is tf3=9 minutes.

[0083] The total time required for the three reactions is T (1-3) =tm1+tb1+tf1+tf2+tf3=1+2+3+6+9=21 minutes;

[0084] Conventional sequential addition requires 5+10+15=30 minutes, so the solution of this embodiment saves 9 minutes, that is, saves 30% of the time.

[0085] Example 3

[0086] Based on the overall concept of the present invention or Example 1 or Example 2, further optimization is performed.

[0087] Reference Figure 1 and Figure 2 In this embodiment, the body 101 of the detection chip includes a bottom plate, an upper cover and a microfluidic channel opened on the bottom plate, and the upper cover is sealed and buckled on the bottom plate, so that the microfluidic channel between the bottom plate and the upper cover forms a detection unit 100;

[0088] A puncture hole is provided on the upper cover above the liquid reagent storage tank, and the pre-sealed bag can be punctured through the puncture hole by an external puncturing device to release the liquid reagent therein.

[0089] Of course, in other embodiments, the rupture of the pre-sealed bag can also be achieved by adopting an external squeezing force. For example, no puncture hole is provided, and the upper part of the liquid reagent storage tank is squeezed by an squeezing device, so that the pre-sealed bag is ruptured to release the liquid reagent therein.

[0090] In a further preferred embodiment, the detection unit 100 further includes a whole blood storage tank 2 and a whole blood injection hole 1 connected to the whole blood storage tank 2 for injecting a blood sample to be tested, and a blood filter membrane 3 is arranged in the whole blood storage tank 2. The blood filter membrane 3 is used to filter the whole blood sample and can filter out at least 80% of the blood cells.

[0091] The detection unit 100 also includes a plasma serum quantitative tank 7 connected to the whole blood storage tank 2 through a whole blood transmission channel 4, a plasma serum transmission channel 8 connecting the plasma serum quantitative tank 7 to the mixing incubation tank 12, a waste liquid tank 16 connected to the mixing incubation tank 12 through a waste liquid transmission channel 15, and a vent 17 connected to the waste liquid tank 16.

[0092] The whole blood transmission channel 4 is connected to the remaining blood cell storage tank 5 at a position upstream of the plasma serum quantitative tank 7;

[0093] An overflow port is provided on one side of the plasma serum quantitative tank 7 close to the rotation center 102, and the overflow port is connected to the plasma serum waste liquid tank 6. The plasma serum quantitative tank 7 is used to achieve quantification to provide a sample of a required volume.

[0094] The whole blood transmission channel 4 includes a radial segment channel connected to the outlet end of the whole blood storage tank 2 and a circumferential segment channel connected to the end of the radial segment channel, the end of the circumferential segment channel is connected to the plasma serum quantitative tank 7, and the plasma serum quantitative tank 7 is located on the side of the circumferential segment channel close to the rotation center 102;

[0095] The remaining blood cell storage groove 5 is connected to the middle part of the circumferential section channel, and the remaining blood cell storage groove 5 is located on the side of the circumferential section channel away from the rotation center 102.

[0096] A microfluidic valve 14 is provided on the waste liquid transmission channel 15 connected to the outlet end of the mixed culture tank 12. The microfluidic valve 14 is a normally closed valve, and the microfluidic valve 14 can have various shapes, such as a capillary valve, which is passively opened when a predetermined pressure is applied; or the microfluidic valve 14 is a valve that is actively operated by receiving electrical power or energy from the outside through an operation signal.

[0097] The plasma serum transmission channel 8 has a first inverted U-shaped section 80, and the waste liquid transmission channel 15 has a second inverted U-shaped section 150. Both the first inverted U-shaped section 80 and the second inverted U-shaped section 150 are slender, and provide the following functions: when the rotation speed of the chip body reaches a certain value, liquid can pass through them; that is, through the cooperation of siphon action and centrifugal action, liquid is allowed to pass only when the rotation speed of the chip body reaches a certain value.

[0098] In this embodiment, the chip body has at least 8 detection units 100, and at least 8 items can be detected simultaneously. The chip body is provided with a dissolving reagent storage tank 9 and two reaction reagent storage tanks, the dissolving reagent storage tank 9 stores the pre-packaged diluent (i.e., dissolving reagent), and the two reaction reagent storage tanks respectively store the pre-packaged first liquid reagent and the second liquid reagent.

[0099] A specific operation step of the chip of the present invention is provided below:

[0100] 1. A whole blood sample (venous blood, peripheral blood) is injected from the whole blood injection port 1 and stored in the whole blood storage tank. The microfluidic chip is controlled to rotate around its rotation center 102 at a first speed (first acceleration). The whole blood sample passes through the blood filter membrane 3, and at least 80% of the blood cells can be filtered out. The plasma / serum and the blood cells that are not filtered cleanly are transmitted to the remaining blood cell storage tank 5 and the plasma serum quantitative tank 7 through the whole blood transmission channel. The blood cells that are not filtered cleanly are stored in the remaining blood cell storage tank 5.

[0101] 2. The microfluidic chip is controlled to rotate at a second rotation speed (second acceleration), and the excess plasma / serum is transferred to the plasma / serum waste liquid tank 6.

[0102] 3. The microfluidic chip is controlled to rotate at a third rotation speed (third acceleration), and a certain amount of plasma / serum is transferred to the mixed culture tank 12 through the plasma-serum transfer channel.

[0103] The diluent (i.e., the dissolving reagent), the first liquid reagent, and the second liquid reagent are pre-packaged in the dissolving reagent storage tank 9, the first reaction reagent storage tank 10, and the second reaction reagent storage tank 11, respectively. The liquid reagent pre-packages can be punctured in sequence by an external spike device, and the liquid reagents can be released in sequence in conjunction with the centrifugal speed (release order: diluent, first liquid reagent, second liquid reagent); solid reagents are pre-packaged in the solid reagent storage tank, including immune microspheres (antibodies are linked to the surface of the microspheres, and the materials can be metal, plastic, silica, etc.), freeze-dried reagent beads, etc., and the reaction can be combined with the oscillation conditions (amplitude, frequency) to control the microfluidic chip for oscillation mixing, and a variety of solid reagent beads can be dissolved in sequence and incubated / reacted with the sample. It should be understood that in addition to N kinds of soluble solid reagents (i.e., freeze-dried reagent beads), the solid reagent storage tank can also include immune microspheres, where the reagents that need to be dissolved are a variety of freeze-dried reagent beads, and the same is true below.

[0104] 4. Manipulate the external spike device to pass through the puncture hole, puncture the pre-sealed bag of the diluent, release the diluent, and manipulate the microfluidic chip to rotate at the fourth speed (fourth acceleration), so that the diluent can be transferred to the mixed cultivation tank 12. At this time, the liquid in the mixed cultivation tank 12 can be retained in the mixed cultivation tank 12 due to the blocking effect of the microfluidic valve 14. As in Example 1 or Example 2, N kinds of freeze-dried reagent beads are dissolved and reacted in sequence. According to Example 1, a series reaction is performed; according to Example 2, a partial parallel reaction is performed; if other reagents need to be added during the reaction, the pre-sealed bag containing the corresponding reagents can be punctured by manipulating the external spike device. Of course, during the reaction process, a variety of oscillation conditions (including set amplitude, frequency, operation time, etc.) can also be configured to manipulate the microfluidic chip for oscillation mixing to assist the reaction, so that the solid reagent reacts with the sample in sequence. The reagents are added sequentially, which greatly simplifies the operation. In addition, according to the scheme of Example 2, partial parallel reactions can also be achieved, thereby shortening the reaction time and improving efficiency.

[0105] 5. The microfluidic chip is controlled to rotate at the fifth speed (fifth acceleration) to break through the microfluidic valve 14 after mixing. Since the liquid pressure is greater than the microfluidic valve 14, the liquid will be transferred from the mixed culture tank 12 (12) to the waste liquid tank 16 through the liquid transfer channel. The N kinds of freeze-dried reagent beads are dissolved and reacted in turn; the immune microspheres are used as loads (the material of the immune microspheres can be metal, plastic silica, etc., and the immune microspheres do not react during the reaction, but only serve as loads, and the solid reagents on them, such as antibodies, participate in the reaction) to bind antibodies, etc. For non-magnetic immune microspheres, their size is larger than the microfluidic valve and can be retained in the mixed culture tank. For magnetic immune microspheres, they can be controlled by an external magnetic field to avoid losing the magnetic immune microspheres during the cleaning stage (in the cleaning stage, a magnetic field is added to the outside of the chip to absorb the magnetic immune microspheres and keep them in the mixed culture tank. The magnetic field is removed during the mixing stage, and the magnetic immune microspheres are mixed and reacted with the system in the mixed culture tank).

[0106] 6. Operate the external spike device to pierce the pre-sealed bag of the first liquid reagent, and control the microfluidic chip to rotate at the sixth speed (sixth acceleration), so that the first liquid reagent can be transferred to the mixing and incubation tank 12. At this time, the liquid in the mixing and incubation tank 12 can be retained in the mixing and incubation tank 12 due to the blocking effect of the microfluidic valve 14. During the reaction process, certain oscillation conditions can also be provided to control the microfluidic chip to oscillate and mix. After the reaction is completed, control the microfluidic chip to rotate at the seventh speed (seventh acceleration) to break through the microfluidic valve 14 after mixing. Since the liquid pressure is greater than the microfluidic valve 14, the liquid will be transferred from the mixing and incubation tank 12 to the waste liquid tank 16 through the liquid transfer channel. Since the size of the immune microspheres is larger than the microfluidic valve 14, they can be retained in the mixing and incubation tank 12. For magnetic immune microspheres, they can be controlled by an external magnetic field to avoid losing the magnetic immune microspheres during the cleaning stage.

[0107] 7. Operate the external spike device to pierce the pre-sealed bag of the second liquid reagent, and operate the microfluidic chip to rotate at the eighth speed (eighth acceleration) to transfer the second liquid reagent to the mixing and incubation tank 12. At this time, the liquid in the mixing and incubation tank 12 can be retained in the mixing and incubation tank 12 due to the blocking effect of the microfluidic valve 14. During the reaction process, certain oscillation conditions can be provided to operate the microfluidic chip to oscillate and mix. After the reaction is completed, it can be detected through an external detection system (such as optical, fluorescent, chemiluminescent equipment, etc.).

[0108] Example 4

[0109] Provided is an application example of the chip of the present invention for chemiluminescent detection of hepatitis A virus antibodies.

[0110] The steps are:

[0111] 1. 50 microliters of venous whole blood (male sample, hematocrit ratio of 69%, i.e., 31% plasma and 69% blood cells) with sodium citrate anticoagulant is injected into the whole blood storage tank through the whole blood injection hole 1, and the disc chip is rotated at 3,000RPM (acceleration of 10,000RPM / s) for 60 seconds to make the whole blood sample pass through the blood filter membrane 3, more than 80% of the blood cells can be filtered out, and the plasma and blood cells that are not filtered cleanly are transmitted to the remaining blood cell storage tank 5 and the plasma serum quantitative tank 7 (quantified to 10 microliters) through the whole blood transmission channel, and the blood cells that are not filtered cleanly are stored in the remaining blood cell storage tank 5. The disc chip is rotated at 1,200RPM (acceleration of 3,400RPM / s) for 10 seconds, and the excess plasma is transmitted to the plasma serum waste liquid tank 6. The disk chip was rotated at 600 RPM (acceleration of 1,250 RPM / s) for 5 seconds, and 10 microliters of plasma was transferred to the mixing culture tank 12 through the plasma / serum transfer channel.

[0112] The diluent (dissolving reagent), the cleaning solution (the first liquid reagent), and the substrate (the second liquid reagent) are pre-packaged in the dissolving reagent storage tank 9, the first liquid reagent storage tank, and the second liquid reagent storage tank, respectively. The magnetic immune microspheres (superparamagnetic microparticles coated with streptavidin), the HAV antigen / lyophilized reagent beads, and the anti-HAVAg antibody / lyophilized reagent beads are pre-packaged in the solid reagent storage tank.

[0113] 2. Operate the external spike device to puncture the pre-packaged diluent, and operate the disc chip to rotate for 5 seconds at 2,500RPM (acceleration of 3,000RPM / s), which can transfer the diluent to the mixing culture tank 12. At this time, the liquid in the mixing culture tank 12 can be retained in the mixing culture tank 12 due to the blocking effect of the microfluidic valve 14. Operate the microfluidic chip to oscillate and mix 15 times with an amplitude of 370 degrees and a frequency of 10Hz, which can dissolve the HAV antigen / lyophilized reagent beads and react with the sample for 2 minutes; operate the microfluidic chip to oscillate and mix 15 times with an amplitude of 250 degrees and a frequency of 8Hz, and react the magnetic immune microspheres with the pre-liquid for 3 minutes; operate the microfluidic chip to oscillate and mix 20 times with an amplitude of 135 degrees and a frequency of 5Hz, which can dissolve the anti-HAVAg antibody / lyophilized reagent beads and react with the pre-liquid for 3 minutes. The disk chip is rotated at 3,500 RPM (acceleration of 5,250 RPM / s) for 5 seconds. Since the liquid pressure of the mixed liquid is greater than the microfluidic valve 14, it will break through the microfluidic valve 14 and be transferred to the waste liquid tank 16 through the liquid transfer channel. The magnetic immunomicrospheres can be controlled by an external magnetic field to avoid losing the magnetic immunomicrospheres during the washing step.

[0114] 3. Operate the external spike device to puncture the pre-packaged cleaning liquid, and rotate the disc chip at 2,500RPM (acceleration of 3,000RPM / s) for 5 seconds to transfer the cleaning liquid to the mixing culture tank 12. At this time, the liquid in the mixing culture tank 12 can be retained in the mixing culture tank 12 due to the blocking effect of the microfluidic valve 14. Operate the microfluidic chip with an amplitude of 180 degrees and a frequency of 15Hz to oscillate and mix 20 times. After the cleaning is completed, operate the disc chip to rotate at 3,500RPM (acceleration of 5,250RPM / s) for 5 seconds. The waste liquid after cleaning will break through the microfluidic valve 14 and be transferred to the waste liquid tank 16 through the liquid transfer channel because the liquid pressure is greater than the microfluidic valve 14. The magnetic immunomicrospheres can be controlled by an external magnetic field to avoid losing the magnetic immunomicrospheres during the cleaning step.

[0115] 4. Operate the external spike device to pierce the substrate pre-package, and operate the disc chip to rotate for 5 seconds at 2,500RPM (acceleration of 3,000RPM / s), and the substrate can be transferred to the mixing culture tank 12. At this time, the liquid in the mixing culture tank 12 can be retained in the mixing culture tank 12 due to the blocking effect of the microfluidic valve 14. Operate the microfluidic chip to oscillate and mix 20 times with an amplitude of 180 degrees and a frequency of 15Hz. After 300 seconds of reaction, it can be detected. The luminescence value is 7,318RLU. Substituting it into the reaction curve, the concentration value is 31.2IU / L, which is not much different from the value of 29.8IU / L ​​measured by the Roche Elecsys2010 immunoassay analyzer. Both are judged to be hepatitis A virus infection or past infection or the presence of antibodies after injection of hepatitis A vaccine.

[0116] Example 5 Analysis of filtration membrane separation efficiency

[0117] The whole blood sample is injected into the whole blood storage tank from the whole blood injection hole 1, and passes through the blood filter membrane 3 to accelerate the whole blood separation efficiency. Experimental parameters: rotation speed (5000RPM, 3000RPM, 1000RPM), sample blood volume ratio (40%, 60%, 80%); fixed parameters: whole blood sample volume is 50 microliters, acceleration is fixed at 10,000RPM / s, and rotation time is fixed at 60 seconds; experimental analysis: using a high-speed camera and image analysis software, the separation efficiency is analyzed by pictures, and the separation efficiency is defined as 80% as separation completion.

[0118] (Experiment 1), inject 50 microliters of whole blood sample (hematocrit 80%; plasma: 10 microliters, blood cells: 40 microliters) into the whole blood storage tank, and control the disk chip to rotate for 60 seconds at 5,000RPM, 3,000RPM, and 1,000RPM (acceleration fixed at 10,000RPM / s), respectively, and take screenshots and analyze them with a high-speed camera. Experimental data: When the disk chip is rotated at 5,000RPM for 60 seconds, 6.5 microliters of plasma can be separated, and the separation efficiency is 65%, which is considered to be incomplete; when the disk chip is rotated at 3,000RPM for 60 seconds, 8.7 microliters of plasma can be separated, and the separation efficiency is 87%, which is considered to be complete; when the disk chip is rotated at 1,000RPM for 60 seconds, 7.4 microliters of plasma can be separated, and the separation efficiency is 74%, which is considered to be incomplete. Experimental results: If the speed of the disc chip is too fast, the whole blood will pass through the filter membrane too fast, which will affect the separation efficiency; if the speed of the disc chip is too slow, the separation efficiency of the filter membrane alone will be too poor, and the separation efficiency cannot be improved. Experiments have shown that the separation efficiency is best when the disc chip is rotated at 3,000RPM (10,000RPM / s).

[0119] (Experiment 2), inject 50 microliters of whole blood sample into the whole blood storage tank, the blood volume ratio is 80%, 60%, and 40% respectively, and rotate the disc chip at 3,000RPM (acceleration 10,000RPM / s) for 60 seconds, and take screenshots and analyze them by high-speed camera. Experimental data: The blood solubility ratio of the sample is 80%. After rotation and filter membrane action, 8.7 microliters of plasma can be separated, and the separation efficiency is 87%, which is considered to be complete; the blood solubility ratio of the sample is 60%, and after rotation and filter membrane action, 18.2 microliters of plasma can be separated, and the separation efficiency is 91%, which is considered to be complete; the blood solubility ratio of the sample is 40%, and after rotation and filter membrane action, 27.8 microliters of plasma can be separated, and the separation efficiency is 93%, which is considered to be complete. Experiments have shown that whole blood samples with a blood volume ratio of up to 80% can be processed by rotating the disc chip at 3,000RPM (10,000RPM / s) for 60 seconds.

[0120] Example 6 Mixing Efficiency Analysis

[0121] The microfluidic chip was manipulated to oscillate back and forth to achieve the function of rapid mixing. Experimental parameters: amplitude (300 degrees, 200 degrees, 100 degrees), frequency (15Hz, 12Hz, 8Hz), mixing tank depth (800μm, 600μm, 400μm); fixed parameters: the working liquid volume was 50 microliters in total, including 25 microliters of colorless deionized water (viscosity 1cP) and 25 microliters of colored glycerol (viscosity 5cP), and the number of oscillations was fixed at 5 times; experimental analysis: using a high-speed camera and image analysis software, the mixing efficiency was analyzed by pictures (0% means no mixing, 100% means complete mixing).

[0122] (Experiment 1), fixed frequency 15Hz, mixing tank depth 800μm, oscillation times 5 times, control the microfluidic chip with amplitude 300 degrees, 200 degrees, and 100 degrees for oscillation mixing, and take screenshots and analyze them with a high-speed camera. Experimental data: The microfluidic chip is controlled with an amplitude of 300 degrees for oscillation mixing, and the mixing efficiency is 98%; the microfluidic chip is controlled with an amplitude of 200 degrees for oscillation mixing, and the mixing efficiency is 45%; the microfluidic chip is controlled with an amplitude of 100 degrees for oscillation mixing, and the mixing efficiency is 12%. Experimental results: The larger the amplitude, the better the mixing efficiency. In addition, the amplitude and mixing efficiency show a quadratic proportional relationship.

[0123] (Experiment 2), fixed amplitude of 300 degrees, mixing tank depth of 800μm, oscillation times 5 times, control the microfluidic chip for oscillation mixing at frequencies of 15Hz, 12Hz, and 8Hz respectively, and take screenshots and analyze them with a high-speed camera. Experimental data: The mixing efficiency of the microfluidic chip for oscillation mixing at a frequency of 15Hz is 98%; the mixing efficiency of the microfluidic chip for oscillation mixing at a frequency of 12Hz is 51%; the mixing efficiency of the microfluidic chip for oscillation mixing at a frequency of 8Hz is 15%. Experimental results: The greater the frequency, the better the mixing efficiency. In addition, the frequency and mixing efficiency are in a cubic proportional relationship.

[0124] (Experiment 3), fixed amplitude of 300 degrees, fixed frequency of 15Hz, oscillation times 5 times, oscillation mixing test was performed at mixing tank depths of 800μm, 600μm, and 400μm, respectively, and screenshots were taken and analyzed by high-speed camera. Experimental data: When the mixing tank depth is 800μm, the mixing efficiency is 98%; when the mixing tank depth is 600μm, the mixing efficiency is 42%; when the mixing tank depth is 400μm, the mixing efficiency is 13%. Experimental results: The deeper the mixing tank depth, the better the mixing efficiency. In addition, the mixing tank depth and mixing efficiency show a cubic proportional relationship.

[0125] Although the embodiments of the present invention have been disclosed as above, they are not limited to the applications listed in the specification and the implementation modes. They can be fully applied to various fields suitable for the present invention. For those familiar with the art, additional modifications can be easily implemented. Therefore, without departing from the general concept defined by the claims and the scope of equivalents, the present invention is not limited to specific details.

Claims

1. A one-time injection high-efficiency microfluidic detection chip, the detection chip has a rotation center, and the detection chip includes at least one detection unit for independent detection, the detection unit includes a mixed culture tank for providing a reaction space, characterized in that: The mixed culture tank is provided with a solid reagent storage tank storing N different solid reagents; The solid reagent includes a first solid reagent, a second solid reagent to an Nth solid reagent, all of which are coated with a protective film on the outside. The first solid reagent is coated with a first protective film on the outside, and the second solid reagent is coated with a first protective film and a second protective film in sequence from the outside to the inside. By analogy, the Nth solid reagent is coated with a first protective film, a second protective film to an Nth protective film in sequence from the outside to the inside. The first to Nth protective films can all be dissolved in the mixed culture tank under the action of the dissolving reagent provided by the detection unit, so that the protective films coated on the outside of the first to Nth solid reagents have different dissolution times; The dissolution time of the innermost protective film of the Nth solid reagent is exactly the same as the reaction time of the N-1th solid reagent body; or, the sum of the dissolution time of the innermost protective film of the Nth solid reagent and the dissolution time of the Nth solid reagent body is the same as the sum of the dissolution time of the N-1th solid reagent body and the reaction time of the N-1th solid reagent body; The high-efficiency microfluidic detection chip also includes at least one dissolving reagent storage tank and at least one reaction reagent storage tank connected to the mixed incubation tank; The first to Nth protective films are made of the same material that can be dissolved by the dissolving reagent stored in the dissolving reagent storage tank.

2. The high-efficiency microfluidic detection chip for one-time injection according to claim 1, characterized in that: The liquid reagents in the dissolving reagent storage tank and the reaction reagent storage tank are sealed and packaged in pre-sealed bags, and the pre-sealed bags can be partially ruptured under the action of external force to release the liquid reagents therein and provide them to the mixed cultivation tank.

3. The high-efficiency microfluidic detection chip for one-time injection according to claim 2, characterized in that: The body of the detection chip includes a bottom plate, an upper cover and a microchannel opened on the bottom plate, and the upper cover is sealed and buckled on the bottom plate, so that the microchannel between the bottom plate and the upper cover forms the detection unit; The upper cover above the liquid reagent storage tank is provided with a puncture hole, and the pre-sealed bag can be punctured through the puncture hole by an external puncturing device to release the liquid reagent therein.

4. The high-efficiency microfluidic detection chip for one-time injection according to any one of claims 1 to 3, characterized in that: The detection unit further comprises a whole blood storage tank and a whole blood injection hole connected to the whole blood storage tank for injecting a blood sample to be tested. A blood filter membrane is arranged in the whole blood storage tank.

5. The high-efficiency microfluidic detection chip for one-time injection according to claim 4, characterized in that: The detection unit also includes a plasma serum quantitative tank connected to the whole blood storage tank through a whole blood transmission channel, a plasma serum transmission channel connecting the plasma serum quantitative tank to the mixed incubation tank, a waste liquid tank connected to the mixed incubation tank through a waste liquid transmission channel, and a vent connected to the waste liquid tank.

6. The high-efficiency microfluidic detection chip for one-time injection according to claim 5, characterized in that: A position on the whole blood transmission channel upstream of the quantitative tank is connected to a residual blood cell storage tank; An overflow port is arranged on one side of the plasma serum quantitative tank close to the rotation center, and the overflow port is connected to a plasma serum waste liquid tank.

7. The high-efficiency microfluidic detection chip for one-time injection according to claim 6, characterized in that: The whole blood transmission channel includes a radial segment channel connected to the outlet end of the whole blood storage tank and a circumferential segment channel connected to the end of the radial segment channel, the end of the circumferential segment channel is connected to the plasma serum quantitative tank, and the plasma serum quantitative tank is located on a side of the circumferential segment channel close to the rotation center; The remaining blood cell storage groove is communicated with the middle part of the circumferential section channel, and the remaining blood cell storage groove is located on a side of the circumferential section channel away from the rotation center.

8. The high-efficiency microfluidic detection chip for one-time injection according to claim 5, characterized in that: A microfluidic valve is provided on the waste liquid transmission channel connected to the outlet end of the mixed cultivation tank.

9. The high-efficiency microfluidic detection chip for one-time injection according to claim 5, characterized in that: The plasma serum transmission channel has a first inverted U-shaped section, and the waste liquid transmission channel has a second inverted U-shaped section.

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