CN116814424A

CN116814424A - Double-chamber three-dimensional biochip and detection method thereof

Info

Publication number: CN116814424A
Application number: CN202211471336.7A
Authority: CN
Inventors: 请求不公布姓名
Original assignee: Jiangsu Aiweide Biotechnology Co ltd
Current assignee: Jiangsu Aiweide Biotechnology Co ltd
Priority date: 2022-09-30
Filing date: 2022-11-22
Publication date: 2023-09-29
Also published as: CN219546980U

Abstract

The invention relates to a double-chamber three-dimensional biochip, which comprises a first channel, a second channel, a fourth channel and a third channel which are sequentially arranged in parallel, wherein the first channel and the third channel are strip-shaped channels, the second channel comprises a first upper culture chamber and a second upper culture chamber which are mutually communicated, and the fourth channel comprises a first lower culture chamber and a second lower culture chamber which are mutually communicated; the chip comprises a connecting layer, an upper culture layer, a lower culture layer, a sealing layer and a circuit layer from top to bottom in sequence, wherein the connecting layer and the circuit layer are respectively positioned on the upper surface and the lower surface of the chip, and the upper culture layer, the lower culture layer and the sealing layer are sequentially positioned between the connecting layer and the circuit layer. The double-chamber three-dimensional biochip and the detection method can realize the serial connection or parallel connection of tissues at different functional parts of a three-dimensional organ.

Description

Double-chamber three-dimensional biochip and detection method thereof

Technical Field

The invention relates to the technical field of microfluidic chips, in particular to a double-chamber three-dimensional biochip and a detection method thereof.

Background

With the development of bioengineering and the improvement of application fields thereof, the concept of cell culture has been newly widened and developed, the interaction between complex structures of internal organs of human body is difficult to simulate by using a cell culture model such as an animal model/organoid chip of a simple two-dimensional model, and the response of two-dimensional cells to external stimuli is very different from the response of three dimensions in human body, and the three-dimensional cell culture has been receiving increasing attention due to the limited capability of simulating human infectious diseases by using a simple two-dimensional model. The three-dimensional cell culture refers to co-culturing different material carriers with three-dimensional structures and cells in vitro, so that the cells migrate and grow in the three-dimensional space structure of the carriers to form a three-dimensional cell-carrier complex, the three-dimensional cell culture can more effectively support biologically relevant experiments, and the limitation of in-vivo environment on cell culture is weakened.

In the related technology, a disease model is constructed by using a model biological mouse, the occurrence and development process of the disease cannot be intuitively monitored, the detection of the efficacy of the drug is insensitive, meanwhile, the existing drug off-target phenomenon still brings great uncertainty and challenges to the new drug development process, and the phenomenon that an animal experiment is effective and a human body inspection is ineffective exists for a long time. The culture system may be formed by injecting a fluid into the culture chamber to culture three-dimensional cells, but is often too simplified in terms of cell-cell and cell-matrix interactions due to its simple structural arrangement, which results in limited use in understanding organs of human complex structures.

Disclosure of Invention

In order to solve the problem that the culture system in the prior art is simple in structure and limited in application in understanding organs of human complex structures, the invention provides a double-chamber three-dimensional biochip and a detection method thereof.

The technical problem to be solved by the invention is to provide a double-chamber three-dimensional biochip, wherein the inside of the biochip comprises a first channel, a second channel, a fourth channel and a third channel which are sequentially arranged in parallel, the first channel and the third channel are strip-shaped channels, the second channel comprises a first upper culture chamber and a second upper culture chamber which are mutually communicated, and the fourth channel comprises a first lower culture chamber and a second lower culture chamber which are mutually communicated; the chip comprises a connecting layer, an upper culture layer, a lower culture layer, a sealing layer and a circuit layer from top to bottom in sequence, wherein the connecting layer and the circuit layer are respectively positioned on the upper surface and the lower surface of the chip, and the upper culture layer, the lower culture layer and the sealing layer are sequentially positioned between the connecting layer and the circuit layer; the first channel and the second channel are positioned on the upper culture layer, the first channel and the second channel are respectively bar-shaped holes penetrating through the upper surface and the lower surface of the upper culture layer, the third channel and the fourth channel are positioned on the lower culture layer, and the third channel and the fourth channel are respectively bar-shaped holes penetrating through the upper surface and the lower surface of the lower culture layer; the first thin film and the second thin film are arranged between the upper culture layer and the lower culture layer, the first thin film is positioned in a gap between the first upper culture chamber and the first lower culture chamber, and the second thin film is positioned in a gap between the second upper culture chamber and the second lower culture chamber.

Preferably, the side wall of the chip sequentially comprises a first side wall, a second side wall, a third side wall, a fourth side wall and a fifth side wall which are connected end to end, wherein the first side wall is opposite to the third side wall, the second side wall is opposite to the fifth side wall, the fourth side wall is a chamfer angle between the third side wall and the fifth side wall, and the first channel, the second channel, the third channel and the fourth channel are respectively parallel to the second side wall.

Preferably, the end part of the first upper culture chamber close to the first side wall is provided with two branched channels, namely a first runner and a second runner, the end part of the second upper culture chamber close to the third side wall is provided with two branched channels, namely a third runner and a fourth runner, the first runner extends along the direction towards the first side wall, the second runner extends firstly towards the second side wall and then extends towards the third side wall after bending, the third runner extends along the direction towards the third side wall, the fourth runner extends firstly towards the second side wall and then extends towards the first side wall after bending, the end part of the second runner is positioned on one side of the first upper culture chamber close to the second side wall, and the end part of the fourth runner is positioned on one side of the second upper culture chamber close to the second side wall; the middle of the second channel is provided with a first accommodating chamber perpendicular to the second channel, the first accommodating chamber is used for discharging electrodes and adding cells, and two sides of the first accommodating chamber are respectively provided with a first upper culture chamber and a second upper culture chamber.

Preferably, the end part of the first lower culture chamber close to the first side wall is provided with two branched channels, namely a fifth channel and a sixth channel, the end part of the second lower culture chamber close to the third side wall is provided with two branched channels, namely a seventh channel and an eighth channel, the fifth channel extends towards the first side wall, the sixth channel extends towards the fifth side wall firstly, then bends and extends towards the second side wall, the seventh channel extends towards the fifth side wall, the eighth channel extends towards the fifth side wall firstly, then bends and extends towards the fifth side wall, the end part of the sixth channel is positioned on one side of the first lower culture chamber close to the fifth side wall, and the end part of the eighth channel is positioned on one side of the second lower culture chamber close to the fifth side wall; the middle of the fourth channel is provided with a second accommodating chamber perpendicular to the fourth channel, the second accommodating chamber is used for discharging electrodes and adding cells, and the two sides of the second accommodating chamber are respectively provided with a first lower culture chamber and a second lower culture chamber.

Preferably, the connecting layer, the upper culture layer and the lower culture layer are respectively provided with a plurality of through holes, and the through holes are respectively correspondingly connected with the end parts of the first channel, the second channel, the third channel and the fourth channel.

Preferably, the position that the tie layer is close to the third lateral wall sets up first circulation group and second circulation group side by side, and first circulation group is close to the fifth lateral wall, and second circulation group is close to the second lateral wall, and first circulation group and second circulation group's structure is the same, and every circulation group sets up respectively to be two rows and two four through holes that arrange in a row, sets up connecting channel between two through holes, and every circulation group includes two connecting channel and is on a parallel with the second lateral wall.

Preferably, two bubble removing films are arranged on the connecting layer, and the two bubble removing films are respectively positioned on the first circulation group and the second circulation group, and the bubble removing films are porous polytetrafluoroethylene films and polydimethylsiloxane films.

Preferably, the first film and the second film are porous films.

Preferably, the chip can measure the transmembrane resistance, the circuit layer is provided with a first observation window, a second observation window and a transmembrane resistance measuring interface, and the connecting layer comprises a connecting hole of the transmembrane resistance measuring interface.

The invention provides a method for detecting a double-chamber three-dimensional biochip, which comprises the following steps: step S1: obtaining the growth of the tissue in the first upper culture chamber, the second upper culture chamber, the first lower culture chamber and the second lower culture chamber from the first observation window and the second observation window; step S2: the transmembrane resistance is measured by connecting the connecting hole of the transmembrane resistance measuring interface with the transmembrane resistance measuring instrument; step S3: and determining the state of the three-dimensional biological tissue model according to the growth condition and the transmembrane resistance of the tissue.

Compared with the prior art, the double-chamber three-dimensional biochip and the detection method have the following beneficial effects:

1. the culture chamber environment required by the three-dimensional organ with the complex multilayer structure can be provided structurally, so that the simulation degree of the constructed model is higher, and the function is closer to that of a real organ of a human body.

2. The three-dimensional organ chip structure constructed by the invention is convenient for observation and detection, and comprises a convenient microscopic observation mode and a transmembrane resistance measurement function.

3. The three-dimensional organ chip constructed by the invention can optimize the effect of bubble removal in a flow path.

Drawings

FIG. 1 is a schematic diagram showing the overall assembly structure of a dual-chamber three-dimensional biochip according to an embodiment of the invention;

FIG. 2 is a schematic diagram showing an exploded structure of a dual-chamber three-dimensional biochip according to an embodiment of the invention;

FIG. 3 is a schematic diagram showing the structure of an upper culture layer according to an embodiment of the present invention;

FIG. 4 is a schematic diagram showing the structure of a lower culture layer according to an embodiment of the present invention;

FIG. 5 is a schematic view of a structure of a connection layer according to an embodiment of the present invention;

FIG. 6 is a schematic diagram of a circuit board layer according to an embodiment of the invention;

FIG. 7 is a schematic view of a structure of a sealing layer according to an embodiment of the present invention;

FIG. 8 is a schematic diagram showing an exploded structure of a dual-chamber three-dimensional biochip according to another embodiment of the invention;

FIG. 9 is a schematic view of a connection layer according to another embodiment of the present invention;

FIG. 10 is a schematic diagram showing the structure of an upper culture layer according to another embodiment of the present invention;

FIG. 11 is a schematic view showing the structure of a lower culture layer according to another embodiment of the present invention;

FIG. 12 is a schematic diagram showing the overall assembly structure of a dual-chamber three-dimensional biochip according to another embodiment of the invention.

Reference numerals illustrate: 11-a first sidewall; 12-a second sidewall; 13-a third sidewall; 14-fourth side wall; 15-a fifth sidewall; a-bubble removal film; a1' -cap; a2' -bubble removal chamber; a21—a first bubble removal chamber; a22-a second bubble removal chamber; B. a B' -connecting layer; C. c' -upper culture layer; d-porous film, first film, second film; E. e' -lower culture layer; F. f' -blocking layer; a G-circuit layer;

b1-fluid inlet of upper culture layer; b2-fluid outlet of upper culture layer; b3-fluid outlet of lower culture layer; b4-fluid inlet of lower culture layer; 19-a first circulation group; 20-a second circulation group; 30-connecting channels; 191-a first through hole; 192-a second through hole; 193-third through hole; 194-fourth through holes; 195-a fifth through hole; 196-sixth through hole; 197-seventh through holes; 198-eighth through holes; b5-fifth aperture; b6-sixth aperture; b7-seventh aperture; b8-eighth aperture;

C1, C1' -first channel; c2, E2' -second channel; 16-a first accommodation chamber; c15, C15' -first upper culture chamber; c16, C16' -second upper culture chamber; 21-a first flow channel; 22-a second flow channel; 23-a third flow channel; 24-fourth flow channel; c3-a first via; c4-a second through hole; c6-a third through hole; c8_fourth through holes; c11_first aperture; c12_second aperture; c13-a third aperture; c14_fourth pores; c10-upper connection channel; c62, C64, C65, C67-wells on upper culture layer C';

e1, E3' -third channel; e2, E4' -fourth channel; 17-a second accommodation chamber; e7, E7' -a first lower culture chamber; e8, E8' -second lower culture chamber; 25-fifth flow channel; 26-sixth flow channel; 27-seventh flow channel; 28-eighth runner; e9-a fifth through hole; e10—seventh through hole; e11—sixth through hole; e6-lower connection channel; e31, E32, E33, E34-wells on lower culture layer E';

f1-a first electrode through hole; f2-a second electrode through hole; f3-a third electrode through hole; f4—fourth electrode via; f5—fifth electrode via; f6—sixth electrode via;

g1-a first electrode; g2—a second electrode; g3—a third electrode; g4—fourth electrode; g5—fifth electrode; g6—sixth electrode; g7-a first viewing window; g8—a second viewing window; 151-connection hole of transmembrane resistance measurement interface.

Detailed Description

The present application will be described in further detail with reference to the drawings and examples, in order to make the objects, technical solutions and advantages of the present application more apparent. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not intended to limit the scope of the application.

For the purpose of making the technical solution and advantages of the present application more apparent, the present application will be further described in detail below with reference to the accompanying drawings and examples of implementation. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not intended to limit the scope of the application.

Referring to fig. 1 and 2, the embodiment of the application provides a dual-chamber three-dimensional biochip 1, which can be used for drug screening detection, biological model construction, disease model construction, environmental assessment, and can be used for in vitro testing of replacement human bodies and living animals. The double-chamber three-dimensional biochip 1 is a plate-shaped box body which is approximately rectangular, and sequentially comprises a connecting layer B, an upper culture layer C, a lower culture layer E and a sealing layer F from top to bottom, wherein positioning holes are respectively formed in the upper and lower corresponding positions of each layer, the layers can be aligned through a jig, and the upper culture layer C and the lower culture layer E are fixed between the connecting layer B and the sealing layer F in a hot-press bonding, laser bonding, liquid glue or solid glue gluing, screw bonding and other modes. Fluids may be introduced into the upper and lower culture layers C and E for culturing cells and the like.

Furthermore, the dual-chamber three-dimensional biochip 1 may further include a circuit board layer G, a connection layer B, an upper culture layer C, a lower culture layer E, a sealing layer F, and positioning holes respectively disposed at the corresponding positions of each layer in the circuit board layer G, where each layer can be aligned by a jig, and the upper culture layer C, the lower culture layer E, and the sealing layer F are fixed between the connection layer B and the circuit board layer G by means of thermocompression bonding, laser bonding, liquid glue or solid glue, screws, and the like.

In some embodiments, the sidewalls of the dual-chamber three-dimensional biochip 1 may include a first sidewall 11, a second sidewall 12, a third sidewall 13, and a fifth sidewall 15 connected end to end in sequence, the first sidewall 11 and the third sidewall 13 being opposite, and the second sidewall 12 and the fifth sidewall 15 being opposite.

Further, the sidewalls of the dual-chamber three-dimensional biochip 1 may further comprise a fourth sidewall 14 in sequence, the fourth sidewall 14 being located at an angular position between the third sidewall 13 and the fifth sidewall 15 for forming a chamfer between the third sidewall 13 and the fifth sidewall 15.

And two bubble removing films A are arranged on the connecting layer B, wherein the bubble removing films A are porous polytetrafluoroethylene films, polydimethylsiloxane films and the like, gas can pass through the films, and liquid cannot pass through the films. After the bubble removal film A is combined with the flow channel on the double-chamber three-dimensional biochip 1, bubbles in the flow channel can be removed, and the influence of the bubbles flowing into the culture chamber on tissue culture is prevented. The circuit board layer G is used for integrating a transmembrane resistance measuring electrode, a measuring circuit and an interface.

Referring to fig. 3, a first channel C1 and a second channel C2 are disposed on the upper culture layer C, the first channel C1 and the second channel C2 are substantially parallel to each other, the first channel C1 is a fluid inflow channel of the upper culture layer C, the first channel C1 is a strip-shaped hole parallel to and close to the second sidewall 12 and penetrates through the upper and lower surfaces of the upper culture layer C, the first channel C1 is a strip-shaped channel, and the second channel C2 includes a first upper culture chamber C15 and a second upper culture chamber C16 that are mutually communicated.

In some embodiments, the second channel C2 is located between the first channel C1 and the fifth side wall 15, a first accommodating chamber 16 perpendicular to the second channel C2 is disposed in the middle of the second channel C2, the first accommodating chamber 16 may be used for adding cells, further, may be used for discharging electrodes, two sides of the first accommodating chamber 16 are respectively the first upper culture chamber C15 and the second upper culture chamber C16, two bifurcated channels are disposed at the end of the first upper culture chamber C15 near the first side wall 11, which are respectively the first flow channel 21 and the second flow channel 22, and the second flow channel 22 is an upper sample adding flow channel of the first culture unit near the inlet and the outlet. The end part of the second upper culture chamber C16, which is close to the third side wall 13, is provided with two branched channels, namely a third flow channel 23 and a fourth flow channel 24, and the fourth flow channel 24 is an upper sample adding flow channel of the second culture unit, which is far away from the inlet and the outlet. The first flow channel 21 extends along the direction towards the first side wall 11, the second flow channel 22 extends along the direction towards the second side wall 12, then bends and extends along the direction towards the third side wall 13, the third flow channel 23 extends along the direction towards the third side wall 13, the fourth flow channel 24 extends along the direction towards the second side wall 12, then bends and extends along the direction towards the first side wall 11, the end part of the second flow channel 22 is positioned on one side of the first upper culture chamber C15, which is close to the second side wall 12, and the end part of the fourth flow channel 24 is positioned on one side of the second upper culture chamber C16, which is close to the second side wall 12. The first accommodating chamber 16 includes an electrode accommodating chamber and a cell accommodating chamber which are communicated with each other, the electrode accommodating chamber is vertically connected to the middle position of the second channel C2, and the cell accommodating chamber is located at one end of the electrode accommodating chamber away from the second channel C2.

The upper culture layer C is provided with an upper connecting channel C10 at a position close to the third side wall 13, and the upper connecting channel C10 is parallel to and close to the strip-shaped holes of the third side wall 13 and penetrates through the upper surface and the lower surface of the upper culture layer C. The two ends of the upper connection channel C10 are respectively close to the ends of the first channel C1 and the second channel C2, and are respectively spaced from the ends of the first channel C1 and the second channel C2.

The upper culture layer C is provided with a first through hole C3 and a second through hole C4 at positions close to the first side wall 11, and the first through hole C3 is positioned between the second channel C2 and the second through hole C4. The middle position of the upper culture layer C is provided with a third through hole C6 and a fourth through hole C8 respectively, the third through hole C6 and the fourth through hole C8 are respectively positioned on one side of the second runner C2 close to the fifth side wall 15, the third through hole C6 is close to the first side wall 11, and the fourth through hole C8 is close to the third side wall 13. The third through hole C6 and the second flow passage 22 have ends respectively located at both sides of the second flow passage C2, and the fourth through hole C8 and the fourth flow passage 24 have ends respectively located at both sides of the second flow passage C2. The first through hole C3, the second through hole C4, the third through hole C6 and the fourth through hole C8 are through holes penetrating through the upper surface and the lower surface of the upper culture layer C. The first through hole C3 and the second through hole C4 are respectively through holes for connecting the fluid outlet B3 and the fluid inlet B4 of the lower culture layer, the third through hole C6 is a through hole for connecting the cell sample inlet of the lower layer of the first culture unit and the sample inlet of the lower layer of the first culture unit, and the fourth through hole C8 is a through hole for connecting the cell sample inlet of the lower layer of the second culture unit and the sample inlet of the lower layer of the second culture unit.

The side of the upper connecting channel C10 is provided with a first small hole C11, a second small hole C12, a third small hole C13 and a fourth small hole C14, the first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 are through holes penetrating through the upper surface and the lower surface of the upper culture layer C respectively, the first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 are arranged in two rows and two columns at intervals, the second small hole C12 and the third small hole C13 are close to the third side wall 13, the connecting line of the second small hole C12 and the third small hole C13 is parallel to the third side wall 13, the first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 are sequentially connected to form a rectangle, and the rectangle formed by connecting lines among the first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 is located between the upper connecting channel C10 and the fifth side wall 15. The third orifice C13 is located between the second orifice C12 and the upper connecting channel C10.

Referring to fig. 4, a third channel E1 and a fourth channel E2 are disposed on the lower culture layer E, the third channel E1 and the fourth channel E2 are substantially parallel to each other, the third channel E1 is a fluid inflow channel of the lower culture layer E, the third channel E1 is a strip-shaped hole parallel to and close to the fifth sidewall 15 and penetrates through the upper and lower surfaces of the upper culture layer E, the third channel E1 is a strip-shaped channel, and the fourth channel E4 includes a first lower culture chamber E7 and a second lower culture chamber E8 that are mutually communicated.

The fourth passageway E2 is located between third passageway E1 and the second lateral wall 12, and the centre of fourth passageway E2 sets up the second accommodation chamber 17 of perpendicular to fourth passageway E2, and second accommodation chamber 17 is used for discharging electrode and cell, and the both sides of second accommodation chamber 17 are respectively cultivate cavity E7 and second down and cultivate cavity E8 under the first, and the tip that second accommodation chamber 17 was kept away from to first cultivation cavity E7 sets up two bifurcation, is fifth runner 25 and sixth runner 26 respectively, sixth runner 26 is the lower floor of the first cultivation unit that is close to the exit and adds the appearance runner. The end of the second lower culture chamber E8 far away from the second accommodating chamber 17 is provided with two branched channels, namely a seventh flow channel 27 and an eighth flow channel 28, and the eighth flow channel 28 is a lower sample adding flow channel of the second culture unit far away from the inlet and the outlet. The fifth flow channel 25 extends towards the first side wall 11, the sixth flow channel 26 extends towards the fifth side wall 15, then bends and extends towards the third side wall 13, the seventh flow channel 27 extends towards the third side wall 13, the eighth flow channel 28 extends towards the fifth side wall 15, then bends and extends towards the first side wall 11, the end of the sixth flow channel 26 is located at one side of the first lower culture chamber E7 close to the fifth side wall 15, and the end of the eighth flow channel 28 is located at one side of the second lower culture chamber E8 close to the fifth side wall 15. The second accommodating chamber 17 includes an electrode accommodating chamber and a cell accommodating chamber which are communicated with each other, the electrode accommodating chamber is vertically connected with the intermediate position of the fourth channel E2, and the cell accommodating chamber is located at one end of the electrode accommodating chamber away from the fourth channel E2.

The lower culture layer E is provided with a lower connecting channel E6 at a position close to the third side wall 13, and the lower connecting channel E6 is parallel to and close to the strip-shaped holes of the third side wall 13 and penetrates through the upper surface and the lower surface of the lower culture layer E. The two ends of the lower connection channel E6 are respectively close to the ends of the third channel E1 and the fourth channel E2, and are respectively spaced from the ends of the third channel E1 and the fourth channel E2.

The middle position of the lower culture layer E is respectively provided with a fifth through hole E9 and a sixth through hole E11, the fifth through hole E9 and the sixth through hole E11 are respectively positioned at one side of the fourth runner E2, which is close to the second side wall 12, the fifth through hole E9 is close to the first lower culture chamber E7, and the sixth through hole E11 is close to the second lower culture chamber E8. The ends of the fifth through hole E9 and the sixth flow channel 26 are respectively located at two opposite sides of the fourth flow channel E2, and the ends of the sixth through hole E11 and the eighth flow channel 28 are respectively located at two opposite sides of the fourth flow channel E2. The fifth through hole E9 and the sixth through hole E11 are through holes penetrating the upper and lower surfaces of the lower culture layer E, respectively. The lower culture layer E is further provided with a seventh through hole E10, and the seventh through hole E10 and the second accommodating chamber 17 are respectively positioned on two opposite sides of the fourth flow channel E2.

When the upper and lower culture layers C and E are attached to each other, the first, second, third and fourth channels C1, C2, E1 and E2 are sequentially positioned in parallel between the second and fifth side walls 12 and 15. The ends of the third and fourth channels E1 and E2 near the first side wall 11 correspond up and down to the positions of the second and first through holes C4 and C3, respectively, and the two ends of the lower connecting channel E6 correspond up and down to the positions of the second and third small holes C12 and C13 near the third side wall 13, respectively.

Referring to fig. 2, two porous membranes D are disposed between the upper and lower culture layers C and E, when the upper and lower culture layers C and E are adhered to each other, a first gap is formed between a first upper culture chamber C15 of the upper culture layer C and a first lower culture chamber E7 of the lower culture layer E, a second gap is formed between a second upper culture chamber C16 of the upper culture layer C and a second lower culture chamber E8 of the lower culture layer E, the two porous membranes D are respectively disposed at positions of the first gap and the second gap between the upper and lower culture layers C and E for separating the upper and lower culture chambers, the porous membrane D adjacent to the first sidewall 11 is used for separating the first upper culture chamber C15 and the first lower culture chamber E7, the porous membrane D adjacent to the third sidewall 13 is used for separating the second upper culture chamber C16 and the second lower culture chamber E8, and at the same time, the upper and lower chamber materials can be exchanged through the porous membrane D. As a modification, the porous membrane D may be another membrane capable of exchanging a substance.

Referring to fig. 5, the connection layer B is disposed at a position close to the first side wall 11 at intervals in sequence with a fluid inlet B1 of the upper culture layer C, a fluid outlet B2 of the upper culture layer C, a fluid outlet B3 of the lower culture layer E, and a fluid inlet B4 of the lower culture layer E, wherein the fluid inlet B1 of the upper culture layer C, the fluid outlet B2 of the upper culture layer C, the fluid outlet B3 of the lower culture layer E, and the fluid inlet B4 of the lower culture layer E are through holes penetrating through the upper and lower surfaces of the connection layer B, and the arrangement direction is parallel to the first side wall 11.

The connecting layer B is disposed adjacent to the third sidewall 13 in parallel with the first circulation group 19 and the second circulation group 20, where the first circulation group 19 is adjacent to the fifth sidewall 15, the second circulation group 20 is adjacent to the second sidewall 12, and the first circulation group 19 and the second circulation group 20 have the same structure. The first flow group 19 includes a first through hole 191, a second through hole 192, a third through hole 193, and a fourth through hole 194, the first through hole 191, the second through hole 192, the third through hole 193, and the fourth through hole 194 are arranged in two rows and two columns at intervals, the first through hole 191, the second through hole 192, the third through hole 193, and the fourth through hole 194 are sequentially connected to form a rectangle, the first through hole 191 and the second through hole 192 are close to the third side wall 13, a connection line of the first through hole 191 and the second through hole 192 is parallel to the third side wall 13, and the first through hole 191 is located between the second through hole 192 and the fifth side wall 15. The first flow-through group 19 is a through hole and a flow passage of a lower culture layer E bubble removal structure, the second flow-through group 20 is a through hole and a flow passage of an upper culture layer C bubble removal structure, the upper connecting channel C10 is a connecting flow passage of the first channel C1, the second channel C2 and the second flow-through group 20, and the lower connecting channel E6 is a connecting flow passage of the third flow passage E1, the fourth flow passage E2 and the first flow-through group 19. The two bubble removal films a arranged on the connecting layer B are respectively positioned on the first circulation group 19 and the second circulation group 20, and the shapes of the first circulation group 19 and the second circulation group 20 are respectively approximate rectangles.

The second through-hole group 20 includes a fifth through-hole 195, a sixth through-hole 196, a seventh through-hole 197, and an eighth through-hole 198, the fifth through-hole 195, the sixth through-hole 196, the seventh through-hole 197, and the eighth through-hole 198 are arranged in two rows and two columns at intervals, the fifth through-hole 195, the sixth through-hole 196, the seventh through-hole 197, and the eighth through-hole 198 are sequentially connected to form a rectangle, the fifth through-hole 195 and the sixth through-hole 196 are close to the third sidewall 13, the connection line of the fifth through-hole 195 and the sixth through-hole 196 is parallel to the third sidewall 13, the fifth through-hole 195 is located between the first through-hole group 19 and the sixth through-hole 196, and the sixth through-hole 196 is located between the fifth through-hole 195 and the second sidewall 12. The positions of the four through holes of the first circulation group 19 respectively correspond to the first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 of the upper culture layer C up and down, the first circulation group 19 is provided with two parallel connecting channels 30, the second circulation group 20 is provided with two parallel connecting channels 30, each connecting channel 30 is positioned between the two through holes to communicate the two connected through holes, and the four connecting channels 30 are respectively parallel to the second side wall 12. The first small hole C11, the second small hole C12, the third small hole C13 and the fourth small hole C14 are through holes of the bubble removing structure of the lower culture layer E.

The middle of the connecting layer B is provided with a fifth small hole B5, a sixth small hole B6, a seventh small hole B7 and an eighth small hole B8, the fifth small hole B5, the sixth small hole B6, the seventh small hole B7 and the eighth small hole B8 are through holes penetrating through the upper surface and the lower surface of the connecting layer B respectively, and the four small holes form a rectangle. The fifth aperture B5 is adjacent to the first and second side walls 11, 12, the sixth aperture B6 is adjacent to the first and fifth side walls 11, 15, the seventh aperture B7 is adjacent to the second and third side walls 12, 13, and the eighth aperture B8 is adjacent to the third and fifth side walls 13, 15. The fifth small hole B5 and the sixth small hole B6 are cell sample adding ports of the first culture unit close to the inlet and outlet (B1-B4), B5 is a cell sample adding port of the upper culture chamber C15 of the first culture unit, and B6 is a cell sample adding port of the lower culture chamber E7 of the first culture unit. The seventh small hole B7 and the eighth small hole B8 are cell sample adding ports of the second culture unit far away from the inlet and the outlet, B7 is a cell sample adding port of the upper culture chamber C16 of the second culture unit, and B8 is a cell sample adding port of the lower culture chamber E8 of the second culture unit.

When the connection layer B, the upper culture layer C and the lower culture layer E are sequentially attached, the fluid inlet B1 of the upper culture layer C and the end portion of the first channel C1 close to the first side wall 11 are vertically corresponding, the fluid outlet B2 of the upper culture layer C and the end portion of the second channel C2 close to the first side wall 11 are vertically corresponding, the fluid outlet B3 of the lower culture layer E and the end portion of the first through hole C3 of the upper culture layer C and the end portion of the fourth channel E2 close to the first side wall 11 are vertically corresponding, the fluid inlet B4 of the lower culture layer E and the second through hole C4 of the upper culture layer C and the end portion of the third channel E1 close to the first side wall 11 are vertically corresponding, the fifth small hole B5 of the connection layer B and the end portion of the second channel 22 of the upper culture layer C are vertically corresponding, the sixth small hole B6 of the connection layer B and the third through hole C6 of the upper culture layer C and the end portion of the sixth channel 26 are vertically corresponding, the seventh small hole B7 of the connection layer B and the end portion of the fourth channel 24 of the upper layer C are vertically corresponding, and the eighth small hole B8 of the eighth layer B and the fourth channel C8 of the eighth layer C are vertically corresponding.

The first through hole 191 of the connection layer B corresponds up and down to the second small hole C12 of the upper culture layer C and the end of the lower connection channel E6 of the lower culture layer E near the fifth side wall 15, the second through hole 192 of the connection layer B corresponds up and down to the third small hole C13 of the upper culture layer C and the end of the lower connection channel E6 of the lower culture layer E far from the fifth side wall 15, the third through hole 193 of the connection layer B corresponds up and down to the fourth small hole C14 of the upper culture layer C and the end of the seventh flow channel 27 of the lower culture layer E, and the fourth through hole 194 of the connection layer B corresponds up and down to the end of the first small hole C11 of the upper culture layer C and the end of the third channel E1 near the third side wall 13.

The fifth through hole 195 of the connection layer B corresponds up and down to an end of the upper connection channel C10 of the upper culture layer C, which is far from the second side wall 12, the sixth through hole 196 of the connection layer B corresponds up and down to an end of the upper connection channel C10 of the upper culture layer C, which is close to the second side wall 12, the seventh through hole 197 of the connection layer B corresponds up and down to an end of the first channel C1 of the upper culture layer C, which is close to the third side wall 13, and the eighth through hole 198 of the connection layer B corresponds up and down to an end of the third flow channel 23 of the upper culture layer C.

Referring to fig. 6, a first observation window G7 and a second observation window G8 are disposed in parallel in the middle of the circuit board layer G, the first observation window G7 is close to the first sidewall 11, and the second observation window G8 is close to the third sidewall 13. The first observation window G7 and the second observation window G8 are through holes penetrating through the circuit board layer G respectively, a first measurement electrode G1 and a second measurement electrode G2 are arranged on one side, far away from the second observation window G8, of the first observation window G7, a third measurement electrode G3 and a fourth measurement electrode G4 are arranged between the first observation window G7 and the second observation window G8, a fifth measurement electrode G5 and a sixth measurement electrode G6 are arranged on one side, far away from the first observation window G7, of the second observation window G8, and the first measurement electrode G1, the second measurement electrode G2, the third measurement electrode G3, the fourth measurement electrode G4, the fifth measurement electrode G5 and the sixth measurement electrode G6 are perpendicular to the upper surface of the circuit board layer G respectively. The middle position of the circuit board layer G, which is close to the fifth side wall 15, is provided with a transmembrane resistance measuring interface G9, the transmembrane resistance measuring interface G9 is perpendicular to the upper surface of the circuit board layer G, and the circuit board layer G is connected with an external transmembrane resistance measuring instrument through a USB interface. The materials of the first measurement electrode G1, the second measurement electrode G2, the third measurement electrode G3, the fourth measurement electrode G4, the fifth measurement electrode G5 and the sixth measurement electrode G6 may be platinum, gold, graphite, etc.

Referring to fig. 7, three sets of electrode through holes are respectively disposed in the middle of the sealing layer F, namely, a first electrode through hole F1, a second electrode through hole F2, a third electrode through hole F3, a fourth electrode through hole F4, a fifth electrode through hole F5 and a sixth electrode through hole F6, wherein the fourth electrode through hole F4 and the first electrode through hole F1 are close to the first sidewall 11, the fifth electrode through hole F5 and the second electrode through hole F2 are located in the middle of the sealing layer F, and the sixth electrode through hole F6 and the third electrode through hole F3 are close to the third sidewall 13.

When the sealing layer F is bonded to the circuit layer G, the first measuring electrode G1 penetrates through the fourth electrode through hole F4, the second measuring electrode G2 penetrates through the first electrode through hole F1, the third measuring electrode G3 penetrates through the fifth electrode through hole F5, the fourth measuring electrode G4 penetrates through the second electrode through hole F2, the fifth measuring electrode G5 penetrates through the sixth electrode through hole F6, and the sixth measuring electrode G6 penetrates through the third electrode through hole F3. The second measuring electrode G2, the third measuring electrode G3 and the sixth measuring electrode G6 are respectively contacted with the fluid of the lower culture layer E through the first electrode through hole F1, the fifth electrode through hole F5, the third electrode through hole F3; the first, fourth and fifth measuring electrodes G1, G4, G5 are in fluid contact with the upper culture layer C through the fourth, second and sixth electrode through holes F4, F2, F6, respectively, and through the fifth, seventh and sixth through holes E9, E10, E11, respectively.

The connecting layer B, the upper culture layer C, the lower culture layer E, the sealing layer F and the upper and lower corresponding positions of the transmembrane resistance measuring interface G9 on the circuit layer G are respectively provided with a strip-shaped through hole, the transmembrane resistance measuring interface G9 sequentially penetrates through the strip-shaped through holes of each layer, and a connecting hole 151 of the transmembrane resistance measuring interface G9 is formed on the upper surface of the connecting layer B.

As shown in fig. 8-12, the present application additionally provides an embodiment of a dual-chamber three-dimensional biochip.

Specifically, the dual-chamber three-dimensional biochip may include a connection layer B ', an upper culture layer C', a lower culture layer E ', and a sealing layer F' in this order from the top down. The corresponding positions on each layer are respectively provided with a positioning hole, each layer can be aligned by a jig, and the upper culture layer C 'and the lower culture layer E' are fixed between the connecting layer B 'and the sealing layer F' by means of hot press bonding, laser bonding, liquid glue or solid glue bonding, screws and the like. Fluids may be introduced into the upper and lower culture layers C 'and E', for culturing cells, etc. The connection layer B' may include a connection male, on which docking holes B1, B2, B3, B4 are provided.

Specifically, the first channel C1' is provided in the upper culture layer C ', and the first channel C1' may include a first upper culture chamber C15' and a second upper culture chamber C16' which are communicated with each other. The lower culture layer E 'is provided with a second channel E2', a third channel E3 'and a fourth channel E4'. The first channel C1', the second channel E2', the third channel E3 'and the fourth channel E4' are approximately parallel two by two. The third channel E3' may include a first lower culture chamber E7' and a second lower culture chamber E8' communicating with each other. The butt joint holes on the connecting layer B ' can be correspondingly communicated with the holes of the channels of the upper culture layer C ' and the lower culture layer E '. The wells C62, C64, C65, C67 on the upper culture layer C ' are respectively communicated with the wells on the corresponding positions on the connecting layer B ', and the wells C62, C64, C65, C67 can be used for adding the sample to the corresponding positions in the upper culture layer C '. The holes E31, E32, E33, E34 on the lower culture layer E ' are respectively communicated with the holes on the corresponding positions on the connecting layer B ' through the upper culture layer C ', and the holes E31, E32, E33, E34 on the lower culture layer E ' can be used for adding samples to the corresponding positions on the lower culture layer E '.

Optionally, the connection layer B ' may further be provided with a bubble removing chamber A2', and the bubble removing chamber A2' may include a first bubble removing chamber a21 and a second bubble removing chamber a22. The bubble removal chamber lid A1' may seal the bubble removal chamber A2' to ensure that fluid enters and exits the first bubble removal chamber a21 through the aperture in the bottom end of the bubble removal chamber A2 '. Two communication ports communicating with the second passage E2', the third passage E3' are provided at the bottom end of the first bubble removal chamber a21, and these two communication ports are at diagonal positions of the first bubble removal chamber a21. The second bubble removal chamber a22 communicates with the first passage C1', the fourth passage E4', respectively. Two communication ports, through which the second bubble removal chamber a22 communicates with the first channel C1', the fourth channel E4', are provided at the bottom end of the second bubble removal chamber a22, and are located at diagonal positions of the second bubble removal chamber a22. By this arrangement, bubbles can rise to the surface of the bubble removal chamber during passage through the bubble removal chamber, and bubbles in the fluid can be greatly reduced by the bubble removal chamber.

The bubble removing chamber can remove bubbles in the flow path, and prevent the bubbles from flowing into the culture chamber to affect tissue culture. For ease of understanding, fig. 12 shows a perspective view of one embodiment of the chip.

The invention relates to a detection method of a double-chamber three-dimensional biochip, which comprises the following steps:

step S1: acquiring the growth conditions of tissues in the first upper culture chamber C15, the second upper culture chamber C16, the first lower culture chamber E7 and the second lower culture chamber E8 from the first observation window G7 and the second observation window G8;

step S2: the transmembrane resistance is measured by connecting the connecting hole 151 of the transmembrane resistance measuring interface G9 with a transmembrane resistance measuring instrument;

step S3: and determining the state of the three-dimensional biological tissue model according to the growth condition and the transmembrane resistance of the tissue.

The invention provides a double-chamber three-dimensional biochip, which can be used for improving development of an external micro-physiological system of a human body, and adopts a micro-fluidic technology (the micro-fluidic technology is a technology capable of controlling or detecting fluid in a micrometer scale, and has the capability of miniaturizing basic functions of a laboratory such as biology, chemistry and the like to a few square centimeters, so that basic operations such as sample preparation, reaction, separation, detection and the like in a biochemical analysis process can be automatically completed), and important structural parts of organs, namely structures such as a culture chamber arranged in the three-dimensional biological tissue chip, a fluid inlet channel, a fluid outlet channel and the like which are communicated with the culture chamber, can effectively simulate functional structural units of the three-dimensional biological tissue and a growing microenvironment, can realize more accurate control in time and space dimensions, further saves labor cost, and realizes high-throughput, large-scale and standardized detection. In addition, the organ cells are cultured on the membrane simultaneously, so that three-dimensional biological tissue culture of various cells can be realized, and the introduction of biological materials can better simulate the interaction between the cells and the matrix, thereby being an ideal in-vitro research model for scientific research and clinical detection. The formed three-dimensional biological tissue chip can obtain a corresponding disease model after being subjected to certain physical, chemical or biological treatment, and can carry out deeper disease research or drug development research.

The above description is only of the preferred embodiments of the present invention and is not intended to limit the invention, but any modifications, equivalents, improvements, etc. within the principles of the present invention should be included in the scope of the present invention.

Claims

1. A dual-chamber three-dimensional biochip, characterized in that: the chip comprises a connecting layer (B, B '), an upper culture layer (C, C'), a lower culture layer (E, E ') and a sealing layer (F, F') which are sequentially arranged; the upper culture (C, C ') layer has a first upper culture chamber (C15, C15 ') and a second upper culture chamber (C16, C16 ') which are in communication with each other, the lower culture layer (E, E ') has a first lower culture chamber (E7, E7 ') and a second lower culture chamber (E8, E8 ') which are in communication with each other, a first membrane (D, D ') and a second membrane (D, D ') are provided between the upper culture layer (C, C ') and the lower culture layer (E, E '), the first membrane (D, D ') is located in a gap between the first upper culture chamber (C15, C15 ') and the first lower culture chamber (E7, E7 '), the second membrane (D, D ') is located in a gap between the second upper culture chamber (C16, C16 ') and the second lower culture chamber (E8, E8 '), the first upper culture chamber (C15, C15 ') and the first lower culture chamber (E7), the second upper culture chamber (E8 ') and the second upper culture chamber (E8, E8 ') constitute a first upper culture unit (C7, C8 ') and the second culture chamber (E8, E8 ').

2. A dual-chamber three-dimensional biochip according to claim 1, wherein: the chip comprises the following components:

a first channel (C1 ') provided to the upper culture layer (C'), the first channel comprising the first upper culture chamber (C15 ') and the second upper culture chamber (C16');

a fourth channel (E4 ') provided on the lower culture pass-through layer (E ') and communicating with the first channel (C1 ');

a second channel (E2 ') arranged in the lower culture layer (E');

a third channel (E3 ') provided in the lower culture layer (E') and communicating with the second channel (E2 '), the third channel (E3') comprising the first lower culture chamber (E7 ') and the second lower culture chamber E8').

3. A dual-chamber three-dimensional biochip according to claim 2, wherein: the connection layer (B') further comprises a first de-bubbling chamber (a 21) and a second de-bubbling chamber (a 22);

the first bubble removing chamber (A21) is communicated with the second channel (E2 ') and the third channel (E3'), two communication ports for communicating the first bubble removing chamber (A21) with the second channel (E2 ') and the third channel (E3') are arranged at the bottom end of the first bubble removing chamber (A21), and the two communication ports are arranged at the diagonal positions of the first bubble removing chamber (A21);

The second bubble removing chamber (A22) is respectively communicated with the first channel (C1 ') and the fourth channel (E4'), and two communication ports for communicating the second bubble removing chamber (A22) with the first channel (C1 ') and the fourth channel (E4') are arranged at the bottom end of the second bubble removing chamber (A22), and the two communication ports are arranged at the diagonal positions of the second bubble removing chamber (A22).

4. A dual-chamber three-dimensional biochip according to claim 1, wherein: the chip comprises the following components:

a first channel (C1) provided in the upper culture layer (C);

a second channel (C2) provided to the upper culture layer (C) and communicating with the first channel (C1), the second channel (C2) including the first upper culture chamber (C15) and the second upper culture chamber (C16);

a third channel (E1) provided in the lower culture pass-through layer (E);

a fourth channel (E2) disposed in the lower culture layer (E) and in communication with the third channel (E1), the fourth channel (E2) comprising the first lower culture chamber (E7) and the second lower culture chamber (E8).

5. A dual-chamber three-dimensional biochip according to claim 4, wherein: the side wall of the chip sequentially comprises a first side wall (11), a second side wall (12), a third side wall (13), a fourth side wall (14) and a fifth side wall (15) which are connected end to end, wherein the first side wall (11) is opposite to the third side wall (13), the second side wall (12) is opposite to the fifth side wall (15), and the fourth side wall (14) is a chamfer angle between the third side wall (13) and the fifth side wall (15);

The position of the connecting layer (B) close to the first side wall is sequentially provided with a fluid inlet (B1) of the upper culture layer (C), a fluid outlet (B2) of the upper culture layer, a fluid outlet (B3) of the lower culture layer (E) and a fluid inlet (B4) of the lower culture layer (E) at intervals;

the fluid inlet (B1) of the upper culture layer (C) is communicated with the end part, close to the first side wall (11), of the first channel (C1), the fluid outlet (B2) of the upper culture layer (C) is communicated with the end part, close to the first side wall (11), of the second channel (C2), the fluid inlet (B4) of the lower culture layer (E) is communicated with the end part, close to the first side wall (11), of the third channel (E1), and the fluid outlet (B3) of the lower culture layer (E) is communicated with the end part, close to the first side wall (11), of the fourth channel (E2).

6. A dual-chamber three-dimensional biochip according to claim 5, wherein: the connection layer (B) further comprises:

a first circulation group (19), wherein the first circulation group (19) comprises a first through hole (191), a second through hole (192), a third through hole (193), a fourth through hole (194) and two connecting channels (30) parallel to the second side wall (12), the first through hole (191), the second through hole (192), the third through hole (193) and the fourth through hole (194) are arranged in two rows and two columns, one connecting channel (30) is communicated with the first through hole (191) and the fourth through hole (194), the other connecting channel (30) is communicated with the second through hole (192) and the third through hole (193), the upper culture layer (C) is further provided with a second small hole (C12), a third small hole (C13), a fourth small hole (C14) and a first small hole (C11) which are respectively communicated with the first through hole (191), the second through hole (192), the third through hole (193) and the fourth through hole (194), the lower culture layer (E) is further provided with a lower connecting channel (E6), two ends of the lower connecting channel (E6) are respectively communicated with the second small hole (C12) and the third small hole (C13), and a fluid inlet (B4) of the lower culture layer (E), the third channel (E1) is close to one end of the first side wall (11), the third channel (E1) is close to one end of the third side wall (13), the first small hole (C11), the fourth through hole (194), one of the connecting channels (30) of the first flow-through group (19), the first through hole (191), the second small hole (C12), the lower connecting channel (E6), the third small hole (C13), the second through hole (192), the other connecting channel (30) of the first flow-through group (19), the third through hole (193), the fourth small hole (C14), the fourth channel (E2) is close to one end of the third side wall (13), the fourth channel (E2) is close to one end of the first side wall (11), and the fluid outlet (B3) of the lower culture layer (E) are sequentially communicated;

A second flow-through group (20), wherein the second flow-through group (20) comprises a fifth through hole (195), a sixth through hole (196), a seventh through hole (197), an eighth through hole (198) and two connecting channels (30) parallel to the second side wall (12), the fifth through hole (195), the sixth through hole (196), the seventh through hole (197) and the eighth through hole (198) are arranged in two rows and two columns, one connecting channel (30) is communicated with the fifth through hole (195) and the eighth through hole (198), the other connecting channel (30) is communicated with the sixth through hole (196) and the seventh through hole (197), the upper culture layer (C) is further provided with an upper connecting channel (C10), two ends of the upper connecting channel (C10) are respectively communicated with the fifth through hole (195) and the sixth through hole (196), a fluid inlet (B1) of the upper culture layer (C) is close to one end of the first side wall (11) in the first channel (C1), one end of the third side wall (13) in the first channel (C1) is close to one end of the third side wall (13) in the seventh through hole (197) in the second through hole (196), one connecting channel (30) of the second through group (20), the sixth through hole (196) in the first through hole (C10) in the first through hole (C1), the fifth through hole (195), the other connecting channel (30) of the second flow-through group (20), and the eighth through hole (198), wherein one end of the second channel (C2) close to the third side wall (13), one end of the second channel (C2) close to the first side wall (11), and a fluid outlet (B2) of the upper culture layer (C) are sequentially communicated;

Wherein, two bubble removing films (A) are arranged on the connecting layer (B), and the two bubble removing films (A) are respectively positioned in a first circulation group (19) and a second circulation group (20).

7. A dual-chamber three-dimensional biochip according to claim 4, wherein: a first accommodating chamber (16) perpendicular to the second channel (C2) is arranged in the middle of the second channel (C2), and the two sides of the first accommodating chamber (16) are respectively provided with the first upper culture chamber (C15) and the second upper culture chamber (C16); the middle of the fourth channel (E2) is provided with a second accommodating chamber (17) perpendicular to the fourth channel (E2), and two sides of the second accommodating chamber (17) are respectively provided with a first lower culture chamber (E7) and a second lower culture chamber (E8).

8. A dual-chamber three-dimensional biochip according to claim 6, wherein: a fifth small hole (B5), a sixth small hole (B6), a seventh small hole (B7) and an eighth small hole (B8) are arranged in the middle of the connecting layer (B);

the end part of the first upper culture chamber (C15) close to the first side wall (11) is provided with a first branched flow passage (21) and a second branched flow passage (22), the first flow passage (21) extends along the direction towards the first side wall (11) and is communicated with a fluid outlet (B2) of the upper culture layer (C), and the second flow passage (22) extends towards the second side wall (12) at first, extends towards the third side wall (13) after being bent and is communicated with the fifth small hole (B5); the end part of the second upper culture chamber (C16) close to the third side wall (13) is provided with a third flow passage (23) and a fourth flow passage (24) which are branched, the third flow passage (23) extends along the direction towards the third side wall (13) and is communicated with the eighth through hole (198), and the fourth flow passage (24) extends along the direction towards the second side wall (12) first, then extends towards the first side wall (11) after being bent and is communicated with the seventh small hole (B7);

The end part of the first lower culture chamber (E7) close to the first side wall (11) is provided with a branched fifth flow passage (25) and a sixth flow passage (26), the fifth flow passage (25) extends along the direction towards the first side wall (11) and is communicated with a fluid outlet (B3) of the lower culture layer (E), and the sixth flow passage (26) extends towards the direction of the fifth side wall (15) first, extends towards the direction of the third side wall (13) after being bent and is communicated with the sixth small hole (B6); the end part of the second lower culture chamber (E8) close to the third side wall (13) is provided with a forked seventh flow passage (27) and an eighth flow passage (28), the seventh flow passage (27) extends along the direction towards the third side wall (13) and is communicated with the fourth small hole (C14), the eighth flow passage (28) extends towards the direction of the fifth side wall (15) first, and then extends towards the direction of the first side wall (11) and is communicated with the eighth small hole (B8) after bending.

9. A dual-chamber three-dimensional biochip according to claim 6, wherein: the chip further comprises a circuit board layer (G), wherein the circuit board layer (G) is arranged on one side, away from the lower culture layer (E), of the sealing layer (F), and a first observation window (G7) and a second observation window (G8) are arranged in parallel in the middle of the circuit board layer (G) and are used for observing the growth conditions of tissues in the first lower culture chamber (E7) and the second lower culture chamber (E8) respectively;

A first measuring electrode (G1) and a second measuring electrode (G2) are arranged on one side, far away from the second observing window (G8), of the first observing window (G7), a third measuring electrode (G3) and a fourth measuring electrode (G4) are arranged between the first observing window (G7) and the second observing window (G8), a fifth measuring electrode (G5) and a sixth measuring electrode (G6) are arranged on one side, far away from the first observing window (G7), of the second observing window (G8), and the first measuring electrode (G1), the second measuring electrode (G2), the third measuring electrode (G3), the fourth measuring electrode (G4), the fifth measuring electrode (G5) and the sixth measuring electrode (G6) are respectively perpendicular to the upper surface of the circuit board layer (G); the first measuring electrode (G1), the fourth measuring electrode (G4) and the fifth measuring electrode (G5) all pass through the closing layer F and the lower culture layer E and penetrate into the second flow channel (22), the first receiving chamber (16) and the fourth flow channel (24), respectively, and the second measuring electrode (G2), the third measuring electrode (G3) and the sixth measuring electrode (G6) all pass through the closing layer (F) and penetrate into the sixth flow channel (26), the second receiving chamber (17) and the eighth flow channel (28), respectively;

And a transmembrane resistance measuring interface (G9) is arranged at the middle position of the circuit board layer (G) close to the fifth side wall (15), and the transmembrane resistance measuring interface (G9) is perpendicular to the upper surface of the circuit board layer (G) and is used for being connected with an external transmembrane resistance measuring instrument.

10. A detection method of a double-chamber three-dimensional biochip is characterized in that: the detection of a chip according to any one of claims 1-9, comprising the steps of:

step S1: obtaining the growth of the tissue in the first upper culture chamber (C15, C15 '), the second upper culture chamber (C16, C16'), the first lower culture chamber (E7, E7 ') and the second lower culture chamber (E8, E8');

step S2: connecting a chip transmembrane resistance measuring instrument to measure transmembrane resistance;

step S3: and determining the state of the three-dimensional biological tissue model according to the growth condition of the tissue and the transmembrane resistance.