CN110684662B - Full liquid phase organ chip and preparation method thereof - Google Patents

Full liquid phase organ chip and preparation method thereof Download PDF

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CN110684662B
CN110684662B CN201910972242.XA CN201910972242A CN110684662B CN 110684662 B CN110684662 B CN 110684662B CN 201910972242 A CN201910972242 A CN 201910972242A CN 110684662 B CN110684662 B CN 110684662B
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顾忠泽
杜鑫
李玉雯
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Southeast University
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Abstract

The invention discloses a full liquid phase organ chip and a preparation method thereof, wherein the full liquid phase organ chip comprises a super-hydrophobic porous membrane with a hydrophilic channel, the porous membrane is sealed in an oil phase, and the hydrophilic channel is provided with a fluid inlet and a fluid outlet which are communicated with the hydrophilic channel; placing the polymer layer porous membrane in a functional modification solution to obtain a hydrophobic polymer layer porous membrane; performing hydrophilic pattern treatment on the hydrophobic polymer layer porous membrane; then carrying out polymer surface super-hydrophobic treatment on the surface of the porous membrane with the hydrophilic channel; and then wetting the hydrophilic channel on the surface of the porous membrane, sealing the hydrophilic channel in the oil phase, and arranging an inlet and an outlet which are communicated with the hydrophilic channel. The invention has mild preparation conditions, can construct cell patterning culture on the prepared substrate to realize the construction of the full liquid phase organ chip, can realize in-situ detection and sampling of the microenvironment in each part of the organ chip, and can realize the spatial editing of the function and the geometric layout of the fluid device according to the requirements.

Description

Full liquid phase organ chip and preparation method thereof
Technical Field
The invention relates to the field of biochips and preparation methods thereof, in particular to a full-liquid-phase organ chip and a preparation method thereof.
Background
The research and development of new medicines are industries which are long in time consumption and large in investment. The average development cost of a new drug at present is about 10 hundred million dollars, and the average time is 10 years, so that the development of the new drug becomes extremely difficult. At present, 90% of drugs passing through cell experiments and animal experiments in drug screening are rejected in clinical test stages, and the reason for the rejection is mainly because the cell experiments and animal models are greatly different from human models, so that the development of a drug test platform closer to the human models is very important. In recent years, with the rapid development of biotechnology and information technology, organ chips have become an emerging field of great features and vitality in biomedicine. An organ chip is a bionic system which can simulate the main functions of human organs and a circulatory system on a micro-fluidic chip by utilizing a micro-processing technology. Besides the characteristics of miniaturization, integration and low consumption of the microfluidic technology, the organ chip technology can accurately control a plurality of system parameters, thereby simulating the complex structure, microenvironment and physiological functions of human organs. Compared with the traditional plane cell culture and animal models, the organoid has the advantages of more approaching the structure and function of in-vivo organs, and can better reflect the physiological characteristics of human bodies.
The organ chip technology still has a lot of technical problems to be solved at present, and because the chip is small in size and low in cell capacity, a convenient and high-sensitivity detection method and device need to be developed. It can achieve accurate real-time detection of cellular processes and biomarkers without affecting cell viability, which requires integration of a large number of microsensors in the design of organ chips. In addition, some tests such as cell sequencing require sampling from the inside of the chip, so the organ chip also needs to integrate sampling flow channels and interfaces, which makes the whole system very complicated, and this greatly restricts the development of the organ chip. In addition, the structure of the existing organ chip is difficult to change after the construction is finished, so that the current organ chip cannot simulate the dynamic change of human tissues, and cannot well realize the bionic function under partial conditions.
Disclosure of Invention
The purpose of the invention is as follows: in view of the deficiencies of the prior art organ chip systems in detection and dynamic changes, it is an object of the present invention to provide an all-liquid organ chip that does not have the solid polymer or glass housing of conventional microfluidic chips; another objective of the invention is to provide a method for preparing an all-liquid-phase organ chip.
The technical scheme is as follows: the full liquid phase organ chip comprises a super-hydrophobic porous membrane with a hydrophilic channel, wherein the porous membrane is sealed in an oil phase, and the hydrophilic channel is provided with a fluid inlet and a fluid outlet which are communicated with the hydrophilic channel.
The invention also provides a preparation method of the whole liquid phase organ chip, which comprises the following steps:
(1) preparing a polymer layer porous membrane;
(2) placing the polymer layer porous membrane in a functional modification solution to obtain a hydrophobic polymer layer porous membrane;
(3) carrying out hydrophilic pattern treatment on the hydrophobic polymer layer porous membrane to obtain a porous membrane with a hydrophilic channel;
(4) then carrying out polymer porous membrane surface super-hydrophobic treatment on the porous membrane surface with the hydrophilic channel;
(5) and then wetting the hydrophilic channel on the surface of the porous membrane, sealing the hydrophilic channel in the oil phase, and arranging an inlet and an outlet which are communicated with the hydrophilic channel. The syringe needle can be used as a water phase channel to be connected with the fluid inlet and the fluid outlet, and the micro-fluidic chip can be formed by pushing the syringe.
Preferably, the method for preparing the polymer layer porous membrane comprises the following steps: carrying out double bond modification treatment on the substrate to obtain a substrate with surface modified double bonds, then adding a prepolymer mixture, and reacting under ultraviolet light; the prepolymer mixture includes a monomer, a crosslinker, a porogen, and an additional photoinitiator. The substrate here may be a conventional substrate such as hydroxyl glass, plastic, metal, or the like.
The preparation method of the polymer layer porous membrane is characterized in that a polymerization-initiated phase separation technology is utilized, so that the prepolymer solution which is originally homogeneous in phase is polymerized under ultraviolet illumination, the polymer and the pore-forming agent are separated, and finally the pore-forming agent is cleaned and removed to obtain the HEMA-EDMA porous membrane. The photoinitiator can be benzoin dimethyl ether, or common ultraviolet initiators such as Bassfer Irgcure 369, 819, 1173, 2959.
Preferably, in order to obtain a porous membrane with good appearance, suitable super-hydrophobicity and relatively stable mechanical property; the prepolymer mixture comprises the following components in percentage by volume: 12-16 vol% of hydroxyethyl methacrylate (HEMA), 4-8 vol% of ethylene glycol dimethacrylate (EDMA), 40-64 vol% of cyclohexanol and 16-40 vol% of n-decanol; benzoin dimethyl ether is used as a photoinitiator, and 0.5 mg-4 mg of the photoinitiator is added into each 1ml of prepolymer mixture.
Preferably, the double bond modification treatment is carried out on the glass sheet by adding a 3- (methacryloyloxy) propyl trimethoxy silane solution with the mass concentration of 10-25 wt%; wherein the solvent of the 3- (methacryloyloxy) propyl trimethoxy silane solution is ethanol.
Preferably, the polymer layer porous membrane is placed in the functional modification solution to react for 3-10 hours; the solvent of the functionalized modification solution is dichloromethane, and the solute is N, N' -diisopropylcarbodiimide, 4-dimethylaminopyridine, 4-pentenoic acid or 4-pentynoic acid; wherein the adding amount of the N, N' -diisopropylcarbodiimide is 2.5-4 mu L, the adding amount of the 4-dimethylaminopyridine is 1-2 mg, and the adding amount of the 4-pentenoic acid or 4-pentynoic acid is 2-3 mu L in each 1mL of dichloromethane by volume of the solvent.
Further, the hydrophilic pattern processing method comprises: dropwise adding a modifying solution to the surface of the hydrophobic polymer layer porous membrane, covering a quartz slide, covering by a photomask, and reacting under ultraviolet light to obtain the porous membrane with a hydrophilic channel; the modification solution is any one of cysteine hydrochloride solution, mercaptoethanol solution, mercaptoethylamine solution and mercaptopropionic acid solution. The preparation process of the cysteine hydrochloride solution comprises the steps of adding cysteine hydrochloride and benzoin dimethyl ether (DMPA) into N, N-Dimethylformamide (DMF), and vortexing until the cysteine hydrochloride is completely dissolved to obtain a functional modification solution; the mercaptoethanol solution is prepared by adding mercaptoethanol and benzoin dimethyl ether (DMPA) into N, N-Dimethylformamide (DMF), and vortexing until the mercaptoethanol is completely dissolved to obtain a functional modification solution; the preparation method of the mercaptoethylamine solution and the mercaptopropionic acid solution is similar.
Preferably, the superhydrophobic treatment method is as follows: dripping 1H, 1H, 2H, 2H-perfluorodecanethiol solution on a porous membrane with a hydrophilic channel, covering a quartz slide, and reacting under ultraviolet light; the preparation method of the 1H, 1H, 2H, 2H-perfluorodecanethiol solution comprises the following steps: toluene is used as a solvent, a solute is perfluorodecyl mercaptan, and the perfluorodecyl mercaptan is added into the toluene.
Preferably, the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol in the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 5-10 wt%.
Preferably, the wetting treatment adopts a nano clay water solution, and each 1mL of the nano clay water solution contains 10-25 mg of nano clay. The oil phase comprises 80-95 vol% of silicone oil and amino-terminated polydimethylsiloxane (H) 2 N-PDMS-NH 2 ) 5-20 vol%. The silicone oil can be replaced by toluene, dodecane, hexadecane, mineral oil and vegetable oil.
The invention principle is as follows: firstly, carrying out modification treatment on a pretreated glass sheet to obtain glass with surface modified double bonds; then synthesizing a HEMA-EDMA polymer layer porous membrane on the glass with surface modified double bonds; performing double bond modification treatment on the polymer layer porous membrane to obtain a hydrophobic polymer layer porous membrane; preparing a patterned hydrophilic channel on the prepared hydrophobic polymer layer porous membrane, adding a modifying solution, quickly covering a glass slide, covering the glass slide by a mask plate, and reacting under ultraviolet light; carrying out surface super-hydrophobic treatment on the porous membrane with the channel; finally, wetting the hydrophilic channel by using a wetting solution, placing the hydrophilic channel in an oil phase to seal the hydrophilic channel so as to form a water flow channel in a closed and oil phase, and then arranging a flow channel inlet and a flow channel outlet which are communicated with the hydrophilic channel so as to form a complete microfluidic system.
The full liquid phase organ chip adopts the oil phase to replace the traditional polymer shell, and various probes can enter and exit the organ chip without damage due to the liquidity of the oil phase, so that various parameters of a microenvironment in the chip can be detected in situ in real time at a position to be detected, and sampling can be carried out at any position in the chip in real time, thereby well solving the problem of difficult detection of the traditional chip; in addition, the full liquid phase chip prepared by the invention can also carry out spatial editing on the function and geometric layout of the fluid device according to the requirement, including channel separation, micro-channel introduction, channel flow path change and the like, and has considerable potential for the development of standardized organ chips.
Has the advantages that: compared with the prior art, the method has the advantages that,
(1) the preparation method of the full liquid phase organ chip has universality, and the method has no selectivity on the type of the organ chip, such as a liver chip, a kidney chip, an intestine chip and the like, and can be used for preparing the full liquid phase organ chip;
(2) the full-liquid-phase organ chip and the preparation method thereof can realize in-situ detection and sampling of microenvironment in each part of the organ chip, the detection is convenient, rapid and lossless, and the problems that the traditional organ chip cannot use the traditional biological detection method due to the solidified shell are solved;
(3) The preparation method is mild in preparation condition, and can construct cell patterning culture on the prepared substrate to realize the construction of the full liquid phase organ chip. In addition, the channels of the fluid device can be separated, introduced into the micro-channels, flow paths can be changed and the like, so that the universality of the fluid device is expanded, and the spatial editing of the function and the geometric layout of the fluid device can be realized according to the requirement;
(4) the chip is sealed by oil phase, and has no solid polymer or glass shell of the traditional micro-fluidic chip, and various probes can enter and exit the chip without damage in the using process, and can detect various parameters of the microenvironment in the chip in situ in real time at the position to be detected, and can also sample at any position in the chip in real time; the method can carry out spatial editing on the function and geometric layout of the fluid device according to the requirement, including channel separation, microchannel introduction and channel flow path change, and has good application prospect.
Drawings
FIG. 1 is a flow chart of a preparation method of the present invention;
FIG. 2 is a schematic view of a superhydrophilic-superhydrophobic patterned substrate provided by the present invention;
FIG. 3 is a schematic view of an apparatus for preparing a whole liquid phase organ chip according to the present invention;
FIG. 4 is a photograph of a full liquid phase organ-chip device.
Detailed Description
As used in the following examples: 3- (methacryloyloxy) propyltrimethoxysilane, hydroxyethyl methacrylate (HEMA), ethylene glycol dimethacrylate (EDMA), cyclohexanol, N-decanol, benzoin dimethyl ether, Dichloromethane (DCM), 4-Pentenoic Acid (PA), 4-pentynoic acid, N' -Diisopropylcarbodiimide (DIC), 1H, 2H, 2H-perfluorodecanethiol, silicone oil, amino-terminated polydimethylsiloxane (H) 2 N-PDMS-NH 2 ) The reagents and materials are all commercially available.
Example 1:
the flow of preparing the whole liquid phase organ chip in this embodiment is shown in fig. 1, and specifically includes the following steps:
(1) preparation of surface-modified double-bond glass
Adding 30 mu L of 3- (methacryloyloxy) propyl trimethoxy silane solution into a gap between two pieces of pretreated hydroxy glass, completely infiltrating the surface between the two pieces of glass slides, ensuring that no bubbles are generated, reacting for 1h, respectively washing with water and ethanol for multiple times, and drying by blowing to obtain the glass slide with surface modified double bonds;
wherein the mass concentration of the 3- (methacryloyloxy) propyl trimethoxy silane in the 3- (methacryloyloxy) propyl trimethoxy silane solution is 20 wt%, and the solvent is ethanol.
(2) Preparation of Polymer layer porous Membrane
Preparing a prepolymer mixture: the prepolymer mixture solution comprises the following components in percentage by volume: HEMA (monomer) accounting for 12 vol%, EDMA (cross-linking agent) accounting for 8 vol%, cyclohexanol (pore-forming agent) accounting for 64 vol%, n-decanol (pore-forming agent) accounting for 16 vol%, and benzoin dimethyl ether (DMPA) serving as a photoinitiator, wherein the addition amount of the photoinitiator accounts for 2mg/mL of the concentration of the mixture; placing a double-bond modified glass sheet on a table top, placing two strips of PI films with the film thickness of 50 mu m on the two sides of the long side of the glass sheet, dropwise adding a proper amount of prepolymer mixed solution, slightly covering a piece of fluorinated glass to ensure that the solution is completely filled between the two glass sheets and no bubbles are generated, and placing the glass sheet under 365nm ultraviolet light for reaction for 30 min. Then washing with ethanol and water for several times to remove unpolymerized monomer, and blow-drying to obtain HEMA-EDMA porous membrane.
(3) Preparation of hydrophobic Polymer layer porous Membrane
Respectively adding 40mL of Dichloromethane (DCM), 100 mu L of 4-Pentenoic Acid (PA) and 120 mu L N, N' -Diisopropylcarbodiimide (DIC) into a 50mL centrifuge tube under a fume hood environment, stirring for 30min, adding the HEMA-EDMA porous membrane prepared in the step (2) into the centrifuge tube, adding 56mg of 4-Dimethylaminopyridine (DMAP) into the centrifuge tube, and stirring for reacting for 4 h; after the reaction is finished, the double-bond modified polymer layer porous membrane can be obtained after the double-bond modified polymer layer porous membrane is washed by ethanol for many times and dried by blowing.
(4) Hydrophilic pattern treatment on hydrophobic polymer layer porous membrane
Preparing cysteine hydrochloride solution: adding 0.2g of cysteine hydrochloride and 0.01g of benzoin dimethyl ether (DMPA) into 4mL of N, N-Dimethylformamide (DMF), and vortexing until the cysteine hydrochloride is completely dissolved to obtain a functional modification solution; dripping a proper amount of prepared cysteine hydrochloride solution on the surface of the porous membrane to be modified, quickly covering a quartz slide, covering the quartz slide by a photomask, reacting for 7min under ultraviolet light, and washing and drying the porous membrane by ethanol for multiple times to obtain the porous membrane with the hydrophilic channel;
(5) super-hydrophobic treatment for polymer porous membrane surface
And (3) dropwise adding a 1H, 1H, 2H, 2H-perfluorodecanethiol solution on the polymer porous membrane with the hydrophilic channel prepared in the step (4), quickly covering a quartz slide, placing the quartz slide under ultraviolet light to irradiate the polymer layer for 2min, and washing and drying the polymer porous membrane with acetone for multiple times to obtain the patterned super-hydrophilic-hydrophobic polymer surface, wherein a gray base part is a HEMA-EDMA porous membrane as shown in a figure 2, and a pattern part in the middle part of the figure 2 is a hydrophilic pattern.
Wherein the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 5 wt%, and the 1H, 1H, 2H, 2H-perfluorodecanethiol is added into toluene by taking the toluene as a solvent.
(6) Preparation of full liquid phase organ chip
Wetting the hydrophilic channel on the surface of the HEMA-EDMA porous membrane prepared in the step (5) by using a 10mg/mL nanoclay aqueous solution, connecting a water phase channel to a fluid inlet and a fluid outlet by using a syringe needle as the water phase channel, adding interfacial force in an oil phase to fix and limit the water phase in the oil phase to form a fluid channel, and pushing and pulling the syringe to form the full-liquid-phase microfluidic chip, as shown in FIGS. 3 and 4; wherein the oil phase comprises 90 vol% of silicone oil and 10 vol% of amino-terminated polydimethylsiloxane. In fig. 3, the right downward arrow indicates the inlet of the fluid channel, the left upward arrow indicates the outlet of the fluid channel, the middle is the sampler, and the lower part is the hydrophilic channel wetted by the nanoclay aqueous solution immersed in the silicone oil-amino terminated polydimethylsiloxane oil phase.
When the full-liquid-phase microfluidic chip prepared by the embodiment is used, two syringe needles are respectively inserted into the water phases at the two ends of the flow channel, the fluid pipeline and the push-pull pump are connected to the back of the syringe needle, and the inlet and the outlet of the flow channel are respectively pushed in and sucked out of the water phases at the same flow rate, so that the microfluidic system can be formed.
Example 2:
(1) preparation of a substrate with surface-modified double bonds
This procedure is essentially the same as example 1, except that: wherein the mass concentration of the 3- (methacryloyloxy) propyl trimethoxy silane in the 3- (methacryloyloxy) propyl trimethoxy silane solution is 10 wt%, and the solvent is ethanol.
Wherein the substrate is plastic, the plastic substrate is pretreated with oxygen plasma to make it hydrophilic, and then the substrate is modified by the same procedure as in example 1.
(2) Preparation of Polymer layer porous Membrane
The preparation of the polymer layer porous film in this step is substantially the same as in example 1 except that: wherein the prepolymer mixture solution comprises 15 vol% HEMA (monomer), 6 vol% EDMA (cross-linking agent), 50 vol% cyclohexanol (pore-forming agent), 29 vol% n-decanol (pore-forming agent), and benzoin dimethyl ether (DMPA) is used as a photoinitiator, and the addition amount of the photoinitiator accounts for 0.5mg/mL of the mixture;
(3) preparation of hydrophobic Polymer layer porous Membrane
Respectively adding 40mL of Dichloromethane (DCM), 90 mu L of 4-pentynoic acid and 160 mu L N, N' -Diisopropylcarbodiimide (DIC) into a 50mL centrifuge tube under a fume hood environment, stirring, adding the HEMA-EDMA porous membrane prepared in the step (2) into the centrifuge tube, adding 60mg of 4-Dimethylaminopyridine (DMAP) into the centrifuge tube, and stirring for reacting for 3 hours; after the reaction is finished, the double-bond modified polymer layer porous membrane can be obtained after the double-bond modified polymer layer porous membrane is washed by ethanol for many times and dried by blowing.
(4) Hydrophilic pattern treatment on hydrophobic polymer layer porous membrane
Preparing a mercaptoethanol solution: adding 0.2g of mercaptoethanol and 0.01g of benzoin dimethyl ether into 4mL of N, N-dimethylformamide, and vortexing until the mercaptoethanol is completely dissolved to obtain a functional modification solution; dropwise adding a proper amount of prepared mercaptoethanol solution on the surface of the porous membrane to be modified, quickly covering a quartz slide, covering by a photomask, reacting for 7min under ultraviolet light, and washing and drying by ethanol for multiple times to obtain the porous membrane with the hydrophilic channel;
(5) Super-hydrophobic treatment for polymer porous membrane surface
The procedure is substantially the same as that of example 1, except that the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 8 wt%;
(6) preparation of full liquid phase organ chip
Wetting the hydrophilic channel on the surface of the HEMA-EDMA porous membrane prepared in the step (5) by using a 15mg/mL nano clay aqueous solution, connecting a syringe needle serving as a water phase channel into a fluid inlet and a fluid outlet, adding interfacial force in an oil phase to fix and limit the water phase in the oil phase to form a fluid channel, and pushing and pulling the syringe to form the full-liquid-phase microfluidic chip; wherein the oil phase comprises 80 vol% of silicone oil and 20 vol% of amino-terminated polydimethylsiloxane.
Example 3:
(1) preparation of surface-modified double-bond glass
This procedure is essentially the same as example 1, except that: wherein the mass concentration of the 3- (methacryloyloxy) propyl trimethoxy silane in the 3- (methacryloyloxy) propyl trimethoxy silane solution is 25 wt%, and the solvent is ethanol.
(2) Preparation of Polymer layer porous Membrane
The preparation of the polymer layer porous film in this step is substantially the same as in example 1 except that: the prepolymer mixture solution comprises 16 vol% HEMA (monomer), 4 vol% EDMA (cross-linking agent), 40 vol% cyclohexanol (pore-foaming agent) and 40 vol% n-decanol (pore-foaming agent), benzoin dimethyl ether (DMPA) is used as a photoinitiator, and the addition amount of the photoinitiator accounts for the concentration of the mixture to be 4 mg/mL;
(3) Preparation of a hydrophobic Polymer layer porous Membrane
Respectively adding 40mL of Dichloromethane (DCM), 80 mu L of 4-pentynoic acid and 100 mu L N, N' -Diisopropylcarbodiimide (DIC) into a 50mL centrifuge tube under a fume hood environment, stirring, adding the HEMA-EDMA porous membrane prepared in the step (2) into the centrifuge tube, adding 40mg of 4-Dimethylaminopyridine (DMAP) into the centrifuge tube, and stirring for reacting for 10 hours; after the reaction is finished, the double-bond modified polymer layer porous membrane can be obtained after the double-bond modified polymer layer porous membrane is washed by ethanol for many times and dried by blowing.
(4) Hydrophilic pattern treatment on hydrophobic polymer layer porous membrane
Preparing a mercaptoethylamine solution: adding 0.2g of mercaptoethylamine and 0.01g of benzoin dimethyl ether into 4mL of N, N-dimethylformamide, and vortexing until mercaptoethanol is completely dissolved to obtain a functional modification solution; dropwise adding a proper amount of prepared mercaptoethylamine solution on the surface of the porous membrane to be modified, quickly covering a quartz slide, covering by using a photomask, reacting for 7min under ultraviolet light, and washing and drying by using ethanol for multiple times to obtain the porous membrane with a hydrophilic channel;
(5) super-hydrophobic treatment for polymer porous membrane surface
The procedure is substantially the same as that of example 1, except that the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 10 wt%;
(6) Preparation of full liquid phase organ chip
Wetting the hydrophilic channel on the surface of the HEMA-EDMA porous membrane prepared in the step (5) by using a nano clay aqueous solution with the concentration of 25mg/mL, connecting a syringe needle serving as a water phase channel into a fluid inlet and a fluid outlet, adding interfacial force in an oil phase to fix and limit the water phase in the oil phase to form a fluid channel, and pushing and pulling the syringe to form the full-liquid-phase microfluidic chip; wherein the oil phase comprises 95 vol% of silicone oil and 5 vol% of amino-terminated polydimethylsiloxane.
Example 4:
(1) preparation of surface-modified double-bond glass
This procedure is essentially the same as example 1, except that: wherein the mass concentration of the 3- (methacryloyloxy) propyl trimethoxy silane in the 3- (methacryloyloxy) propyl trimethoxy silane solution is 15 wt%, and the solvent is ethanol.
(2) Preparation of Polymer layer porous Membrane
The preparation of the polymer layer porous film in this step is substantially the same as in example 1 except that: wherein the prepolymer mixture solution comprises 15 vol% HEMA (monomer), 5 vol% EDMA (cross-linking agent), 60 vol% cyclohexanol (pore-forming agent) and 20 vol% n-decanol (pore-forming agent), and benzoin dimethyl ether (DMPA) is used as a photoinitiator, and the addition amount of the photoinitiator accounts for 3mg/mL of the mixture;
(3) Preparation of hydrophobic Polymer layer porous Membrane
Respectively adding 40mL of Dichloromethane (DCM), 120 mu L of 4-pentynoic acid and 140 mu L N, N' -Diisopropylcarbodiimide (DIC) into a 50mL centrifuge tube under a fume hood environment, stirring, adding the HEMA-EDMA porous membrane prepared in the step (2) into the centrifuge tube, adding 80mg of 4-Dimethylaminopyridine (DMAP) into the centrifuge tube, and stirring for reacting for 10 hours; after the reaction is finished, the double-bond modified polymer layer porous membrane can be obtained after the double-bond modified polymer layer porous membrane is washed by ethanol for many times and dried by blowing.
(4) Hydrophilic pattern treatment on hydrophobic polymer layer porous membrane
The procedure is as in example 1;
(5) super-hydrophobic treatment for polymer porous membrane surface
The procedure is substantially the same as that of example 1, except that the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 6 wt%;
(6) preparation of full liquid phase organ chip
This procedure is essentially the same as example 1, except that the nanoclay in aqueous nanoclay solution contains 20mg of nanoclay per 1ml of solution.

Claims (10)

1. A full liquid phase organ chip, characterized in that: the organ chip comprises a super-hydrophobic porous membrane with hydrophilic channels, wherein the porous membrane is sealed in the oil phase, and the hydrophilic channels are connected with the inlet and the outlet of fluid.
2. A method for preparing the full liquid phase organ chip according to claim 1, which comprises the steps of:
(1) preparing a polymer layer porous membrane;
(2) placing the polymer layer porous membrane in a functional modification solution to obtain a hydrophobic polymer layer porous membrane;
(3) carrying out hydrophilic pattern treatment on the hydrophobic polymer layer porous membrane to obtain a porous membrane with a hydrophilic channel;
(4) performing polymer surface super-hydrophobic treatment on the surface of the porous membrane with the hydrophilic channel;
(5) and (3) wetting the hydrophilic channel on the surface of the porous membrane, sealing the hydrophilic channel in the oil phase, and connecting a fluid inlet and a fluid outlet which are communicated with the hydrophilic channel.
3. The method for preparing a full liquid phase organ chip according to claim 2, characterized in that: the preparation method of the polymer layer porous membrane comprises the following steps: treating the substrate, modifying double bonds, adding the prepolymer mixture, and reacting under ultraviolet light; the prepolymer mixture includes a monomer, a crosslinker, a porogen, and a photoinitiator.
4. The method for preparing a full liquid phase organ chip according to claim 3, characterized in that: the prepolymer mixture comprises the following components in percentage by volume: 12-16 vol% of hydroxyethyl methacrylate, 4-8 vol% of ethylene glycol dimethacrylate, 40-64 vol% of cyclohexanol and 16-40 vol% of n-decanol; benzoin dimethyl ether is used as a photoinitiator, and 0.5 mg-4 mg of the photoinitiator is added into each 1ml of prepolymer mixture.
5. The method of preparing a full liquid phase functional chip according to claim 3, wherein: performing double bond modification treatment on the substrate by adding 3- (methacryloyloxy) propyl trimethoxy silane solution; the mass concentration of the 3- (methacryloyloxy) propyl trimethoxy silane in the 3- (methacryloyloxy) propyl trimethoxy silane solution is 10-25 wt%.
6. The method for preparing a full liquid phase organ chip according to claim 2, characterized in that: the polymer layer porous membrane is placed in the functional modification solution to react for 3-10 hours; the solvent of the functionalized modification solution is dichloromethane, and the solute is N, N' -diisopropylcarbodiimide, 4-dimethylaminopyridine, 4-pentenoic acid or 4-pentynoic acid; wherein the adding amount of the N, N' -diisopropylcarbodiimide is 2.5-4 mu L, the adding amount of the 4-dimethylaminopyridine is 1-2 mg, and the adding amount of the 4-pentenoic acid or 4-pentynoic acid is 2-3 mu L in each 1mL of dichloromethane by volume of the solvent.
7. The method for preparing a full liquid phase organ chip according to claim 2, characterized in that: the hydrophilic pattern treatment method comprises the following steps: dropwise adding a modifying solution to the surface of the hydrophobic polymer layer porous membrane, covering a glass slide, covering by a photomask, and reacting under ultraviolet light to obtain the porous membrane with a hydrophilic channel; the modification solution is any one of cysteine hydrochloride solution, mercaptoethanol solution, mercaptoethylamine solution and mercaptopropionic acid solution.
8. The method for preparing a full liquid phase organ chip according to claim 2, characterized in that: the super-hydrophobic treatment method comprises the following steps: dripping 1H, 1H, 2H, 2H-perfluorodecanethiol solution on the porous membrane with the hydrophilic channel, covering a quartz slide, and reacting under ultraviolet light.
9. The method for preparing a full liquid phase organ chip according to claim 8, characterized in that: the mass concentration of the 1H, 1H, 2H, 2H-perfluorodecanethiol in the 1H, 1H, 2H, 2H-perfluorodecanethiol solution is 5-10 wt%.
10. The method for preparing a full liquid phase organ chip according to claim 2, characterized in that: the wetting treatment adopts a nano clay aqueous solution, and each 1mL of the nano clay aqueous solution contains 10-25 mg of nano clay; the oil phase comprises 80-95 vol% of silicone oil and 5-20 vol% of amino-terminated polydimethylsiloxane.
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