CN117282482B

CN117282482B - Method for preparing test tube and method for using test tube

Info

Publication number: CN117282482B
Application number: CN202311575408.7A
Authority: CN
Inventors: 李楠; 周天平; 王军波; 陈德勇
Original assignee: Aerospace Information Research Institute of CAS
Current assignee: Aerospace Information Research Institute of CAS
Priority date: 2023-11-23
Filing date: 2023-11-23
Publication date: 2024-02-13
Anticipated expiration: 2043-11-23
Also published as: CN117282482A

Abstract

The application discloses a preparation method of a test tube and a use method of the test tube, wherein the preparation method of the test tube comprises the following steps: sequentially storing at least two reactants in a sealing way; wherein storing each reactant in a sealed manner comprises: adding the reaction reagent into a test tube; adding a barrier layer into the test tube, wherein the barrier layer seals the reaction reagent, and the barrier layer can allow the magnetic beads to pass through; the reaction reagent and the barrier layer are sequentially distributed along the axial direction of the test tube, any two reaction reagents are sequentially distributed along the axial direction of the test tube, and two adjacent reaction reagents are separated by the barrier layer. In the preparation method of the test tube, the adjacent reaction reagents are sealed through the barrier layer, so that the sealed storage of at least two reaction reagents is realized, and no cross contamination exists; the volume required for storing the reaction reagent is reduced; the adaptability to the volume of the reaction reagent is improved, the adaptability to the density of the reaction reagent is also improved, and the universality is improved; flexibility is also improved.

Description

Method for preparing test tube and method for using test tube

Technical Field

The application relates to the technical field of in-vitro diagnosis, in particular to a preparation method of a test tube and a use method of the test tube.

Background

In the field of in vitro diagnosis, nucleic acid detection provides accurate judgment basis for disease prediction, diagnosis, prevention and treatment. Nucleic acid extraction equipment and nucleic acid detection equipment based on magnetic bead extraction and purification often need a plurality of reaction reagents, and correspondingly, a plurality of cavities for storing the reaction reagents need to be arranged, so that the whole structure is complex, and the volume of the whole equipment is large; furthermore, after the nucleic acid extraction apparatus and the nucleic acid detecting apparatus are provided with the cavities, the nucleic acid extraction apparatus and the nucleic acid detecting apparatus can be adapted to a specific volume of the reaction reagent only, resulting in poor adaptability to the volume of the reaction reagent.

In addition, the interface stability between different reagents is poor, and the reaction result is affected.

In addition, nucleic acid extraction equipment and nucleic acid detection equipment based on magnetic bead extraction and purification often need complicated liquid operation parts, so that the equipment is large in size, and the risk of cross contamination exists, so that the requirements of the area behind the medical infrastructure and the on-site immediate detection are hardly met.

In view of the above, how to store multiple reagents to reduce the volume required for storing the reagents and to improve the adaptability to the volume of the reagents is a problem to be solved by those skilled in the art.

Disclosure of Invention

In view of this, it is an object of the present application to provide a method of manufacturing a test tube and a method of using a test tube to reduce the volume required for storing a reaction reagent and to improve the adaptability to the volume of a reaction reagent.

In order to achieve the above purpose, the present application provides the following technical solutions:

a method of manufacturing a test tube comprising: sequentially storing at least two reactants in a sealing way;

wherein hermetically storing each of the reagents comprises: adding the reaction reagent into the test tube; adding a barrier layer into the test tube, wherein the barrier layer seals the reaction reagent, and the barrier layer can pass through magnetic beads;

the reaction reagents and the blocking layers are sequentially distributed along the axial direction of the test tube, any two reaction reagents are sequentially distributed along the axial direction of the test tube, and two adjacent reaction reagents are separated by the blocking layers.

Optionally, the first end of the test tube is provided with an opening, and the second end of the test tube is closed;

the blocking layer comprises a blocking piece and a communicating liquid, at least one channel structure is arranged on the circumferential side wall of the blocking piece, and a magnetic bead channel for the magnetic beads to pass through is formed by the channel structure and the inner wall of the test tube; the communication liquid can pass through the magnetic beads;

Adding a barrier layer into the test tube, specifically comprising: placing the blocking piece into the test tube, wherein the blocking piece is positioned above the reaction reagent, a gap is formed between the blocking piece and the inner wall of the test tube, and the gap is annular; adding the communication liquid into the test tube, wherein the communication liquid fills a gap between the blocking piece and the inner wall of the test tube;

wherein, will the separation piece is put into the test tube specifically is: securing the barrier within the tube;

an installation table is fixed on the inner wall of the test tube; fix the separation piece in the test tube, specifically: securing the barrier to the mounting table; or the bottom wall of the test tube is fixed with a mounting column; fix the separation piece in the test tube, specifically: securing the barrier to the mounting post; the number of the blocking parts is at least two, the blocking parts are sequentially distributed along the axial direction of the test tube, and any two blocking parts are fixed on the same mounting column;

the circumferential side wall of the blocking piece is provided with at least one concave opening, and the concave opening forms the channel structure; alternatively, the circumferential side wall of the barrier is provided with at least one protrusion forming the channel structure.

Optionally, the barrier layer comprises only a communication liquid;

adding a barrier layer into the test tube, specifically: and adding the communication liquid into the test tube, wherein the communication liquid seals the reaction reagent, and the communication liquid can be used for the magnetic beads to pass through.

Optionally, the test tube comprises: test tube barrel and piston push rod;

the first end of the test tube barrel is provided with a first opening for inserting the first end of the piston push rod, and the second end of the test tube barrel is provided with a second opening for allowing liquid to enter and exit;

the first end of the piston push rod is in sealing connection with the inner wall of the test tube barrel, the second end of the piston push rod is positioned outside the test tube barrel, and the piston push rod can reciprocate along the axial direction of the test tube barrel so as to realize the addition and release of the reaction reagent and the communication liquid.

Optionally, in storing at least one of the reagents in a sealed manner, before adding the reagent into the tube, the method further comprises: adding a freeze-drying reagent into the test tube, adding molten paraffin into the test tube, covering the freeze-drying reagent by the molten paraffin, and solidifying the molten paraffin after cooling;

the barrier layer comprises a communicating liquid, and the communicating liquid is paraffin;

Adding the communication liquid into the test tube, specifically: and adding molten paraffin into the test tube, wherein the paraffin solidifies to form the barrier layer or part of the barrier layer after cooling, and the molten paraffin can pass through the magnetic beads.

Optionally, at least two reagents are hermetically stored in sequence, specifically: sequentially storing two reaction reagents in a sealing way, wherein the two reaction reagents are a first reaction reagent and a second reaction reagent respectively;

sequentially storing a first reactant and the second reactant in a sealed manner, comprising:

adding the first reactant into the test tube;

adding a first blocking member into the test tube, wherein the first blocking member is positioned above the first reactant, a gap is formed between the first blocking member and the inner wall of the test tube, and the gap is annular; adding the communication liquid into the test tube, wherein the communication liquid fills a gap between the first barrier and the inner wall of the test tube;

adding the second reactant into the test tube;

adding a second blocking piece into the test tube, wherein the second blocking piece is positioned above the second reactant, a gap is formed between the second blocking piece and the inner wall of the test tube, and the gap is annular; adding the communication liquid into the test tube, wherein the communication liquid fills a gap between the second barrier and the inner wall of the test tube;

Wherein the first barrier and the second barrier are both the barriers.

sequentially storing a first reactant and the second reactant in a sealing way, and specifically comprising:

pushing the piston push rod to the bottom of the test tube barrel, then inserting the test tube barrel into a container filled with the first reactant, and pulling the piston push rod upwards to suck the first reactant into the test tube barrel;

inserting the tube into a container containing the communication liquid, and pulling the piston push rod upwards to suck the communication liquid into the tube, wherein the communication liquid seals the first reactant;

inserting the tube into a container containing the second reactant, and pulling the piston push rod upwards to suck the second reactant into the tube;

and inserting the test tube barrel into a container filled with the communication liquid, and pulling the piston push rod upwards to suck the communication liquid into the test tube barrel, wherein the communication liquid seals the second reactant.

The application also provides a use method of the test tube, wherein the test tube is prepared by the preparation method of the test tube;

the using method of the test tube comprises the following steps:

adding a third reaction reagent into the test tube, wherein the third reaction reagent comprises a sample to be detected and magnetic beads, and scattering the magnetic beads in the third reaction reagent so as to uniformly disperse the magnetic beads and fully combine with the sample to be detected to complete a first-step reaction;

moving the magnetic beads to the second reaction reagent through the magnetic bead channel, and scattering the magnetic beads in the second reaction reagent to complete a second-step reaction;

moving the magnetic beads into the first reaction reagent through the magnetic bead channel, and scattering the magnetic beads in the first reaction reagent to complete the third reaction;

and detecting the reaction result in the test tube.

Optionally, in the case that the test tube has at least two detection cavities, the magnetic bead channels are in one-to-one correspondence with the detection cavities;

before moving the magnetic beads into the second reaction reagent through the magnetic bead channel, further comprising: uniformly dividing the magnetic beads into at least two magnetic bead stacks;

before moving the magnetic beads into the first reaction reagent through the magnetic bead channel, further comprising: uniformly dividing the magnetic beads into at least two magnetic bead stacks;

Wherein the magnetic bead stacks and the magnetic bead channels are in one-to-one correspondence;

the magnetic beads are scattered through a magnetic bead scattering device outside the test tube, wherein the magnetic bead scattering device is a device capable of applying vibration force, an ultrasonic field or a magnetic field transformation outside the test tube;

moving the magnetic beads and uniformly dividing the magnetic beads into at least two magnetic bead stacks through a magnetic bead moving device outside the test tube, wherein the magnetic bead moving device can generate an equal-intensity magnetic field distributed in a space multipoint manner;

detecting a reaction result in the test tube by a detection device outside the test tube, wherein the detection device is an optical signal detection device, an electric signal detection device or a magnetic signal detection device;

the using method of the test tube further comprises a heating device, wherein the heating device is used for heating the first reactant, and in the case that paraffin is included in the test tube, the heating device is also used for heating the paraffin; the paraffin is the communication liquid and/or the paraffin-covered freeze-drying reagent.

The application also provides a use method of the test tube, wherein the test tube is prepared by the preparation method of the test tube, the first reaction reagent comprises a sample to be detected and magnetic beads, and the second reaction reagent is a magnetic bead cleaning solution;

The using method of the test tube comprises the following steps: inserting the test tube cartridge into a container containing a third reactant, the piston push rod being pulled upwards to draw the third reactant into the test tube cartridge, the third reactant being a nucleic acid eluent; scattering the magnetic beads in the first reaction reagent, and fully combining the magnetic beads with a sample to be detected to complete a first-step reaction; the magnetic beads are downwards moved into the second reaction reagent along the inner wall of the test tube barrel, and the magnetic beads in the second reaction reagent are scattered to complete the second-step reaction; the magnetic beads are downwards moved into the third reaction reagent along the inner wall of the test tube barrel, and the magnetic beads in the third reaction reagent are scattered to complete the third reaction; the magnetic beads are upwards moved to a communicating liquid at the upper side of the third reaction reagent along the inner wall of the test tube barrel; pressing down on the piston push rod to expel the third reactant from the test tube;

alternatively, the method for using the test tube comprises the following steps: inserting the test tube barrel into a container filled with the third reaction reagent, and pulling the piston push rod upwards to suck the third reaction reagent into the test tube barrel, wherein the third reaction reagent is nucleic acid amplification solution; inserting the tube into a container containing the communication liquid, and pulling the piston push rod upwards to suck the communication liquid into the tube, wherein the communication liquid seals the third reactant; scattering the magnetic beads in the first reaction reagent, and fully combining the magnetic beads with a sample to be detected to complete a first-step reaction; the magnetic beads are downwards moved into the second reaction reagent along the inner wall of the test tube barrel, and the magnetic beads in the second reaction reagent are scattered to complete the second-step reaction; the magnetic beads are downwards moved into the third reaction reagent along the inner wall of the test tube barrel, and the magnetic beads in the third reaction reagent are scattered to complete the third reaction; detecting the reaction result in the test tube.

In the preparation method of the test tube, the adjacent reaction reagents are sealed through the barrier layer, so that the reaction reagents of different types are stably separated through the barrier layer, namely, the sealed storage of at least two reaction reagents is realized, and no cross contamination is caused; because the reaction reagent and the barrier layer are sequentially distributed along the axial direction of the test tube, any two reaction reagents are sequentially distributed along the axial direction of the test tube, compared with the prior art, a plurality of cavities are not required to be arranged for storing the reaction reagents, and the volume required for storing the reaction reagents is effectively reduced; in addition, the volume of the reaction reagent can be adjusted by adjusting the height of the reaction reagent (the height of the reaction reagent in the axial direction of the test tube), so that the adaptability to the volume of the reaction reagent is effectively improved, the adaptability to the density of the reaction reagent is also improved, and the universality is improved; furthermore, any kind of reactant can be arranged in a building block mode, so that the flexibility is improved.

Meanwhile, the preparation method of the test tube can realize that reagents required by various reactions are stored in a single test tube, is simple to operate, is short in time, and is stable and controllable in interface, so that the method is expected to promote wide application of on-site instant detection.

According to the application method of the test tube, the magnetic beads in the test tube can be shuttled among various reaction reagents through the manipulation of the magnetic beads outside the test tube, the whole nucleic acid/immune analysis process based on the magnetic bead method can be realized under the airtight condition, no complex microfluid manipulation component is used, the equipment required by in-vitro diagnosis is greatly simplified, cross contamination is avoided, the size is small, the cost is low, and the popularization of the portable in-vitro diagnosis technology can be promoted.

Drawings

In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings that are required to be used in the embodiments or the description of the prior art will be briefly described below, and it is obvious that the drawings in the following description are only embodiments of the present application, and that other drawings may be obtained according to the provided drawings without inventive effort to a person skilled in the art.

FIG. 1 is a schematic flow chart of a method for preparing a test tube according to an embodiment of the present disclosure;

FIG. 2 is a partial enlarged view of the internal structure of a test tube in the method for manufacturing a test tube according to the first embodiment of the present application;

FIG. 3 is a schematic view showing an internal structure of a test tube in a method for manufacturing a test tube according to an embodiment of the present disclosure;

FIG. 4 is a schematic view of the barrier of FIG. 3;

FIG. 5 is a schematic view showing another internal structure of a test tube in the method for manufacturing a test tube according to the first embodiment of the present application;

FIG. 6 is a schematic view of the barrier of FIG. 5;

FIG. 7 is a schematic flow chart of a method for using a test tube according to an embodiment of the present disclosure;

FIG. 8 is a schematic flow chart of another method for preparing a test tube according to the first embodiment of the present disclosure;

FIG. 9 is a schematic flow chart of another method for using a test tube according to the first embodiment of the present disclosure;

fig. 10 is a schematic flow chart of a method for preparing a test tube according to a second embodiment of the present disclosure;

FIG. 11 is a schematic flow chart of a portion of a method for using a test tube according to a second embodiment of the present disclosure;

fig. 12 is a schematic structural diagram of a test tube in the method for using a test tube according to the third embodiment of the present application;

fig. 13 is a schematic flow chart of a method for preparing a test tube according to a fourth embodiment of the present application;

fig. 14 is a schematic flow chart of a method for using a test tube according to the fourth embodiment of the present application;

fig. 15 is a schematic flow chart of another method for using the test tube according to the fourth embodiment of the present application.

Reference numerals illustrate:

100-test tube; 101-a detection chamber; 102-test tube cap; 103-mounting table; 104-mounting posts; 105-a second projection; 106-testing tube; 107-piston push rod; 200-a first reactant; 300-barrier; 300 a-a first barrier; 300 b-a second barrier; 301-a recessed opening; 302-a first projection; 400-communicating liquid; 500-a second reactant; 600-third reactant; 700-magnetic beads; 800-a magnetic bead moving device; 801-a magnet; an 802-support ring; 900-heating device; 1000-detecting means; 1100-lyophilizing the reagent; 1200-paraffin; 1300-a magnetic bead scattering device; 01-magnetic bead channel.

Detailed Description

The following description of the embodiments of the present application will be made clearly and fully with reference to the accompanying drawings, in which it is evident that the embodiments described are only some, but not all, of the embodiments of the present application. All other embodiments, which can be made by one of ordinary skill in the art without undue burden from the present disclosure, are within the scope of the present disclosure.

The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application. The terminology used in the following embodiments is for the purpose of describing particular embodiments only and is not intended to be limiting of the application. As used in the specification and the appended claims, the singular forms "a," "an," "the," and "the" are intended to include, for example, "one or more" such forms of expression, unless the context clearly indicates to the contrary. It should also be understood that in embodiments of the present application, "one or more" means one, two, or more than two; "and/or", describes an association relationship of the association object, indicating that three relationships may exist; for example, a and/or B may represent: a alone, a and B together, and B alone, wherein A, B may be singular or plural. The character "/" generally indicates that the context-dependent object is an "or" relationship.

Reference in the specification to "one embodiment" or "some embodiments" or the like means that a particular feature, structure, or characteristic described in connection with the embodiment is included in one or more embodiments of the application. Thus, appearances of the phrases "in one embodiment," "in some embodiments," "in other embodiments," and the like in the specification are not necessarily all referring to the same embodiment, but mean "one or more but not all embodiments" unless expressly specified otherwise. The terms "comprising," "including," "having," and variations thereof mean "including but not limited to," unless expressly specified otherwise.

The plurality of the embodiments of the present application refers to greater than or equal to two. It should be noted that, in the description of the embodiments of the present application, the terms "first," "second," and the like are used for distinguishing between the descriptions and not necessarily for indicating or implying a relative importance, or alternatively, for indicating or implying a sequential order.

Example 1

The first embodiment provides a preparation method of a test tube and a use method of the test tube.

As shown in fig. 1, the preparation method of the test tube provided in the first embodiment includes: s11), S12), S13), S14), S15), and S16).

S11) the first reaction reagent 200 is added to the test tube 100.

S12) adding a first blocking member 300a to the test tube 100, the first blocking member 300a being located above the first reactive agent 200, a gap being provided between the first blocking member 300a and the inner wall of the test tube 100, the gap being annular.

S13) filling the gap between the first barrier 300a and the inner wall of the test tube 100 with the communication liquid 400.

S14) adding the second reaction reagent 500 to the test tube 100.

S15) adding a second blocking member 300b into the test tube 100, the second blocking member 300b being located above the second reactant 500, a gap being provided between the second blocking member 300b and the inner wall of the test tube 100, the gap being annular.

S16) filling the gap between the second barrier 300b and the inner wall of the test tube 100 with the communication liquid 400.

In the first embodiment, the first end (top end) of the test tube 100 has an opening, and the second end (bottom end) of the test tube 100 is closed. A first end of the test tube 100 may be provided with a tube cap 102, which cap 102 is adapted to close the opening of the test tube 100. It should be noted that, during the preparation of the test tube, the opening of the test tube 100 is opened.

In the first embodiment, the first reagent 200, the second reagent 500, and the communicating liquid 400 may be added to the test tube 100 by a pipette or the like.

In S11), the first reaction reagent 200 is a liquid reagent. In order to avoid contact between the first reactive agent 200 and the inner wall of the test tube 100 during the addition of the first reactive agent 200, the above-mentioned test tube 100 may be selected to be vertically placed, i.e., the first reactive agent 200 is added to the vertically placed test tube 100. Accordingly, in S14), the second reaction reagent 500 is a liquid reagent. In order to avoid contact between the second reagent 500 and the inner wall of the test tube 100 during the addition of the second reagent 500, the test tube 100 may be selected to be vertically placed, i.e., the second reagent 500 is added to the vertically placed test tube 100.

The first barrier 300a and the second barrier 300b may each be referred to as a barrier 300, and the first reactant 200 and the second reactant 500 may each be referred to as a reactant. The gap between the barrier 300 and the inner wall of the test tube 100 indicates that the barrier 300 does not completely cover the liquid surface of the reactant located thereunder, i.e., the outer contour of the barrier 300 is located at the inner periphery of the outer contour of the liquid surface of the reactant located thereunder.

The communicating liquid 400 is located at the periphery of the blocking member 300, the communicating liquid 400 can allow the magnetic beads to pass through, and the communicating liquid 400 is fixedly connected with the blocking member 300 and the inner wall of the test tube 100, so that the blocking member 300 is fixed in the test tube 100. It is understood that the communication 400 and the first reactant 200 are not compatible with each other, and that the communication 400 and the second reactant 500 are not compatible with each other.

As shown in fig. 2, the first reactive agent 200 and the second reactive agent 500 are stably separated into upper and lower layers by the first barrier 300a and the communicating liquid 400. In this way, by increasing the contact area of the communicating liquid 400 with the first barrier 300a and the contact area of the communicating liquid 400 with the inner wall of the test tube 100, stability can be improved.

As shown in fig. 3 to 6, the circumferential side wall of the barrier 300 is provided with at least one channel structure, which forms a magnetic bead channel 01 for passing magnetic beads with the inner wall of the cuvette 100. In this way, at least one magnetic bead channel 01 can be formed. It will be appreciated that the channel structure corresponds one-to-one to the bead channel 01.

In the circumferential side wall of the barrier 300, the gap between the position where the channel structure is not provided and the inner wall of the cuvette 100 is small, and the gap between the position where the channel structure is provided and the inner wall of the cuvette 100 is large. Thus, the larger gap is the magnetic bead channel 01.

As shown in fig. 3 and 4, three recess openings 301 may be optionally provided on the circumferential side wall of the barrier 300, and the recess openings 301 form a channel structure, in this case, three magnetic bead channels 01, and the cross section of the test tube 100 may be circular or other shapes. It will be appreciated that the recess 301 corresponds to the bead channel 01 one-to-one.

As shown in fig. 5 and 6, three first protruding portions 302 may be provided on the circumferential side wall of the barrier 300, and the first protruding portions 302 may form a channel structure, in this case, three magnetic bead channels 01 are provided, the circumferential side wall of the test tube 100 is provided with the second protruding portions 105, and the magnetic bead channels 01 are formed between the first protruding portions 302 and the second protruding portions 105. It will be appreciated that the first protrusions 302 are in one-to-one correspondence with the magnetic bead channels 01.

In the structures shown in fig. 3 and 5, the contact area between the communicating liquid 400 and the blocking member 300 and the contact area between the communicating liquid 400 and the inner wall of the test tube 100 are increased, and the interface stability between the communicating liquid 400 and the reactant is improved.

In practical situations, the magnetic bead channel 01 can also be one, two, four or more than five, and is selected according to practical situations.

Under the condition that the number of the magnetic bead channels 01 is more than two, the test tube 100 can be selected to be provided with at least two detection cavities 101, and the detection cavities 101 are in one-to-one correspondence with the magnetic bead channels 01, so that the magnetic beads can enter each detection cavity 101, and the normal reaction in the detection cavities 101 can be ensured. It will be appreciated that in the corresponding detection chamber 101 and bead channel 01, the detection chamber 101 is located directly below the bead channel 01.

In the case of at least two detection chambers 101, multi-index detection of the same sample to be detected is achieved.

In order to improve the interface stability, the blocking member 300 is further fixed in the test tube 100 by a fixing structure, and the fixing structure of the blocking member 300 can be selected according to practical situations.

As shown in fig. 3, the inner wall of the test tube 100 is fixed with a mounting table 103, the barrier 300 is placed on the mounting table 103, and the barrier 300 is fixed to the mounting table 103. The blocking member 300 and the mounting table 103 may be fixedly connected by bonding or clamping, which is not limited in this embodiment.

The number of the mounting bases 103 may be one or two or more. In the case where there are two or more mounting bases 103, any two of the mounting bases 103 are distributed in sequence along the circumferential direction of the test tube 100.

In the structure shown in fig. 3, the positions of the bead channel 01 and the mount 103 in the circumferential direction of the cuvette 100 are different. In this way, the mount 103 is prevented from interfering with the bead channel 01.

When the number of the blocking members 300 is two or more, there are at least two mounting stages 103 in the axial direction of the test tube 100; moreover, the number of mounting stages 103 and the number of stoppers 300 are the same in the axial direction of the test tube 100.

As shown in fig. 5, the bottom wall of the test tube 100 is fixed with a mounting post 104, and the barrier 300 is fixed to the mounting post 104. Wherein, the blocking member 300 and the mounting post 104 can be fixedly connected in a clamping, inserting or threaded connection manner.

When there are more than two barriers 300, all of the barriers 300 are secured to the same mounting post 104.

To avoid the mounting post 104 affecting the bead channel, the mounting post 104 and cuvette 100 may optionally be coaxially arranged.

The communicating liquid 400 may be an oily liquid insoluble in water, such as silicone oil, mineral oil, silicone gel or paraffin, and the density of the communicating liquid 400 is close to that of the reactant, the viscosity of the communicating liquid 400 is relatively high, and the gap between the blocking member 300 and the inner wall of the test tube 100 can be stably sealed by means of the viscous force between the communicating liquid 400 and the inner wall of the test tube 100 and the viscous force between the communicating liquid 400 and the blocking member 300. In some embodiments, the viscosity of the communication 400 may be selected to be greater than ten times the viscosity of water.

Illustratively, the densities and fluid viscosities (viscosities) of water, silicone oil, mineral oil, silicone gel, paraffin are shown in table one.

List one

Solution	Density (g/mL)	Fluid viscosity (cps)
			Water and its preparation method	1	1
Silicone oil	0.963	10-100000
			Mineral oil	0.877	10.37
Silica gel	0.97	70000-160000
			Paraffin wax	0.9	110-230

In the case where the communicating liquid 400 is paraffin, the paraffin is heated and melted, and when the communicating liquid 400 is added, the melted paraffin is added, and after the addition is completed, the melted paraffin is cooled and solidified, and the solidified paraffin seals the gap between the barrier 300 and the inner wall of the test tube 100. Therefore, in the case where the communication liquid 400 is paraffin, the outside of the test tube 100 is further provided with a heating means for heating the paraffin to melt the paraffin, and the melted paraffin can pass through the magnetic beads. It will be appreciated that the location of the heating means corresponds to the location of the wax.

In the first embodiment, the blocking member 300 improves the interface stability between the communicating liquid 400 and the reactant by mainly reducing the contact area between the communicating liquid 400 and the reactant, increasing the contact area between the communicating liquid 400 and the blocking member 300, and increasing the contact area between the communicating liquid 400 and the inner wall of the test tube 100.

In the first embodiment, the communication liquid 400 has three main functions: firstly, the characteristic that the oil and water are not contained is utilized to isolate different reaction reagents on the upper layer and the lower layer; secondly, by utilizing the characteristic that the magnetic beads can flow at normal temperature or by heating, a flowable medium for the magnetic beads to pass through is provided; thirdly, the blocking member 300 is fixed to the test tube 100 by using the characteristic of having a certain viscosity.

The sealed storage of the first reactant 200 is completed through the above-described S11), S12), and S13), it can be understood that the above-described S11), S12), and S13) are steps of sealed storage of the first reactant 200; the sealed storage of the second reactant 500 is completed through the above-described S14), S15), and S16), and it is understood that the above-described S14), S15), and S16) are steps of storing the second reactant 500 in a sealed manner. Thus, the above-described test tube manufacturing method completes the sealed storage of the two reagents of the first reagent 200 and the second reagent 500.

In actual practice, the above-mentioned test tube manufacturing method may further hermetically store at least three reagents, and referring to the step of hermetically storing the first reagent 200 and the step of hermetically storing the second reagent 500, for example, sealing the third reagent, the fourth reagent, the fifth reagent, and the like may be completed. Therefore, the preparation method of the test tube provided in the first embodiment can form a multilayer stable liquid interface, so that the isolation, sealing and storage of any number of reaction reagents in the test tube 100 can be realized, and further the long-term stable storage of a plurality of reaction reagents in the test tube 100 and the operation of subsequent reactions can be realized.

The communication liquid 400 is insoluble in any of the reagents.

In the first embodiment, by using the blocking member 300 and the method of filling the communicating liquid 400 in the gap between the blocking member 300 and the inner wall of the test tube 100, a stable interface formed between the reactant and the communicating liquid 400 can be realized without being limited by the shape and the volume of the test tube 100, so that any multilayer stable liquid interface can be formed in the gravity direction without being limited by the density of each layer of liquid and the volume of the liquid, and stable sealed storage of the multilayer reactant and no cross contamination are ensured; and can realize that the reagent that multiple reactions need is stored to single test tube 100, easy operation, the time spent is short, and the interface is stable controllable to hopefully promote the wide application of on-the-spot instant detection.

In the foregoing, both the barrier 300 and the communication liquid 400 may be understood as being part of the barrier layer. The first reactant 200, the second reactant 500, and the communication liquid 400 may all be referred to as a liquid.

In the first embodiment, the reaction reagents and the barrier layers (the barrier 300 and the communicating liquid 400) are sequentially distributed along the axial direction of the test tube 100, any two reaction reagents are sequentially distributed along the axial direction of the test tube 100, and the two adjacent reaction reagents are separated by the barrier layer.

In the preparation method of the test tube 100, the adjacent reaction reagents are sealed through the barrier layer, so that the stable separation of different reaction reagents through the barrier layer is realized, namely, the sealed storage of at least two reaction reagents is realized, and no cross contamination is caused; because the reaction reagents and the barrier layers are sequentially distributed along the axial direction of the test tube 100, any two reaction reagents are sequentially distributed along the axial direction of the test tube 100, compared with the prior art, a plurality of cavities are not required to be arranged for storing the reaction reagents, and the volume required for storing the reaction reagents is effectively reduced; in addition, the volume of the reaction reagent can be adjusted by adjusting the height of the reaction reagent (the height of the reaction reagent in the axial direction of the test tube), so that the adaptability to the volume of the reaction reagent is effectively improved, the adaptability to the density of the reaction reagent is also improved, and the universality is improved; furthermore, any kind of reactant can be arranged in a building block mode, so that the flexibility is improved.

In other embodiments, if the communicating liquid 400 and the inner wall of the test tube 100 can have a larger specific surface area, a larger viscous force is provided between the communicating liquid 400 and the inner wall of the test tube 100, i.e. the communicating liquid 400 and the reactant can be stably stored in multiple layers without using the barrier 300. In this case, the communication liquid 400 may be understood as a barrier layer, i.e. the barrier layer comprises only the communication liquid 400.

In the preparation method of the test tube, a barrier layer is added into the test tube 100, specifically: the communicating liquid 400 is added into the test tube 100, the communicating liquid 400 seals the reaction reagent, and the communicating liquid 400 can pass through the magnetic beads.

In other embodiments, the freeze-drying reagent may be added to the test tube 100, then the melted paraffin is added to the test tube 100, the melted paraffin covers the freeze-drying reagent, the paraffin solidifies to form a dense paraffin protection layer (paraffin) after cooling, and then the reaction reagent is added to the test tube 100, so that the storage stability and the use flexibility of the reagent can be further improved.

The lyophilized reagent and the paraffin wax covering the lyophilized reagent may be provided in any one of the reaction reagents, and this is not limited in this embodiment.

In the above embodiment, the outside of the test tube 100 is provided with a heating device whose position corresponds to that of the paraffin, and the heating device is used to heat the paraffin to melt the paraffin, thereby achieving release of the lyophilized reagent.

The first embodiment of the invention also provides a using method of the test tube, wherein the test tube is prepared by adopting the preparation method of the test tube. As shown in fig. 7, the method for using the test tube includes: s21), S22), S23), and S24).

S21) adding the third reactant 600 into the test tube 100, the third reactant 600 comprising the magnetic beads 700 and the sample to be detected, scattering the magnetic beads 700 in the third reactant 600 so that the magnetic beads 700 are uniformly dispersed and sufficiently combined with the sample to be detected to complete the first-step reaction.

S22) moving the magnetic beads 700 into the second reaction reagent 500 through the magnetic bead channel 01, and scattering the magnetic beads 700 in the second reaction reagent 500 to complete the second reaction.

S23) moving the magnetic beads 700 into the first reaction reagent 200 through the magnetic bead channel 01, scattering the magnetic beads 700 in the first reaction reagent 200, to complete the third reaction.

S24) moving the magnetic beads 700 into the second reaction reagent 500 through the magnetic bead channel 01, and detecting the reaction result in the test tube 100.

In S21) above, the third reaction reagent 600 is a liquid. The addition of the third reagent 600 into the tube 100 can be understood as: the magnetic beads 700 and the sample to be detected are added into the test tube 100, specifically: adding a sample to be detected mixed with the magnetic beads 700 into the test tube 100; alternatively, the magnetic beads 700 and the sample to be detected are sequentially added into the test tube 100; or the sample to be detected and the magnetic beads 700 are sequentially added to the test tube 100.

In addition, in S21) above, the magnetic beads 700 may be stored in the test tube 100 in advance, and S21) only the sample to be detected is added. In this case, the third reagent 600 includes only the sample to be detected.

In S21), the magnetic beads 700 in the third reaction agent 600 are scattered, specifically: the beads 700 in the sample to be detected are scattered by a bead scattering device (not shown) outside the test tube 100.

The above-described magnetic bead scattering device is used to apply a vibration force, an ultrasonic field, or a transformed magnetic field to the outside of the test tube 100 to scatter the magnetic beads 700.

In S22), the magnetic beads 700 are moved to the second reaction reagent 500 through the magnetic bead channel, specifically: the magnetic beads 700 are moved into the second reaction reagent 500 through the magnetic bead channel by the magnetic bead moving means 800 outside the cuvette 100.

The magnetic bead moving device 800 may be a permanent magnet or an electromagnet. Of course, the magnetic bead moving device 800 may be another type of device, which is not limited in this embodiment.

In the case of at least two magnetic bead channels 01, the magnetic bead moving device 800 is further configured to uniformly divide the magnetic beads 700 into at least two magnetic bead stacks, where the magnetic bead stacks are in one-to-one correspondence with the magnetic bead channels 01, so that the magnetic beads can uniformly enter each magnetic bead channel 01. Illustratively, the magnetic bead moving device 800 is capable of generating a spatially multi-point distributed, uniform magnetic field to uniformly split the magnetic beads 700. The constant magnetic field means: the magnetic fields generated by the magnetic bead moving device 800 at each magnetic bead channel 01 have the same magnitude and the same direction; in the case where the number of detection chambers 101 is two or more, the magnetic fields generated by the magnetic bead moving device 800 at each detection chamber 101 are equal in magnitude and the directions of the magnetic fields are the same.

Based on the above, the above S21) and S22) further include: s21') uniformly dividing the magnetic beads 700 into at least two magnetic bead stacks, wherein the magnetic bead stacks are in one-to-one correspondence with the magnetic bead channels 01; the above steps S22) and S23) further include: s22') uniformly divides the magnetic beads 700 into at least two magnetic bead stacks, wherein the magnetic bead stacks are in one-to-one correspondence with the magnetic bead channels 01.

In the above embodiment, the magnetic beads 700 are uniformly stacked, which is particularly suitable for the case where the test tube 100 has more than two detection chambers 101.

In S22) above, in order to facilitate the magnetic beads 700 entering the magnetic bead channel 01, the magnetic beads 700 may be driven by the magnetic bead moving device 800 outside the cuvette 100 to be concentrated on the inner wall of the cuvette 100, and then the magnetic beads 700 may be manipulated into the magnetic bead channel 01 and the second reaction reagent 500 by moving the magnetic bead moving device 800. Accordingly, in S23) above, in order to facilitate the entry of the magnetic beads 700 into the magnetic bead channel 01, the magnetic beads 700 may be driven to be enriched on the inner wall of the cuvette 100 by the magnetic bead moving means 800 outside the cuvette 100, and then the magnetic beads 700 may be manipulated into the magnetic bead channel 01 as well as the first reaction reagent 200 by moving the magnetic bead moving means 800.

In the above S22) and S23), the magnetic beads 700 may be scattered by the aforementioned magnetic bead scattering device.

In S24) above, the reaction result in the test tube 100 can be detected by the detection device 1000 outside the test tube 100.

The position of the detection device 1000 corresponds to the position of the first reagent 200. The detection device 1000 may be an optical signal detection device, an electrical signal detection device, or a magnetic signal detection device.

In the case where the detection device 1000 is an optical signal detection device, the cuvette 100 may be selected to be a transparent tube so that the detection device 1000 detects the reaction result.

In the above embodiment, the heating device 900 may be disposed outside the test tube 100, and the position of the heating device 900 corresponds to the position of the first reaction reagent 200, and the heating device 900 is used to heat the first reaction reagent 200 to satisfy the temperature required for the third reaction. Based on this, S23) above specifically includes: the magnetic beads 700 are concentrated on the inner wall of the test tube 100 by driving the magnetic bead moving device 800 outside the test tube 100, the magnetic beads 700 are controlled to enter the first reaction reagent 200 through the magnetic bead channel 01 by moving the magnetic bead moving device 800, the magnetic beads 700 are scattered by the magnetic bead scattering device outside the test tube 100 so that the magnetic beads 700 are uniformly dispersed and fully mixed with the first reaction reagent 200, and the first reaction reagent 200 is heated by the heating device 900 outside the test tube 100, so that the third reaction is completed.

In the above embodiment, in the case where the communication liquid 400 is paraffin, the paraffin and the first reactant 200 may be heated by the same heating device 900.

The preparation method of the test tube and the use method of the test tube provided in the first embodiment can be used for nucleic acid detection.

Taking the nucleic acid detection application as an example, first a test tube 100 is prepared, and the test tube 100 may be referred to as a nucleic acid detection test tube 100. The first reaction reagent 200 is a nucleic acid amplification solution, the second reaction reagent 500 is a magnetic bead cleaning solution, and the communication solution 400 is silicone oil.

As shown in fig. 1 (1), the test tube 100 is vertically placed, and the first reaction reagent 200 is added to the test tube 100; as shown in fig. 1 (2), a first barrier 300a is placed in the cuvette 100; as shown in fig. 1 (3), the sealed storage of the first reagent 200 can be completed by filling the gap between the first barrier 300a and the inner wall of the test tube 100 with the communication liquid 400; as shown in fig. 1 (4), a second reactant 500 is added into the test tube 100, a second blocking member 300b is placed into the test tube 100, and a gap between the second blocking member 300b and the inner wall of the test tube 100 is filled with a full communicating liquid 400, so that the sealed storage of the second reactant 500 is completed, and the preparation of the test tube 100 is completed.

After the preparation of the test tube 100 is completed, the prepared test tube 100 is used for detecting the total integrated nucleic acid. As shown in fig. 7 (1), a third reaction reagent 600 is added into the test tube 100, wherein the third reaction reagent 600 comprises magnetic beads 700, a lysate and a sample to be detected, and the magnetic beads 700 are uniformly dispersed under the actions of external oscillation, ultrasound, a magnetic field and the like of the test tube 100 to complete the first-step reaction (sample lysis, magnetic bead 700 binding nucleic acid reaction); as shown in fig. 7 (2), the magnetic beads 700 are enriched on the inner wall of the cuvette 100 by moving the magnetic bead moving device 800 outside the cuvette 100; as shown in fig. 7 (3), the magnetic bead moving device 800 is moved to operate the magnetic beads 700 to pass through the magnetic bead channel 01 filled with the communication liquid 400 and enter the second reaction reagent 500, completing the second step reaction (magnetic bead washing reaction); as shown in fig. 7 (4), the magnetic beads 700 are moved to the first reaction reagent 200 to elute nucleic acids; as shown in fig. 7 (5), the magnetic beads 700 are moved into the second reaction reagent 500; as shown in fig. 7 (6), the heating device 900 is turned on to control the temperature of the first reaction reagent 200, and the detection device 1000 may perform temperature-variable PCR amplification (polymerase chain amplification), isothermal amplification, such as LAMP (loop-mediated isothermal amplification), RPA (recombinase polymerase amplification), RCA (rolling circle amplification), etc., to scan and detect the signal value of the nucleic acid amplification solution in real time, thereby obtaining the detection result.

The preparation method of the test tube and the use method of the test tube provided in the first embodiment can also be used for immunodetection. This embodiment employs a tube 100 having a smaller inside diameter, or a larger inside surface area, with the barrier layer comprising only the communication fluid 400.

Taking the immunoassay application as an example, first, a test tube 100 is prepared, and the test tube 100 may be referred to as an immunoassay test tube. Wherein the first reaction reagent 200 is an enzyme-labeled primary antibody or a fluorescent-labeled primary antibody, the second reaction reagent 500 is a magnetic bead cleaning solution, and the communicating liquid 400 is silicone oil.

As shown in fig. 8 (1), the test tube 100 is vertically placed, and the first reaction reagent 200 is added to the test tube 100; as shown in fig. 8 (2), the addition of the communication liquid 400 to the test tube 100 can be understood as filling the first reaction reagent 200 with the communication liquid 400 having a predetermined volume, thereby completing the sealed storage of the first reaction reagent 200; as shown in fig. 8 (3), the second reagent 500 is added to the test tube 100, and the communication liquid 400 is added to the test tube 100, and it is understood that the communication liquid 400 is added above the second reagent 500, and the second reagent 500 is covered and sealed with the communication liquid 400.

It should be noted that, the method for realizing stable isolation of multiple layers of liquid in the test tube by using the barrier 300 and the communicating liquid 400 is universal, and is applicable to test tubes with arbitrary shapes and also applicable to the condition that the inner diameter of the test tube 100 is smaller; the method for realizing stable isolation of the multilayer liquid only by the communicating liquid 400 is only suitable for specific scenes, and the conditions of smaller inner diameter of the test tube 100, larger specific surface area of the communicating liquid 400 and the like are required to be met simultaneously.

After the preparation of the test tube 100, the method for performing an immunoassay on the prepared test tube 100 includes: as shown in fig. 9 (1), a sample to be detected (serum sample and magnetic beads 700 modified with primary antibodies) 600 mixed with magnetic beads 700 is added into a test tube 100, and the test tube 100 is vibrated to allow the magnetic beads 700 modified with primary antibodies to be resuspended and fully combined with an antigen to be detected in serum; as shown in fig. 9 (2), the magnetic beads 700 are enriched in the inner sidewall of the cuvette 100 by the magnetic bead moving apparatus 800 outside the cuvette 100; as shown in fig. 9 (3), the beads 700 are manipulated by the bead moving device 800 to reach the second reaction reagent (cleaning liquid) 500 through the communication liquid 400, and the beads 700 are sufficiently cleaned; as shown in fig. 9 (4), the magnetic beads 700 are manipulated through the communication liquid 400 by the magnetic bead moving device 800 to reach the first reaction reagent 200, so that the magnetic beads 700 having the antigen captured therein are sufficiently mixed with the enzyme-labeled primary antibody or the fluorescent-labeled primary antibody; as shown in fig. 9 (5), a detection device (optical detection device) 1000 detects a signal value in the cuvette 100, and obtains a detection result.

Example two

As shown in fig. 10 and 11, the method for preparing a test tube and the method for using a test tube according to the second embodiment are different from the first embodiment in that: paraffin 1200, which is solid at normal temperature and is heated to be liquid, is used as the communicating liquid 400 and is used for sealing the lyophilization agent 1100.

As shown in fig. 10, in the preparation method of the test tube 100, in the case where the barrier layer includes the barrier 300 (the first barrier or the second barrier mentioned above) and the communication liquid 400, the communication liquid 400 is filled in the gap between the barrier 300 and the inner wall of the test tube 100, specifically: the heated and melted paraffin 1200 fills the gap between the barrier 300 and the inner wall of the test tube 100.

In the case that the barrier layer comprises only the communication liquid 400, or the communication liquid 400 is added into the test tube 100, specifically: the heated and melted paraffin 1200 is added to the test tube 100, and the melted paraffin 1200 is directly covered with the reaction reagent.

In the preparation method of the test tube 100, after the paraffin 1200 is cooled and solidified to form a stable isolation interface, the subsequent reaction reagent is added.

In the preparation process of the test tube 100, the freeze-drying reagent 1100 can be added in the preparation process of any reaction reagent, then the freeze-drying reagent 1100 is covered by the paraffin 1200 melted by heating, after the paraffin is cooled and solidified, a compact paraffin protection layer can be formed on the ball surface of the freeze-drying reagent 1100, and then the reaction reagent is added; the subsequent steps are the same as those of the first embodiment, and will not be described here again.

As shown in fig. 11, in the method of using the test tube 100, it is necessary to heat the paraffin 1200 (the communication liquid 400) at a position corresponding to the paraffin 1200 in the test tube 100 outside the test tube 100, so that the magnetic bead moving device can manipulate the magnetic beads to pass through the liquid paraffin 1200 and enter the next layer of reaction reagent.

In the use process of the test tube 100, only the heating device 900 is used to heat the position of the test tube 100 corresponding to the paraffin 1200, and the paraffin 1200 is melted to release the lyophilized reagent 1100, and the other steps are the same as those of the first embodiment, and will not be repeated here.

The method is compatible with the storage of liquid and fixed reagents, further increases the flexibility and stability of reagent storage, and ensures the detection performance of the test tube 100.

Example III

The preparation method of the test tube and the use method of the test tube provided in the third embodiment are mainly different from those in the first embodiment and the second embodiment: and realizing multi-index detection by adopting a spatially uniform magnetic field.

As shown in fig. 12, in the third embodiment, three magnetic bead channels are disposed between the blocking member 300 and the inner wall of the test tube 100, and the manipulation magnetic field outside the test tube 100 is a uniform magnetic field with a spatial multipoint distribution (linear distribution, spatial circumferential distribution, or spatial array distribution), which can be understood that the magnetic bead moving device 800 can generate a uniform magnetic field with a spatial multipoint distribution (linear distribution, spatial circumferential distribution, or spatial array distribution). In this way, the magnetic beads 700 can be uniformly divided into three magnetic bead stacks, and the three magnetic bead stacks respectively enter the next layer of reaction reagent through three magnetic bead channels; the bottom of the test tube 100 is provided with three mutually isolated detection cavities 101, and the detection cavities 101 are in one-to-one correspondence with the magnetic bead channels.

In the third embodiment, the number of the magnetic bead channels and the detection chamber 101 may be two, four or more than five, which is not limited in the third embodiment.

In the third embodiment, by uniformly stacking the magnetic beads 700, each magnetic bead stack enters one detection cavity 101 to detect one index, so that parallel detection of multiple indexes of a single sample can be realized.

In the third embodiment, the test tube is prepared in the same manner as in the first embodiment.

In the third embodiment, the method of using the test tube is different from the first embodiment in that: the magnetic beads 700 in the test tube 100 are uniformly piled up by an operating magnetic field (a magnetic bead moving device 800) outside the test tube 100, each magnetic bead pile is synchronously operated to sequentially pass through each layer of reaction reagent, and finally the reaction reagent enters three detection cavities 101 to realize multi-index detection.

To facilitate uniform bead stacking and moving, the bead moving device 800 includes: support ring 802 and magnet 801, wherein, support ring 802 overcoat is in test tube 100, and magnet 801 is fixed at the circumference inside wall of support ring 802, and magnet 801 is located between support ring 802 and the test tube 100 promptly, and magnet 801 is two at least and distribute in proper order along the circumference of support ring 802, and magnet 801 and detection chamber 101 one-to-one, and magnet 801 and magnetic bead passageway one-to-one.

In practical cases, the bead moving device 800 may be selected to have other structures, and is not limited to the above-described structure.

Example IV

The fourth embodiment differs from the three previous embodiments in that: the test tube 100 has different structures, different barrier layers, and different addition modes of the reaction reagent and the communication liquid.

As shown in fig. 13, in the fourth embodiment, the test tube 100 includes: a test tube 106 and a piston push rod 107; wherein, the first end of the test tube barrel 106 is provided with a first opening for inserting the first end of the piston push rod 107, and the second end of the test tube barrel 106 is provided with a second opening for leading in and out liquid; the first end of the piston push rod 107 is in sealing connection with the inner wall of the test tube barrel 106, the second end of the piston push rod 107 is located outside the test tube barrel 106, and the piston push rod 107 can reciprocate along the axial direction of the test tube barrel 106 to realize the addition and release of the reaction reagent and the communication liquid 400.

In the fourth embodiment, the barrier layer includes only the communication liquid 400.

As shown in fig. 13, the preparation method of the test tube provided in the fourth embodiment includes:

sequentially sealing and storing at least two reactants, specifically: two reagents, namely a first reagent 200 and a second reagent 500, are sequentially stored in a sealed manner.

The first reactant 200 and the second reactant 500 are stored in a sealed manner in sequence, and specifically include: s31), S32), S33), and S34).

S31) pushing the piston rod 107 to the bottom of the test tube cartridge 106, and then inserting the test tube cartridge 106 into a container (not shown) containing the first reagent 200, and pulling the piston rod 107 upward to suck the first reagent 200 into the test tube cartridge 106.

S32) the test tube 106 is inserted into a container (not shown) containing the communication liquid 400, and the piston rod 107 is pulled upward to suck the communication liquid 400 into the test tube 106, and the communication liquid 400 seals the first reactive agent 200.

S33) the cuvette holder 106 is inserted into a container (not shown) containing the second reagent 500, and the piston rod 107 is pulled upward to suck the second reagent 500 into the cuvette holder 106.

S34) the test tube 106 is inserted into a container (not shown) containing the communication liquid 400, and the piston rod 107 is pulled upward to suck the communication liquid 400 into the test tube 106, and the communication liquid 400 seals the second reactant 500.

It should be noted that S32 is not performed until the first reagent 200 is sucked; s32), after the end of the suction of the communicating liquid 400, the process proceeds to S33); after the second reagent 500 is sucked up, the process proceeds to S34).

In S31 above), the piston rod 107 is pushed to the bottom of the test tube 106 to exhaust the air in the test tube 106, so as to avoid the air from affecting the volume of the first reaction agent 200 and from affecting the normal reaction in the test tube 106.

In S32) above, the communication liquid 400 covers the first reaction reagent 200 to seal the communication liquid 400 to the first reaction reagent 200.

In S33), the communicating liquid 400 can ensure complete isolation between the first reactant 200 and the second reactant 500, and the reactant in the test tube 106 and the communicating liquid 400 move up or down integrally during the process of pulling the piston rod 107, so that the adjacent liquid surfaces remain stable.

In S34) above, the communication liquid 400 covers the second reaction reagent 500 to seal the second reaction reagent 500 by the communication liquid 400.

In the above method, the volumes of the first reactant 200, the second reactant 500, and the communication liquid 400 may be controlled by controlling the displacement of the piston rod 107.

In the above method, the movement of the piston rod 107 may be manually driven or may be driven by a driving device, which is not limited in this embodiment.

The test tube prepared by the preparation method of the test tube has two using methods, wherein in the two using methods, the first reaction reagent comprises a sample to be detected and magnetic beads, and the second reaction reagent is a magnetic bead cleaning solution.

As shown in fig. 14, the method for using the test tube provided in the fourth embodiment includes: s41), S42), S43), S44), S45), and S46).

S41) the cartridge 106 is inserted into a container (not shown) containing the third reagent 600, and the piston rod 107 is pulled upward to draw the third reagent 600 into the cartridge 106, the third reagent 600 being a nucleic acid eluent.

S42) scattering the magnetic beads 700 in the first reaction reagent 200, and fully combining the magnetic beads 700 with the sample to be detected to complete the first reaction.

S43) moves the magnetic beads 700 down the inner wall of the tube 106 into the second reaction reagent 500, and breaks up the magnetic beads 700 in the second reaction reagent 500 to complete the second reaction.

S44) moves the magnetic beads 700 down the inner wall of the test tube 106 into the third reaction reagent 600, and breaks up the magnetic beads 700 in the third reaction reagent 600 to complete the third reaction.

S45) moves the magnetic beads 700 up along the inner wall of the test tube cartridge 106 into the communication liquid 400 at the upper side of the third reaction reagent 600.

S46) presses down on the piston push rod 107 to expel the third reaction agent 600 from the cuvette 106.

In the above method, the volumes of the third reactant 600 and the communication liquid 400 may be controlled by controlling the displacement of the piston rod 107.

In S41), after the third reagent 600 is sucked, the process proceeds to S42).

In S42), the magnetic beads 700 in the first reaction reagent 200 are scattered by the magnetic bead scattering device 1300, and the magnetic beads 700 are sufficiently combined with the nucleic acid to be detected to complete the first reaction.

The above S43) is specifically: moving the magnetic bead moving device 800 to the test tube cartridge 106 to enrich the magnetic beads 700 in the first reaction reagent 200 in the inner wall of the test tube cartridge 106; moving the magnetic bead moving means 800 downward to transfer the magnetic beads 700 downward along the inner wall of the cuvette holder 106 into the communication solution 400 at the lower side of the first reaction reagent 200; and continuing to move the magnetic bead moving device 800 downwards so that the magnetic beads 700 are downwards transferred into the second reaction reagent 500 along the inner wall of the test tube 106, removing the magnetic bead moving device 800 and driving the magnetic bead scattering device 1300 to sufficiently scatter the magnetic beads 700 in the second reaction reagent 500, so as to realize the sufficient cleaning of the magnetic beads 700, namely, complete the second-step reaction.

The above S44) is specifically: moving the magnetic bead moving device 800 to the tube 106 to enrich the magnetic beads 700 in the second reaction reagent 500 in the inner wall of the tube 106; the magnetic bead moving device 800 is moved downwards to realize that the magnetic beads 700 are transferred downwards to the communication liquid 400 at the lower side of the second reaction reagent 500 along the inner wall of the test tube barrel 106; continuing to move the magnetic bead moving device 800 downwards to transfer the magnetic beads 700 into the third reaction reagent 600 downwards along the inner wall of the test tube 106, removing the magnetic bead moving device 800 and driving the magnetic bead scattering device 1300 to sufficiently scatter the magnetic beads 700 in the third reaction reagent 600, so as to realize sufficient elution of the magnetic beads 700, and complete the third reaction.

The step S45) is specifically: moving the magnetic bead moving device 800 to the test tube cartridge 106 to enrich the magnetic beads 700 in the third reaction agent 600 in the inner wall of the test tube cartridge 106; the magnetic bead moving device 800 is moved upward so that the magnetic beads 700 are transferred upward along the inner wall of the test tube cartridge 106 into the communication liquid 400 on the upper side of the third reaction agent 600.

The above S46) is specifically: a driving device (not shown) presses the piston rod 107 downwardly at a distance to expel the third reagent 600 from the test tube 106, and the third reagent 600 can be used for subsequent detection analysis.

The method of using the test tube, which may be referred to as a test tube for nucleic acid extraction, is a method of extracting nucleic acid, which completes nucleic acid extraction.

As shown in fig. 15, another method for using a test tube according to the fourth embodiment includes: s51), S52), S53), S54), S55), and S56).

S51) inserting the test tube 106 into the container containing the third reagent 600, and pulling the piston rod 107 upward to suck the third reagent 600 into the test tube 106, the third reagent 600 being a nucleic acid amplification solution.

S52) inserting the test tube 106 into the container containing the communication liquid 400, and pulling up the piston rod 107 to suck the communication liquid 400 into the test tube 106, the communication liquid sealing the third reactant 600.

S53) scattering the magnetic beads 700 in the first reaction reagent 200, and fully combining the magnetic beads 700 with the sample to be detected to complete the first reaction.

S54) moves the magnetic beads 700 down the inner wall of the tube 106 into the second reaction reagent 500, and breaks up the magnetic beads 700 in the second reaction reagent 500 to complete the second reaction.

S55) moves the magnetic beads 700 down the inner wall of the test tube 106 into the third reaction reagent 600, and breaks up the magnetic beads 700 in the third reaction reagent 600 to complete the third reaction.

S56) detecting the reaction result in the test tube.

In S51), after the third reagent 600 is sucked, the process proceeds to S52).

In S52), after the end of the suction of the communication liquid 400, the process proceeds to S53).

S53) above may refer to S42) above and S54) above may refer to S43) above, and will not be described here again.

In S55), the magnetic bead moving device 800 is moved to the tube 106 to enrich the magnetic beads 700 in the second reaction reagent 500 in the inner wall of the tube 106; the magnetic bead moving device 800 is moved downwards to realize that the magnetic beads 700 are transferred downwards to the communication liquid 400 at the lower side of the second reaction reagent 500 along the inner wall of the test tube barrel 106; and continuing to move the magnetic bead moving device 800 downwards to transfer the magnetic bead 700 into the third reaction reagent 600 along the inner wall of the test tube 106, removing the magnetic bead moving device 800 and driving the magnetic bead scattering device 1300 to sufficiently scatter the magnetic bead 700 in the third reaction reagent 600, so as to fully elute the magnetic bead 700, transferring the magnetic bead 700 to the communication liquid 400 on the upper side of the third reaction reagent 600, and heating the region of the test tube 106 where the third reaction reagent 600 is positioned by adopting the heating device 900 so as to enable the third reaction reagent 600 to perform a temperature control reaction, thereby completing the third-step reaction.

In S56), the optical signal generated by the third reagent 600 is detected by the detection device (optical detection device) 1000.

In the above method, the communication liquid 400 adjacent to the third reaction reagent 600 in the up-down direction plays a role of sealing and isolating, so that the third reaction reagent 600 is ensured not to leak to complete the third reaction step.

The method for using the test tube, which may be referred to as a nucleic acid detection test tube, is a method for detecting nucleic acid by completing the nucleic acid detection.

The specific structures of the magnetic bead moving device 800, the magnetic bead scattering device 1300, the heating device 900 and the detecting device 1000 can be referred to as the first embodiment, and will not be described herein.

In the fourth embodiment, the number of test tubes 100 may be two or more. For example, at least two test tubes 100 are sequentially distributed along a set direction; alternatively, a plurality of test tubes 100 are arranged along an array.

In order to reduce equipment and cost, all test tubes 100 can be selected to share the same driving device to drive the piston push rod 107, i.e. the same driving device is used to complete the synchronous preparation process of all test tubes 100.

After the preparation of all test tubes 100 is completed, all test tubes 100 share the same set of external mechanism (driving device, magnetic bead moving device, magnetic bead scattering device), so that the synchronous nucleic acid extraction flow of all test tubes 100 can be realized, and the method greatly improves the nucleic acid extraction flux.

Moreover, after the preparation of all test tubes 100 is completed, all test tubes 100 share the same set of external mechanism (driving device, magnetic bead moving device, magnetic bead scattering device, heating device and optical detection device), so that the synchronous detection flow of all test tubes 100 can be realized.

In practical situations, it is also possible to select that several test tubes 100 share the same driving device and share the same external mechanism, and it is not limited to all test tubes 100 share the same driving device and share the same external mechanism.

The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the application. Thus, the present application is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method of manufacturing a test tube, comprising: sequentially storing at least two reactants in a sealing way;

the reaction reagents and the blocking layers are sequentially distributed along the axial direction of the test tube, any two reaction reagents are sequentially distributed along the axial direction of the test tube, and two adjacent reaction reagents are separated by the blocking layer;

the first end of the test tube is provided with an opening, and the second end of the test tube is closed;

The gap between the position where the channel structure is not arranged and the inner wall of the test tube is smaller, the gap between the position where the channel structure is arranged and the inner wall of the test tube is larger, and the larger gap is the magnetic bead channel;

2. The method for preparing a test tube according to claim 1, wherein,

In hermetically storing at least one of the reagents, prior to adding the reagent into the tube, further comprising: adding a freeze-drying reagent into the test tube, adding molten paraffin into the test tube, covering the freeze-drying reagent by the molten paraffin, and solidifying the molten paraffin after cooling;

3. The method for preparing a test tube according to claim 1, wherein,

sequentially sealing and storing at least two reactants, specifically: sequentially storing two reaction reagents in a sealing way, wherein the two reaction reagents are a first reaction reagent and a second reaction reagent respectively;

adding the first reactant into the test tube;

Adding the second reactant into the test tube;

wherein the first barrier and the second barrier are both the barriers.

4. A method of using a test tube, wherein the test tube is prepared by the method of preparing a test tube according to claim 3;

the using method of the test tube comprises the following steps:

and detecting the reaction result in the test tube.

5. A method of using a test tube according to claim 4, wherein,

under the condition that the test tube is provided with at least two detection cavities, the magnetic bead channels are in one-to-one correspondence with the detection cavities;