CN217103835U - Card box with nucleic acid molecule amplification structure - Google Patents

Abstract

The utility model relates to the technical field of biological medical treatment, and discloses a card box with a nucleic acid molecule amplification structure, which comprises a card box plate, a front membrane and a rear membrane; the front film and the rear film are respectively attached to the front surface and the back surface of the card box plate and are sealed in the cavity on the surface of the card box plate to form a uniform mixing cavity, a blocking cavity, a PCR cavity, a seventh air passage and a flow passage; the mixing cavity is communicated with the PCR cavity through a runner, the blocking cavity is positioned between the mixing cavity and the PCR cavity, the bottom of the blocking cavity is connected into the runner, and the blocking cavity seals the runner after the PCR cavity finishes amplification reaction. The utility model discloses set up each reaction chamber and runner on the card box board to it is sealed by preceding membrane and back membrane, make nucleic acid molecule amplification process go on in closed card box, avoid polluting. And, set up between mixing chamber and PCR chamber and block the chamber structure, seal it after the PCR chamber accomplishes the amplification reaction to avoid the amplification result in the PCR chamber to leak outside the card box through the card box runner, pollute instrument or environment.

Description

Card box with nucleic acid molecule amplification structure

Technical Field

The utility model relates to the technical field of biological medical treatment, in particular to a card box with a nucleic acid molecule amplification structure.

Background

Nucleic acid detection plays a very important role in many fields of biochemical analysis, and has been widely applied in the field of biomedical science.

In the prior art, nucleic acid extraction is generally carried out by using a centrifugal column method or a magnetic bead method, and four steps of lysis, combination, washing, elution and the like are generally required. After nucleic acid extraction is carried out on the sample, subsequent steps of nucleic acid molecular hybridization, Polymerase Chain Reaction (PCR), biochip and the like are carried out continuously to complete nucleic acid detection.

The mainstream technology of molecular detection is a fluorescent quantitative PCR technology, and because the PCR technology has the characteristic of exponential amplification template, the existing open consumables easily cause PCR aerosol pollution in the whole operation process and influence the purity of the extract. Moreover, after the PCR chamber completes the amplification reaction, the structure of the sealed PCR chamber is lacked, which easily causes the amplification product in the PCR chamber to leak to the external environment through the flow channel, and pollutes the instrument or the environment.

Thus, improvements in the prior art are needed.

SUMMERY OF THE UTILITY MODEL

The utility model aims at: the utility model provides a card box with nucleic acid molecule amplification structure to solve prior art's open consumptive material and make whole operation process cause PCR aerosol to pollute easily, influence the purity of extract product, and accomplish the amplification reaction back in the PCR chamber, lack the structure in sealed PCR chamber, lead to the amplification product in the PCR chamber to leak the technical problem of outer environment through the runner easily.

In order to achieve the above object, the present invention provides a cartridge having a nucleic acid molecule amplification structure, comprising a cartridge plate, a front membrane and a rear membrane;

the surface of the cassette board is provided with a plurality of cavities, and the front film and the rear film are respectively attached to the front surface and the back surface of the cassette board to seal the cavities, so that a blending cavity, a blocking cavity, a PCR cavity, a seventh air passage and a flow passage are formed;

the mixing cavity is communicated with the PCR cavity through the runner, one end of the seventh air passage is arranged at the top of the mixing cavity, and the other end of the seventh air passage is provided with an air vent;

the blocking cavity is positioned between the blending cavity and the PCR cavity, and the bottom of the blocking cavity is connected to the flow channel; and a blocking structure is arranged in the blocking cavity so as to seal the flow channel after the PCR cavity finishes the amplification reaction.

In some embodiments of the present application, the blocking cavity is a wax valve cavity, and solid wax melted by heating is preset inside the blocking cavity.

In some embodiments of the present application, the PCR chamber comprises a central chamber, an amplification chamber, and a zone flow channel; the central cavity is a concave cavity arranged on the front surface of the cassette plate, the amplification cavity is a through hole arranged on the cassette plate, and the flow channel and the area flow channel are grooves arranged on the front surface or the back surface of the cassette plate;

the central cavity is communicated with the flow passage; the amplification cavity is provided with at least one, and the amplification cavity is communicated with the central cavity through the region flow channel.

In some embodiments of the present application, the PCR chamber further comprises a central flow channel groove, a gas containing region, a waterproof gas permeable membrane, and a region gas channel;

the central runner groove is arranged on the front surface of the central cavity, the runner is arranged on the back surface of the central cavity, the runner is communicated with one end of the central runner groove, and the other end of the central runner groove is communicated with the zone runner; the waterproof breathable film is attached to the central cavity to seal the central runner groove; one end of the air passage is arranged at the edge of the sealing area of the waterproof breathable film, and the other end of the air passage is communicated with the air containing area.

In some embodiments of the present application, two of the region channels are provided for one of the amplification chambers, and the two region channels are an entry region channel and an exit region channel; one end of the entering area flow channel is communicated with the central flow channel groove, and the other end of the entering area flow channel is communicated with one end of the amplification cavity; one end of the outlet flow channel is communicated with the other end of the amplification cavity, and the port of the other end of the outlet flow channel is positioned in the central cavity and sealed by the waterproof breathable film.

In some embodiments of the present application, a first barrier is disposed on the entrance flow channel, and a second barrier is disposed on the exit flow channel.

In some embodiments of the present application, the number of the amplification chambers is four, and the amplification chambers are arranged circumferentially at equal intervals with the center of the central chamber as a circle center.

In some embodiments of the present application, one amplification chamber and the corresponding entry flow channel and exit flow channel thereof form a wing-shaped amplification unit, and four amplification units are symmetrically arranged to form a butterfly-wing-shaped amplification unit group.

In some embodiments of the present application, the gas-containing areas are at least four and are respectively located between every two adjacent amplification units; a transition gas area is also arranged between every two adjacent gas containing areas; the transition gas area is communicated with the gas containing area adjacent to the transition gas area through a gas distributing channel.

In some embodiments of the present application, the gas containment zone and the transition gas zone are both hollow areas.

Compared with the prior art, the embodiment of the utility model provides a card box with nucleic acid molecule amplification structure, its beneficial effect lies in:

the utility model discloses card box with nucleic acid molecule amplification structure sets up each reaction chamber and runner on the card box board to sealed by preceding membrane and back membrane, make nucleic acid molecule amplification process go on in closed card box, avoid polluting. And, set up between mixing chamber and PCR chamber and block the chamber structure, seal it after the PCR chamber accomplishes the amplification reaction to avoid the amplification result in the PCR chamber to leak outside the card box through the card box runner, pollute instrument or environment.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for the embodiments or the prior art descriptions will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings without creative efforts.

Fig. 1 is an exploded view of a cartridge according to an embodiment of the present invention;

FIG. 2 is a schematic front view of the cassette plate;

FIG. 3 is a schematic view of the back side of the cassette plate;

FIG. 4 is a schematic view of a perspective structure of a main flow channel of a cassette plate;

FIG. 5 is a front isometric view of the cassette plate;

FIG. 6 is a schematic front view of the cartridge with the front film removed;

FIG. 7 is a schematic view of the back side of the cartridge of FIG. 6;

FIG. 8 is a front top structural schematic view of the cartridge of FIG. 6;

FIG. 9 is a side view schematic of the cartridge of FIG. 6;

FIG. 10 is a schematic diagram of the front structure of the PCR chamber;

FIG. 11 is a schematic diagram I of the front axial side structure of the PCR chamber;

FIG. 12 is a schematic diagram II of the front axial side structure of the PCR chamber;

FIG. 13 is a schematic view of the structure of a central chamber and one amplification chamber;

in the figure:

1. a card box body; 11. a cassette plate; 12. a front film; 13. a rear film;

101. a sample addition chamber; 102. an internal reference freeze-dried bead cavity; 103. a sample processing chamber; 104. a magnetic bead storage chamber; 105. a waste fluid chamber; 106. a magnetic bead capture chamber; 107. a mixing chamber; 108. a blocking chamber; 109. a PCR chamber; 110. siphon bending pipe; 111. a sample addition port;

2. a reagent pack group; 21. a housing; 22. sealing the film; 23. attaching an accessory; 24. a limiting strip; 121. a lysis solution bag; 122. combining the liquid bag; 123. a first cleaning solution bag; 124 an eluate package; 125. a second cleaning liquid bag;

3. sealing the cover; 4. breaking the bag;

a1, a first gas flow passage; a2, a second air channel; a3, a third air channel; a4, a fourth gas flow channel; a5, a fifth air channel; a6, sixth airway; a7, seventh airway; a8, eighth airway;

141. a first branch; 142. a second branch circuit; 143. a third branch; 144. a fourth branch; 145. a fifth branch; 146. a sixth branch; 147. a seventh branch; 148. an eighth branch; n1, a positive groove; n2, back groove; n3, a communicating hole;

b1, a first valve; b2, a second valve; b3, a third valve; b4, a fourth valve; b5, a fifth valve; b6, a sixth valve; b7, a seventh valve; b8, eighth valve; b9, ninth valve;

161. a central lumen; 162. an amplification chamber; 163. a zone flow channel; 1631. an inlet flow channel; 1632. an outlet flow channel; 164. a central runner channel; 165. a gas containing area; 1651. a first gas containment zone; 1652. a second gas containment zone; 166. a waterproof breathable film; 167. a zone gas duct; 168. an air distributing channel; 169. a transition gas zone; m1, first barrier; m2, second barrier.

Detailed Description

The following detailed description of the embodiments of the present invention is made with reference to the accompanying drawings and examples. The following examples are intended to illustrate the invention, but are not intended to limit the scope of the invention.

In the description of the present invention, it is to be understood that the terms "upper", "lower", "left", "right", "top", "bottom", and the like indicate orientations or positional relationships based on those shown in the drawings, and are only for convenience of description and simplification of description, but do not indicate or imply that the device or element referred to must have a particular orientation, be constructed and operated in a particular orientation, and thus should not be construed as limiting the present invention.

It will be understood that the terms "length," "width," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," "outer," and the like are used in an orientation or positional relationship indicated in the drawings for convenience in describing the invention and to simplify the description, and are not intended to indicate or imply that the referenced device or element must have a particular orientation, be constructed and operated in a particular orientation, and are therefore not to be considered limiting of the invention.

Furthermore, the terms "first", "second" and "first" are used for descriptive purposes only and are not to be construed as indicating or implying relative importance or implicitly indicating the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of that feature. In the description of the present invention, "a plurality" means two or more unless specifically limited otherwise.

As shown in fig. 1-4, the cartridge of the preferred embodiment of the present invention comprises a cartridge body 1, a reagent pack group 2 and a cover 3.

Specifically, the card box body 1 is provided with a main flow channel, a valve group, a reaction cavity and an airflow channel. The valve set comprises a plurality of valves arranged at the preset position of the main flow channel, so that one ends of the reaction chamber and the air flow channel are selectively communicated with the main flow channel, and the other end of the air flow channel is provided with an air vent for connecting an air pump.

The reaction chamber comprises a sample chamber, a magnetic bead storage chamber 104, a magnetic bead capture chamber 106, a waste liquid chamber 105, a mixing chamber 107 and a PCR chamber 109. The sample cavity is respectively communicated with the first ends of the magnetic bead storage cavity 104 and the magnetic bead capture cavity 106, the first end of the magnetic bead capture cavity 106 is also communicated with the mixing cavity 107, the opposite second end of the mixing cavity is communicated with the waste liquid cavity 105, and the mixing cavity 107 is communicated with the PCR cavity 109. The sample chamber has a sample addition port 111, and the cover 3 is used to seal the sample addition port 111. Magnetic particles, such as freeze-dried magnetic beads, are pre-disposed in the magnetic bead storage chamber 104. Freeze-dried beads are pre-placed in the mixing chamber 107.

The reagent pack group 2 is attached to the surface of the cartridge body 1, and includes at least four independently sealed reagent pack units, where the four reagent pack units are a lysis solution pack 121, a binding solution pack 122, a first cleaning solution pack 123 and an eluent pack 124, respectively, and may further preferably include a second cleaning solution pack 125 (in fig. 2 and 3, the reference numerals are only schematic of the positions of the reagent pack units, and do not indicate complete reagent pack units).

The gas flow channels include at least a first gas flow channel A1, a second gas flow channel A2, a third gas flow channel A3 and a fourth gas flow channel A4 corresponding to the lysis pad 121, the binding pad 122, the first wash pad 123 and the elution pad 124, respectively, and may preferably further include a fifth gas flow channel A5 corresponding to the second wash pad 125.

The lysate bag 121 is broken and flows into the first gas flow channel a1, and the first gas flow channel a1 may be communicated with the magnetic bead storage chamber 104 or the sample chamber.

The binding packet 122 is broken and flows into the second flow channel A2, and the second flow channel A2 can be in communication with the magnetic bead storage chamber 104 or in communication with the sample chamber.

The above steps are mainly for mixing the lysis solution, the magnetic beads (magnetic particles) in the magnetic bead storage chamber 104, and the binding solution in the sample chamber. When the first air flow channel a1 is communicated with the magnetic bead storage chamber 104, i.e. the lysis solution carries the magnetic beads to the sample chamber, the binding solution directly flows into the sample chamber, and thus the second air flow channel a2 is communicated with the sample chamber. When the first air flow channel a1 directly communicates with the sample chamber, i.e. the binding solution carries the magnetic beads to the sample chamber, the second air flow channel a2 needs to communicate with the magnetic bead storage chamber 104.

The first cleaning solution bag 123 is broken and flows into the third flow channel A3, and the third flow channel A3 can be connected to the first end of the sample chamber or the magnetic bead capture chamber 106. This step is mainly to wash the magnetic beads, and to make the washing solution pass through the magnetic bead capture chamber 106 to wash the magnetic beads, and then inject the washing solution into the waste liquid chamber 105. Therefore, the third flow channel a3 can be directly connected to the first end of the magnetic bead capture chamber 106 for magnetic bead cleaning, or connected to the sample chamber, so that the cleaning solution flows through the sample chamber first and then flows into the magnetic bead capture chamber 106 for magnetic bead cleaning.

The second cleaning solution bag 125 is broken and flows into the fifth flow channel a5, and the fifth flow channel a5 can be connected to the first end of the sample chamber or the magnetic bead capturing chamber 106. The principle and the steps of this step are the same as those of the first washing, and are not described herein again. The second washing of the magnetic beads is a preferred embodiment and is not an essential step.

After the eluent packet 124 is broken, the eluent flows into the fourth gas flow channel a4, and the fourth gas flow channel a4 is communicated with the second end of the magnetic bead capture cavity 106. This step is mainly to elute the nucleic acid molecules adsorbed on the surface of the magnetic beads, and then push the eluent into the mixing chamber 107 to mix the eluent with the lyophilized beads in the mixing chamber 107.

After the eluent and the freeze-dried beads are mixed in the mixing chamber 107, the mixture is pushed to the PCR chamber 109 for amplification.

In the above steps, the valve set and the air pump control the flow of the liquid.

According to the above process steps, the present application proposes a preferred embodiment of the cartridge as follows:

referring to fig. 1-4, the cartridge comprises a cartridge body 1, a reagent pack set 2 and a cover 3, wherein the cartridge body 1 comprises a cartridge plate 11, a front membrane 12 and a rear membrane 13. The front film 12 and the rear film 13 are respectively attached to the front surface and the back surface of the cassette board 11 for sealing the cavity opening, the slot and the hollow-out position on the front surface and the back surface of the cassette board 11. In the following description, the descriptions relating to the left, right, up, and down orientations are based on the front view of the cartridge board 11.

In this embodiment, the sample chamber preferably includes a sample addition chamber 101, a reference freeze-dried bead chamber 102, a sample processing chamber 103, and a siphon trap 110, which are disposed at predetermined positions in the middle of the front surface of the cartridge plate 11. The sample addition chamber 101 has a sample addition port 111 at the top, a sample to be tested is injected into the cartridge from the sample addition port 111, and the cover 3 is used for closing the sample addition port 111. The internal reference freeze-dried bead cavity 102 is located below the sample adding cavity 101, the top of the internal reference freeze-dried bead cavity 102 is communicated with the bottom of the sample adding cavity 101, and the bottom of the internal reference freeze-dried bead cavity 102 is communicated with one end of the siphon elbow 110. The sample processing cavity 103 is located at the left side of the sample adding cavity 101, the other end of the siphon bend 110 is communicated with the top of the sample processing cavity 103 at the right side, a sixth air passage A6 is arranged at the top of the left side of the sample processing cavity 103, and an air vent is arranged at the other end of the sixth air passage A6. The internal reference freeze-dried beads are preset in the internal reference freeze-dried bead cavity 102 and serve as internal reference samples, and the internal reference freeze-dried beads have the function of monitoring the whole card box process. If the internal reference freeze-dried bead sample can have a fluorescence reaction, and the sample to be detected does not have a fluorescence reaction, the problem of the sample to be detected is indicated.

The magnetic bead storage cavity 104 is positioned at the left side of the sample processing cavity 103, the mixing cavity 107 is positioned below the siphon bent pipe 110, a seventh air passage A7 is arranged at the central line of the top of the mixing cavity 107, and an air vent is arranged at the other end of the seventh air passage A7. The waste liquid cavity 105 is positioned at the right side of the sample adding cavity 101, the top of the waste liquid cavity 105 is provided with an eighth air passage A8, and the other end of the eighth air passage A8 is provided with a vent hole which can be used as an exhaust hole. The PCR chamber 109 is provided at the right side of the front face of the cassette plate 11.

Furthermore, a blocking cavity 108 is arranged between the mixing cavity 107 and the PCR cavity 109. The blocking chamber 108 is located below the sample addition chamber 101 and is connected to a branch (main channel) between the mixing chamber 107 and the PCR chamber 109. Further, the solid wax is arranged at the inner top of the blocking cavity 108, and is melted when heated to a preset temperature, and the wax seals the main flow channel, so that the communication between the blending cavity 107 and the PCR cavity 109 is blocked, and the mixed solution cannot continuously circulate between the blending cavity 107 and the PCR cavity 109.

The reagent pack group 2 is attached to the back of the left upper side of the cassette board 11 and includes five independent sealed reagent pack units, and the five reagent pack units are a lysis solution pack 121, a binding solution pack 122, a first cleaning solution pack 123, an elution solution pack 124 and a second cleaning solution pack 125 (in the figure, the reference numerals are only schematic for the positions of the reagent pack units, and do not represent the complete reagent pack units).

Referring to fig. 4, the main flow path on the cassette board 11 includes a first branch 141, a second branch 142, a third branch 143, a fourth branch 144, a fifth branch 145, a sixth branch 146, a seventh branch 147, and an eighth branch 148.

One end of the first branch 141 is connected to the liquid outlet end of the first air flow channel a1, and the other end of the first branch 141 is connected to the magnetic bead storage chamber 104, the first valve B1, and the sample processing chamber 103 in sequence.

One end of the second branch 142 is connected to the liquid outlet end of the second flow channel a2, and the other end of the second branch 142 is connected to the first branch 141 after being connected to the second valve B2, where the connection point is marked as C1.

One end of the third branch 143 is connected to the liquid outlet end of the third flow channel a3, and the other end of the third branch 143 is connected to the first branch 141 after being sequentially connected to the fourth valve B4 and the fifth valve B5, where the access point is marked as C2.

One end of the fourth branch 144 is connected to the liquid outlet of the fourth gas flow channel a4, and the other end of the fourth branch 144 is connected to the seventh valve B7 and one end (second end) of the magnetic bead capturing cavity 106 near the left side of the cartridge board 11.

One end of the fifth branch 145 is connected to the liquid outlet end of the fifth flow channel a5, the other end of the fifth branch 145 is connected to the third branch 143, and the access point C3 is located between the fourth valve B4 and the fifth valve B5.

One end of the sixth branch 146 is communicated with the bottom of the sample processing chamber 103 (or connected to the section of the first branch 141 close to the sample processing chamber 103), and the other end of the sixth branch 146 is communicated with the third valve B3 and then communicated with one end (first end) of the magnetic bead capturing chamber 106 close to the right side of the cassette plate 11.

One end of the seventh branch 147 is connected to the upper portion of the waste liquid chamber 105, and the other end of the seventh branch 147 is connected to the sixth valve B6 and one end (second end) of the magnetic bead capturing chamber 106 near the left side of the cartridge plate 11.

One end of the eighth branch 148 is communicated with one end (first end) of the magnetic bead capturing cavity 106 close to the right side of the cartridge plate 11, and the other end of the eighth branch 148 is sequentially communicated with the eighth valve B8, the mixing cavity 107, the blocking cavity 108, the ninth valve B9 and the PCR cavity 109.

In the above embodiment, referring to fig. 1 to 9, the sample adding chamber 101, the reference freeze-dried bead chamber 102, the sample processing chamber 103, the waste liquid chamber 105, the mixing chamber 107, and the blocking chamber 108 are cavities formed by protruding the front surface of the cartridge board 11 toward the back surface thereof, and the cavities are sealed when the front film 12 is attached to the front surface of the cartridge board 11 to form the above reaction chambers. Further, the sections of the sample addition chamber 101 and the sample processing chamber 103 near the bottom are inclined toward the front surface of the cartridge plate 11 to facilitate the liquid outflow. A groove is cut in the front face of the cartridge plate 11 and sealed by the front membrane 12 to form a siphon trap 110. Referring to fig. 7, a hollow hole is formed in the cartridge board 11, a protrusion 1041 slightly protruding out of the back surface of the cartridge board 11 is disposed at a position around the edge of the hollow hole on the back surface of the cartridge board 11, the protrusion 1041 is provided with a flow channel to communicate with the first valve B1, and the front surface of the hollow hole is sealed by the front film 12 to form the magnetic bead storage chamber 104.

In the above embodiments, the main flow channel includes the first branch 141, the second branch 142, the third branch 143, the fourth branch 144, the fifth branch 145, the sixth branch 146, the seventh branch 147 and the eighth branch 148, which are all formed by grooves formed on one surface (front surface or back surface) of the cassette board 11. Further, this application provides a positive and negative groove structure, and above-mentioned branch road all can comprise this positive and negative groove structure of at least one. Referring to fig. 10, taking the eighth branch 148 as an example, the front-back groove structure includes a front groove N1, a back groove N2 and a communication hole N3. The front groove N1 is a groove (solid line in the figure) opened on the front surface of the cartridge board 11, the back groove N2 is a groove (dotted line in the figure) opened on the back surface of the cartridge board 11, and the communication hole N3 is provided at the intersection of the ports of the front groove N1 and the back groove N2, and is used for communicating the ports of the front groove N1 and the back groove N2. The front film 12 seals the front groove N1, the rear film 13 seals the rear groove N2, and the communication hole N3 communicates the front groove N1 and the rear groove N2, thereby forming a flow channel.

In the above embodiment, referring to FIGS. 10-12, the PCR chamber 109 comprises a central chamber 161, an amplification chamber 162, a zone flow channel 163, a central flow channel groove 164, a gas containing zone 165, a waterproof gas permeable membrane 166, a zone gas channel 167, a gas distributing channel 168, and a transition gas zone 169.

The central channel groove 164 is formed on one surface (front surface) of the central chamber 161, the channel (i.e., the section of the back recess N2 of the eighth branch 148) is formed on the other surface (back surface) of the central chamber 161 opposite to the central chamber 161, the eighth branch 148 is communicated with one end of the central channel groove 164, and the other end of the central channel groove 164 is communicated with the block channel 163. A waterproof, breathable membrane 166 is attached to the central cavity 161 to seal the central runner channel 164. The regional air channel 167 is arranged at the edge of the sealing area of the waterproof breathable film 166, and the regional air channel 167 is communicated with the air containing region 165. Referring to fig. 12, the height of the central channel 164 is about half of the height of the central cavity 161. when the waterproof membrane 166 is attached to the central cavity 161, the upper half of the central cavity 161 is not sealed by the waterproof membrane 166, and the gas channel 167 is actually a channel between the upper half of the central cavity 161 and the gas containment region 165. In this embodiment, a waterproof air-permeable membrane 166 is disposed on the opposite surface (front surface) of the liquid inlet end (the eighth branch 148 disposed on the back surface), and the sealed edge thereof is communicated with an air-containing region 165. When the mixed liquid in the mixing chamber 107 enters the central channel groove 164 of the central chamber 161, the carried air bubbles will be released from the waterproof and air-permeable membrane 166 and enter the air-holding region 165, so that the mixed liquid entering the amplification chamber 162 does not carry air bubbles, thereby facilitating the amplification step.

Further, referring to fig. 10 and 13, one amplification chamber 162 is correspondingly provided with two region flow channels 163, and the two region flow channels 163 are an inlet region flow channel 1631 and an outlet region flow channel 1632, respectively. One end of the inlet flow channel 1631 is communicated with the central flow channel groove 164, and the other end of the inlet flow channel 1631 is communicated with one end of the amplification chamber 162. One end of the exit flow channel 1632 is communicated with the other end of the amplification chamber 162, and the port of the other end of the exit flow channel 1632 is located in the central chamber 161 and sealed by the waterproof and breathable membrane 166. The mixed liquid in the central channel groove 164 enters the amplification chamber 162 through the inlet channel 1631, when the amplification chamber 162 is filled with the mixed liquid, the excess mixed liquid flows into the outlet channel 1632, and when the outlet channel 1632 is filled, the mixed liquid cannot pass through the waterproof air-permeable membrane 166, so that the mixed liquid stops entering the inlet channel 1631.

Further, a first barrier M1 is disposed on the entrance runner 1631, and a second barrier M2 is disposed on the exit runner 1632. In practice, the first barrier M1 and the second barrier M2 are flow path interrupting structures of a section of the film 12 before welding. Referring specifically to fig. 13, when the front film 12 is welded on the surface of the cassette board 11 by laser, the welding line is about 0.1mm away from the outside of the flow channel, i.e., the area outside the dotted line strip circle X in the figure is the welding area. Since the first barrier M1 and the second barrier M2 are non-welded regions, and the front film 12 is only attached to the surface of the cartridge board 11, when the flow rate or pressure of the mixture is high, the front film 12 at the barrier is pressed to arch the front film 12 outwards, and the mixture can still flow through the barrier (as shown by the arrows in the figure). First barrier M1 and second barrier M2 can act as a flow barrier but do not completely block the flow of liquid so that mixed liquid can flow between inlet channel 1631, amplification chamber 162, and outlet channel 1632. When the amplification chamber 162 is filled and before the temperature rise and drop cycle is performed, it is necessary to completely block the flow of the inlet flow channel 1631 and the outlet flow channel 1632, and at this time, the waterproof and breathable membrane 166 is a first barrier, and the top pressure rod of the matching instrument is used to press the front membrane 12 at the first barrier M1 and the second barrier M2 to form a second barrier, which ensures that the flow of the inlet flow channel 1631 and the outlet flow channel 1632 is completely blocked.

Further, four amplification chambers 162 are provided, and are arranged circumferentially at equal intervals around the center of the central chamber 161. One amplification cavity 162 and the corresponding entry flow channel 1631 and exit flow channel 1632 form a wing-shaped amplification unit, and the four amplification units are symmetrically arranged to form a wing-shaped amplification unit group. Further, at least four gas-containing regions 165 are provided and are respectively located between every two adjacent amplification units. A transition air area 169 is also arranged between every two adjacent air containing areas 165, and the transition air area 169 is communicated with the air containing area 165 adjacent to the transition air area through an air distributing channel 168. The central cavity 161 and the waterproof breathable membrane 166 are both circular, the center of the central cavity 161 is taken as the circle center, the position of the flow channel is 0 degree, and the four amplification cavities 162 are respectively positioned at the positions of 45 degrees, 135 degrees, 225 degrees and 315 degrees on the circle. Specifically, the air-containing region 165 includes a first air-containing region 1651 and a second air-containing region 1652. The first air-containing area 1651 is provided with two air-containing areas, which are respectively located at the axes of 90 ° and 270 °, and the shape of each air-containing area is similar to a triangle corresponding to the fin-shaped gap, one angle of each air-containing area is communicated with the central cavity 161 through the air-distributing channel 168, and the other two angles of each air-containing area are respectively communicated with one end of the transition air area 169 through the air-distributing channel 168. The second air-containing area 1652 is provided with four air-distributing channels 168, two of which are respectively located at two sides of the flow channel and are communicated with each other, and the other two air-distributing channels 168 and the air-distributing channels 167 are symmetrically arranged at two opposite sides of the flow channel and are communicated with the central cavity 161. The other end of transition gas zone 169 communicates with second gas containing zone 1652 via gas dividing duct 168. Referring to fig. 11-12, the air containing region 165 and the transition air region 169 are hollow regions, the air dividing duct 168 and the air dividing duct 167 are grooves formed on the card case body 1, and the arrangement form of the region flow passage 163 is a positive and negative groove structure form. After the mixed liquid in the mixing chamber 107 flows into each amplification chamber 162, the optical module of the matched instrument is positioned at the corresponding position of the front PCR chamber 109 of the card box, the thermal module is positioned at the corresponding position of the back PCR chamber 109 of the card box, and the optical module and the thermal module clamp the card box from the two sides of the card box. During amplification, the thermal module cyclically heats and cools the amplification chamber 162. After amplification is completed, it is necessary to excite the amplified reactant in the amplification chamber 162 with light of a different wavelength to detect the concentration of the amplified reactant in the amplification chamber 162. Specifically, the optical module includes four small modules, namely a red module, a green module, a yellow module and a blue module, the four small modules are respectively disposed on the four amplification chambers 162 to detect different types of fluorescence after reaction of reagents in the amplification chambers 162, and each amplification chamber 162 needs to receive detection of the four small modules in turn. Because the four amplification cavities 162 are designed on one circumference, the optical module can correspondingly detect different amplification cavities 162 by the four small modules in turn only by rotating, and the optical module does not need to move an extra stroke, thereby effectively improving the detection efficiency.

In the above embodiment, referring to fig. 1, the reagent pack group 2 is attached to the back surface of the left upper side of the cassette board 11, and the reagent pack group 2 includes the housing 21, the sealing film 22, and the attachment 23. The casing 21 is provided with five liquid containing cavities with openings facing the front, and the sealing film 22 is attached to the front of the casing 21 to seal the liquid containing cavities, so as to form five reagent pack units, namely a corresponding lysis solution pack 121, a binding solution pack 122, a first cleaning solution pack 123, an elution solution pack 124 and a second cleaning solution pack 125. One surface of the attachment 23 is used for connecting the front surface of the shell 21, the other surface of the attachment 23 is used for connecting the back surface of the cassette board 11, and a hollow part is formed at a position corresponding to the liquid containing cavity on the attachment 23, namely the attachment 23 does not shield the sealing film 22.

Further, a bale breaking piece 4 is also included. The bag breaking piece 4 is arranged at a position with a preset distance from the sealing film 22, and the bag breaking piece 4 breaks the sealing film 22 when being subjected to preset pressure.

Further, referring to fig. 3 and 7, the back of the cassette board 11 is provided with a plurality of position-limiting strips 24 fitting the shape of the housing 21, and the edges of the attachment 23 are located in the position-limiting strips 24 to prevent the reagent pack set 2 from shifting.

The utility model also provides a detection device, including above-mentioned card box to and the supporting instrument that uses with the card box cooperation. The supporting instrument mainly includes: the air pump is connected with the air port of each air passage; the positions corresponding to the valves are provided with driving modules (central rods) which are used for driving the valves to open and close; magnetic attraction modules are arranged at positions corresponding to the magnetic bead capture cavity 106 and the sample processing cavity 103 and used for providing magnetic attraction force and controlling attraction or release of magnetic beads; an ultrasonic module and a thermal cycle module are arranged at the positions corresponding to the sample processing cavity 103 and can be used for uniformly mixing and heating the sample mixed liquid; a pressing module is arranged at the position corresponding to the bag breaking piece 4 and used for providing preset pressing force and extruding the bag breaking piece 4 to break the sealing film of the reagent bag; and an amplification heating module and an optical detection module are arranged at the position corresponding to the PCR cavity 109, the amplification heating module is used for heating and cooling the PCR cavity, and the optical detection module is used for detecting the amplification condition in the PCR cavity.

The working process of the cartridge structure provided by the above embodiment is as follows:

first step, loading sample: the sample of the patient to be tested is introduced into the sample addition chamber 101 from the sample addition port 111, and then the cover 3 is fastened, thereby sealing the cartridge. Because the siphon bend 110 is disposed between the sample addition chamber 101 and the sample processing chamber 103, the sample temporarily stays in the siphon bend 110 and does not directly flow into the sample processing chamber 103.

Step two, connecting a matched instrument: the cartridge is inserted into a mating instrument. Specifically, the matched instrument is provided with a slot, so that the card box can be vertically inserted into the matched instrument. Vents of the first air channel A1, the second air channel A2, the third air channel A3, the fourth air channel A4, the fifth air channel A5, the sixth air channel A6 and the seventh air channel A7 on the cartridge body 1 are all connected with an air pump of a matched instrument, and an air pressure valve can be arranged at the connection position. Moreover, the center rods of the matching apparatus are respectively aligned with the valves (the first valve B1, the second valve B2, the third valve B3, the fourth valve B4, the fifth valve B5, the sixth valve B6, the seventh valve B7, the eighth valve B8 and the ninth valve B9), and the initial states of all the center rods are all the states of pressing the elastic pad, so as to close all the valves, and make all the channels in the cartridge body 1 in the closed state.

Step three, sample pretreatment: sample pretreatment: after sample application, the sample is mixed with the internal reference lyophilized beads in the internal reference lyophilized bead cavity, and then reaches the siphon elbow 110 to stay. After the test is initiated, the sample residing in the siphon bend 110 is drawn into the sample processing chamber 103 by using the associated instrument air pump to draw air from the vent of the sixth air passage a6 (the following description will not refer to the operation of the associated instrument air pump, but will only describe the state of the air passage being vented or evacuated).

Step four, releasing lysate: the pressing module of the kit presses the area of the bag breaking member 4 corresponding to the lysis solution bag 121, so as to puncture the sealing membrane 22 of the lysis solution bag 121, and the lysis solution flows out of the lysis solution bag 121 to the first air flow channel a1 (the following description does not refer to the operation of the pressing module of the kit, and only describes the state of the reagent bag). Then, the center lever corresponding to the first valve B1 is released, thereby opening the first valve B1 (the following description does not refer to the operation of the center lever of the associated instrument, and only the on-off state of the valve is described). Meanwhile, air is introduced from the first air flow channel A1, so that the lysate flows into the magnetic bead storage cavity 104, freeze-dried magnetic beads in the magnetic bead storage cavity 104 are uniformly mixed, and all the magnetic beads are carried to flow into the sample processing cavity 103. By continuing to introduce air from the first air flow channel a1, the lysate is thoroughly mixed with the sample by the continuous generation of air bubbles from the bottom of the sample processing chamber 103, and then the first valve B1 is closed. In the mixing process, the mixed solution can be heated to 85 ℃ through a thermal cycle module of a matched instrument for thermal cracking.

Step five, releasing the binding solution: the binding liquid pack 122 is punctured, and the binding liquid flows out of the binding liquid pack 122 to the second flow channel a 2. The second valve B2 is opened and air is bled from the second flow channel a2 to push the binding fluid into the sample processing chamber 103. And continuously introducing air from the second air flow channel a2, mixing the binding solution and magnetic beads with the lysed sample by using the bubbles continuously generated from the bottom of the sample processing chamber 103, and then closing the second valve B2.

In the above steps, a mode of firstly releasing the lysis solution and mixing the lysis solution with the magnetic beads in the sample processing chamber 103, and then releasing the binding solution to the sample processing chamber 103 for mixing together is adopted. However, in the present embodiment, a manner of releasing the lysis solution into the sample processing chamber 103, releasing the binding solution to mix with the magnetic beads, and then mixing with the magnetic beads in the sample processing chamber 103 may also be adopted, and the specific ventilation and valve control in this manner are the same as in the above manner, and will not be described again here.

Sixthly, cleaning magnetic beads: in this embodiment, there are two types of magnetic bead cleaning methods.

The first magnetic bead cleaning mode is to adsorb the magnetic bead in the magnetic bead capturing cavity 106 for cleaning, and the magnetic module is only arranged at the corresponding position of the magnetic bead capturing cavity 106 by the matched instrument. The method comprises the following specific steps: the third valve B3 and the sixth valve B6 are opened, and simultaneously, the magnetic module is started by the matched instrument. Air is introduced through the sixth air passage a6 to push the mixed solution in the sample processing chamber 103 into the magnetic bead capture chamber 106 and the waste liquid chamber 105 in sequence. When the mixed liquid passes through the magnetic bead capturing cavity 106, the magnetic beads are uniformly adsorbed on the inner wall of the magnetic bead capturing cavity 106 under the action of the magnetic attraction force of the magnetic attraction module, and the rest liquid part of the mixed liquid is completely pushed into the waste liquid cavity 105, and then the third valve B3 and the sixth valve B6 are closed. The first cleaning liquid pack 123 is punctured, and the cleaning liquid flows out from the first cleaning liquid pack 123 to the third flow channel a 3. The fourth valve B4 and the fifth valve B5 are opened, air is introduced from the third air flow channel A3, the wash solution is pushed into the sample processing chamber 103, and then the fourth valve B4 and the fifth valve B5 are closed. And opening a third valve B3 and a sixth valve B6, introducing air from a sixth air channel A6, and sequentially pushing the cleaning solution into the magnetic bead capture cavity 106 and the waste liquid cavity 105 to complete the first cleaning of the magnetic beads. The second bag of wash solution 125 is further punctured, the fifth valve B5 is opened, air is vented from the fifth flow channel a5, the wash solution is pushed into the sample processing chamber 103, and the fifth valve B5 is closed. And opening a third valve B3 and a sixth valve B6, introducing air from a sixth air channel A6, and sequentially pushing the cleaning solution into the magnetic bead capture cavity 106 and the waste liquid cavity 105 to finish the second magnetic bead cleaning. Air is continuously introduced from the sixth air passage A6 to dry the magnetic bead capture cavity 106.

The second kind of magnetic bead cleaning method is to adsorb the magnetic bead and carry out a washing in sample processing chamber 103, carries out the secondary and washs in magnetic bead capture chamber 106, and supporting instrument all sets up magnetism and inhales the module in magnetic bead capture chamber 106 and sample processing chamber 103 corresponding position department. The method comprises the following specific steps: the mating instrument starts the magnetic module at the corresponding position of the sample processing chamber 103, opens the third valve B3 and the sixth valve B6, and introduces air from the sixth air passage a 6. Under the action of the magnetic attraction of the magnetic module, the magnetic beads are uniformly adsorbed on the inner wall of the sample processing chamber 103, the rest of the liquid of the mixture is pushed into the waste liquid chamber 105, and then the third valve B3 and the sixth valve B6 are closed. The first cleaning liquid pack 123 is punctured, and the cleaning liquid flows out from the first cleaning liquid pack 123 to the third flow channel a 3. The fourth valve B4 and the fifth valve B5 are opened to allow air to flow through the third flow channel A3 and to push the wash solution into the sample processing chamber 103. Then, the magnetic module is turned off, air is continuously introduced from the third air flow channel a3, the cleaning solution and the magnetic beads are fully mixed by the bubbles continuously generated from the bottom of the sample processing chamber 103, and then the fourth valve B4 and the fifth valve B5 are closed. The magnetic module in the magnetic bead capturing chamber 106 is opened, and the third valve B3 and the sixth valve B6 are opened, so that air is introduced from the sixth air channel a 6. Under the magnetic attraction of the magnetic module, the magnetic beads are uniformly adsorbed on the inner wall of the magnetic bead capturing chamber 106, the remaining liquid portion of the mixture is pushed into the waste liquid chamber 105, and then the third valve B3 and the sixth valve B6 are closed. The second bag of wash solution 125 is further punctured, the fifth valve B5 is opened, air is vented from the fifth flow channel a5, the wash solution is pushed into the sample processing chamber 103, and the fifth valve B5 is closed. And opening a third valve B3 and a sixth valve B6, introducing air from a sixth air channel A6, and sequentially pushing the cleaning solution into the magnetic bead capture cavity 106 and the waste liquid cavity 105 to finish the second magnetic bead cleaning. And continuously introducing air from the sixth air channel A6 to blow dry the reagents remained on the surfaces of the magnetic beads in the magnetic bead capture cavity 106.

Seventh step, releasing the eluent: the eluent packet 124 is punctured such that eluent flows from the eluent packet 124 to the fourth gas flow path a 4. The seventh valve B7 and the eighth valve B8 are opened, and air is introduced from the fourth air channel a4, so that the eluents are sequentially pushed into the magnetic bead capture chamber 106 and the mixing chamber 107. And continuously introducing air from the fourth air channel A4, mixing the eluent with the freeze-dried beads in the blending cavity 107 by utilizing the bubbles continuously generated from the bottom of the blending cavity 107, and then closing the seventh valve B7 and the eighth valve B8.

Eighth step, pushing the eluent into the PCR cavity: the ninth valve B9 is opened, and air is introduced from the seventh air passage A7 to push the eluent in the mixing chamber 107 into the PCR chamber 109. Because the waterproof and air permeable membrane 166 is disposed in the PCR chamber 109, when the four amplification chambers 162 are filled, the mixture stops flowing into the PCR chamber 109, and the ninth valve B9 is closed.

Ninth step, liquid flow separation: the pressurizing module of the matching instrument is used to press against the first partition M1 and the second partition M2, so that the PCR cavity 109 is separated from other flow channels.

Step ten, performing an amplification cycle: the 4 amplification zones 162 of the PCR chamber 109 are subjected to temperature rise and decrease cycles by an amplification heating module of a matching instrument, and the amplification result is detected by an optical detection module.

Step ten, sealing a flow passage: after the experiment is finished, the heating module of the matched instrument is utilized to heat the wax in the wax valve cavity (blocking the cavity 108) so that the wax is melted and naturally falls down under the action of gravity, and the runner between the uniformly mixing cavity 107 and the PCR cavity 109 is blocked after solidification, so that the effect of sealing the PCR cavity 109 is achieved, and the phenomenon that an amplification product in the PCR cavity leaks out of the card box through the runner of the card box to pollute the instrument or the environment is avoided.

To sum up, the utility model provides a card box, with necessary reaction chamber if sample chamber, magnetic bead storage chamber 104, magnetic bead capture chamber 106, waste liquid chamber 105, mixing chamber 107 and PCR chamber 109 rationally locate card box body 1 on to independently seal the required reagent of reaction and establish in pasting the reagent package on card box body 1, solved the sealed problem of depositing of reagent. Preferably, thermoplastic elastic glue is used as the bale breaking piece 4 to break the reagent bag.

In addition, the card box body 1 is reasonably provided with an air flow channel, a main flow channel and a valve (a specially designed needle ejection valve), the flowing direction and the stopping position of the sample and the reagent are controlled through the valve, and the flowing of the liquid in the card box body 1 is controlled in a mode of driving the liquid by an air pump, so that the reagent bag is selectively communicated with the reaction cavity, and the control is simple and reliable.

Also, when the liquid in the cartridge is controlled using the air pump, or when the liquid reacts in the reaction chamber, bubbles are easily generated. The prior art uses cartridges of horizontal construction, where the bubbles, which are generated when pushed by a reaction or air pump, are not easily removed. The card box of the utility model is of a vertical structure, and bubbles generated inside during reaction can be emitted from the bottom to the top, and can be easily eliminated. And a waterproof air-permeable membrane 166 is arranged at the PCR cavity 109, so that the mixed solution of the eluent and the freeze-dried beads can remove air bubbles before entering the PCR cavity 109, and the mixed solution entering the PCR cavity 109 is ensured not to contain air bubbles.

Therefore, the utility model provides a card box and detection device collects molecular diagnosis nucleic acid extraction, amplification, detects in an organic whole, has realized automatic, the closed testing process. The device is used for detection, the detection time is less than one hour, the manual operation is not more than 2 minutes, and the device can be used for detecting pathogen genome targets of various human clinical samples and has wide applicability. The preservation and the transportation of this application device are normal atmospheric temperature condition, need not the cold chain, and the economic nature is high. The utility model discloses the device card box is totally closed at the testing process, and its testing result is identical with conventional method completely moreover, need not the professional training and can operate, and is safe convenient.

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, a plurality of modifications and replacements can be made without departing from the technical principle of the present invention, and these modifications and replacements should also be regarded as the protection scope of the present invention.

Claims (10)

1. A cartridge having a nucleic acid molecule amplification structure, characterized by comprising a cartridge plate (11), a front membrane (12) and a rear membrane (13);

the surface of the cassette board (11) is provided with a plurality of cavities, the front film (12) and the rear film (13) are respectively attached to the front surface and the back surface of the cassette board (11) to seal the cavities, so that a uniform mixing cavity (107), a blocking cavity (108), a PCR cavity (109), a seventh air passage (A7) and a flow passage are formed;

the blending cavity (107) is communicated with the PCR cavity (109) through the flow channel, one end of the seventh air channel (A7) is arranged at the top of the blending cavity (107), and the other end of the seventh air channel is provided with an air vent;

the blocking cavity (108) is positioned between the blending cavity (107) and the PCR cavity (109), and the bottom of the blocking cavity (108) is connected to the flow channel; a blocking structure is arranged in the blocking cavity (108) to seal the flow channel after the PCR cavity (109) completes the amplification reaction.

2. The cartridge with a nucleic acid molecule amplification structure according to claim 1, wherein the blocking chamber (108) is a wax valve chamber (151) inside which a solid wax melted by heat is pre-disposed.

3. The cartridge having a nucleic acid molecule amplification structure according to claim 1, wherein the PCR chamber (109) comprises a central chamber (161), an amplification chamber (162), and a zone flow channel (163); the central cavity (161) is a cavity arranged on the front surface of the cassette plate (11), the amplification cavity (162) is a through hole arranged on the cassette plate (11), and the flow channel and the zone flow channel (163) are grooves arranged on the front surface or the back surface of the cassette plate (11);

the central cavity (161) is in communication with the flow passage; the amplification cavity (162) is provided with at least one, and the amplification cavity (162) is communicated with the central cavity (161) through the region flow channel (163).

4. The cartridge having a nucleic acid molecule amplification structure according to claim 3, wherein the PCR chamber (109) further comprises a central flow channel groove (164), a gas containing region (165), a water-proof gas-permeable membrane (166), and a regional gas channel (167);

the central runner groove (164) is arranged on the front surface of the central cavity (161), the runner is arranged on the back surface of the central cavity (161), the runner is communicated with one end of the central runner groove (164), and the other end of the central runner groove (164) is communicated with the zone runner (163); the waterproof breathable film (166) is attached to the central cavity (161) to seal the central runner groove (164); one end of the area air passage (167) is arranged at the edge of the sealing area of the waterproof breathable film (166), and the other end of the area air passage is communicated with the air containing area (165).

5. The cartridge of claim 4, wherein one amplification chamber (162) is provided with two zone flow paths (163), and the two zone flow paths (163) are an inlet zone flow path (1631) and an outlet zone flow path (1632); one end of the inlet flow channel (1631) is communicated with the central flow channel groove (164), and the other end of the inlet flow channel (1631) is communicated with one end of the amplification cavity (162); one end of the outlet flow channel (1632) is communicated with the other end of the amplification cavity (162), and the port of the other end of the outlet flow channel (1632) is positioned in the central cavity (161) and sealed by the waterproof breathable film (166).

6. The cartridge of claim 5, wherein the entrance flow path (1631) is provided with a first barrier (M1), and the exit flow path (1632) is provided with a second barrier (M2).

7. The cartridge of claim 5, wherein the amplification chambers (162) are four and are circumferentially arranged at equal intervals around the center of the central chamber (161).

8. The cartridge of claim 7, wherein one amplification chamber (162) and the corresponding entry flow channel (1631) and exit flow channel (1632) form a wing-shaped amplification unit, and four amplification units are symmetrically arranged to form a wing-shaped amplification unit group.

9. The cartridge having a nucleic acid molecule amplification structure according to claim 8, wherein the gas-containing regions (165) are provided in at least four and are respectively located between each two adjacent amplification units; a transition gas area (169) is also arranged between every two adjacent gas containing areas (165); the transition gas area (169) is communicated with the gas containing area (165) adjacent to the transition gas area through a gas distributing channel (168).

10. The cartridge of claim 9, wherein the gas-containing region (165) and the transition gas region (169) are hollow regions.

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