CN110773244A - Micro-fluidic chip for high-throughput screening of nano-particles in cross-vascular transport and preparation method thereof - Google Patents

Micro-fluidic chip for high-throughput screening of nano-particles in cross-vascular transport and preparation method thereof Download PDF

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CN110773244A
CN110773244A CN201911016382.6A CN201911016382A CN110773244A CN 110773244 A CN110773244 A CN 110773244A CN 201911016382 A CN201911016382 A CN 201911016382A CN 110773244 A CN110773244 A CN 110773244A
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史岩彬
高岩
孔德印
付孟广
蒋波
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Qilu University of Technology
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Abstract

一种纳米颗粒跨血管输运高通量筛选的微流控芯片及制备方法,所述微流控芯片包括流体通道层以及与流体通道层连接的玻璃支撑层,所述流体通道层的下端面与玻璃支撑层的上端面接触,所述流体通道层内设置有流体通道Ⅰ和流体通道Ⅱ,所述流体通道Ⅰ的一端设置有进样口,所述流体通道Ⅰ的另一端设置有出样口,所述流体通道Ⅰ的中部通过间隙通道与流体通道Ⅱ连通,所述流体通道Ⅱ的一端设置有渗出口。所述微流控芯片能够用来筛选适合穿透血管壁的载药纳米颗粒,其筛选的准确率高。

Figure 201911016382

A microfluidic chip for high-throughput screening of nanoparticle transvascular transport and preparation method, the microfluidic chip comprises a fluid channel layer and a glass support layer connected with the fluid channel layer, the lower end surface of the fluid channel layer is In contact with the upper end surface of the glass support layer, the fluid channel layer is provided with a fluid channel I and a fluid channel II, one end of the fluid channel I is provided with a sample inlet, and the other end of the fluid channel I is provided with a sample outlet The middle part of the fluid channel I communicates with the fluid channel II through a gap channel, and one end of the fluid channel II is provided with a seepage outlet. The microfluidic chip can be used to screen drug-loaded nanoparticles suitable for penetrating blood vessel walls, and the screening accuracy is high.

Figure 201911016382

Description

纳米颗粒跨血管输运高通量筛选的微流控芯片及制备方法Microfluidic chip for high-throughput screening of nanoparticle transvascular transport and preparation method

技术领域technical field

本发明涉及将微流控芯片技术应用到生物医学检测筛选的技术领域,具体涉及一种纳米颗粒跨血管输运高通量筛选的微流控芯片及制备方法。The invention relates to the technical field of applying microfluidic chip technology to biomedical detection and screening, in particular to a microfluidic chip for high-throughput screening of nanoparticle transvascular transport and a preparation method.

背景技术Background technique

肿瘤目前已成为威胁人类健康的头号杀手,高效抗肿瘤药物已成为人民健康生活的重大需求。近年来随着纳米技术的发展,纳米颗粒载药治疗肿瘤已经被广泛的研究。通过纳米技术,不仅可以提高药物的水溶性、改善药物在体内的药代动力学参数还可以控制药物的释放速度。但是,就目前来说治疗效果依旧不理想。其原因在于肿瘤微环境在理化性质方面与人体正常内环境存在着许多不相同的地方,比较显著的是其低氧、低pH、高渗透压以及高间质流体压力的特征。也正是因为这些特征,致使药物无法以均匀有效的浓度到达肿瘤组织,导致局部药物浓度不足,这种异质性的药物分布是载药纳米颗粒治疗效果不佳的主要原因。研究如何使纳米颗粒最大限度的到达肿瘤是治疗肿瘤的关键,因此有必要充分的认识纳米颗粒在人体内的输运过程。Tumor has now become the number one killer threatening human health, and high-efficiency anti-tumor drugs have become a major demand for people's healthy life. In recent years, with the development of nanotechnology, nanoparticle-loaded drug therapy for tumors has been widely studied. Nanotechnology can not only improve the water solubility of the drug, improve the pharmacokinetic parameters of the drug in the body, but also control the release rate of the drug. However, so far, the treatment effect is still not ideal. The reason is that there are many differences between the tumor microenvironment and the normal internal environment of the human body in terms of physicochemical properties, the most notable being its characteristics of low oxygen, low pH, high osmotic pressure and high interstitial fluid pressure. It is precisely because of these characteristics that the drug cannot reach the tumor tissue in a uniform and effective concentration, resulting in insufficient local drug concentration. This heterogeneous drug distribution is the main reason for the poor therapeutic effect of drug-loaded nanoparticles. Studying how to make the nanoparticles reach the tumor to the maximum extent is the key to the treatment of tumors, so it is necessary to fully understand the transport process of nanoparticles in the human body.

纳米颗粒在人体中的运输过程大致可以分为四个阶段,在血管中的输运、穿透血管壁进入组织间隙、在组织间隙中的输运以及进入肿瘤细胞。其中穿透血管壁的输运在整个输运过程中扮演着重要的角色,由于肿瘤血管壁缺少基膜和粘附蛋白导致其血管壁上出现很多孔洞,所以纳米颗粒就可以通过这些孔洞穿透血管壁。然而研究人员却很少进行这方面的研究,对于纳米颗粒的物理化学性质以及血管中的环境对其穿透血管壁效率的影响仍然缺乏一个清晰的认识。因此,建立一种器件来筛选适合穿透血管壁的纳米颗粒和适宜的血管环境是当前肿瘤治疗研究的首要任务。The transport process of nanoparticles in the human body can be roughly divided into four stages: transport in blood vessels, penetrating the blood vessel wall into the interstitial space, transporting in the interstitial space and entering tumor cells. Among them, the transport through the vascular wall plays an important role in the entire transport process. Due to the lack of basement membrane and adhesion proteins in the tumor vascular wall, there are many holes in the vascular wall, so the nanoparticles can penetrate through these holes. Vascular wall. However, few studies have been conducted in this area, and a clear understanding of the physicochemical properties of nanoparticles and the influence of the environment in the blood vessel on their efficiency of penetrating the vessel wall is still lacking. Therefore, establishing a device to screen nanoparticles suitable for penetrating the vascular wall and suitable vascular environment is the primary task of current tumor therapy research.

微流控技术是近年来发展迅速的新兴交叉学科技术,通过在芯片上构建微型通道可以处理或操作微小流体,实现一系列常规方法所难以完成的微加工和微操作。在空间维度上能够提供更为准确的操作,尺度同实际尺度更相近。因此微流控芯片可以作为模拟人体内环境的平台。目前,利用微流控芯片来筛选适合穿透血管壁的纳米颗粒和适宜的血管环境的研究尚处于空白。Microfluidics is an emerging interdisciplinary technology that has developed rapidly in recent years. By constructing microchannels on a chip, tiny fluids can be processed or manipulated, and a series of microfabrication and micromanipulations that are difficult to accomplish by conventional methods can be realized. It can provide more accurate operations in the spatial dimension, and the scale is closer to the actual scale. Therefore, the microfluidic chip can be used as a platform for simulating the internal environment of the human body. At present, the use of microfluidic chips to screen nanoparticles suitable for penetrating the vascular wall and suitable vascular environment is still in the blank.

发明内容SUMMARY OF THE INVENTION

本发明所要解决的技术问题,就是针对现有技术所存在的不足,而提供一种纳米颗粒跨血管输运高通量筛选的微流控芯片及制备方法,该微流控芯片能够用来筛选适合穿透血管壁的载药纳米颗粒,其筛选的准确率高,可以满足现代载药纳米颗粒穿透血管屏障医学研究的需要。The technical problem to be solved by the present invention is to provide a microfluidic chip for high-throughput screening of nanoparticle transvascular transport and a preparation method for the deficiencies in the prior art. The microfluidic chip can be used for screening The drug-loaded nanoparticles suitable for penetrating the blood vessel wall have high screening accuracy and can meet the needs of modern drug-loaded nanoparticles for medical research on vascular barrier penetration.

本方案是通过如下技术措施来实现的:一种纳米颗粒跨血管输运高通量筛选的微流控芯片,其特征是:包括流体通道层以及与流体通道层连接的玻璃支撑层,所述流体通道层的下端面与玻璃支撑层的上端面接触,所述流体通道层内设置有流体通道Ⅰ和流体通道Ⅱ,所述流体通道Ⅰ的一端设置有进样口,所述流体通道Ⅰ的另一端设置有出样口,所述流体通道Ⅰ的中部通过间隙通道与流体通道Ⅱ连通,所述流体通道Ⅱ的一端设置有渗出口。载药纳米颗粒溶液经压力施加装置从进样口注入流体通道Ⅰ,在经过间隙通道时,一部分载药纳米颗粒溶液会通过流体通道Ⅰ从出样口流出,另一部分载药纳米颗粒溶液通过间隙通道进入流体通道Ⅱ并从渗出口流出,收集渗出口流出的样品溶液。更换载药纳米颗粒溶液重复上述过程,对比收集样品溶液中的载药纳米颗粒含量,即可筛选出更适合穿透肿瘤血管壁的载药纳米颗粒。The solution is realized by the following technical measures: a microfluidic chip for high-throughput screening of nanoparticle transvascular transport, which is characterized by comprising: a fluid channel layer and a glass support layer connected to the fluid channel layer; the The lower end surface of the fluid channel layer is in contact with the upper end surface of the glass support layer, the fluid channel layer is provided with a fluid channel I and a fluid channel II, one end of the fluid channel I is provided with a sample inlet, and the fluid channel I is provided with a sample inlet. The other end is provided with a sample outlet, the middle part of the fluid channel I is communicated with the fluid channel II through a gap channel, and one end of the fluid channel II is provided with a seepage outlet. The drug-loaded nanoparticle solution is injected into the fluid channel I from the injection port through the pressure application device. When passing through the gap channel, a part of the drug-loaded nanoparticle solution will flow out from the sample outlet through the fluid channel I, and the other part of the drug-loaded nanoparticle solution will pass through the gap. The channel enters fluid channel II and flows out from the permeate port, and the sample solution flowing out of the permeate port is collected. The above process is repeated by replacing the drug-loaded nanoparticle solution, and the drug-loaded nanoparticles that are more suitable for penetrating the tumor blood vessel wall can be screened by comparing the content of the drug-loaded nanoparticles in the collected sample solution.

所述微流控芯片采用模塑法进行制备,包括以下步骤:The microfluidic chip is prepared by a molding method, including the following steps:

(1)清洗单晶硅片:用去离子水冲洗,完毕后用氮气吹干,然后置于加热台上静置;(1) Cleaning the single crystal silicon wafer: rinse with deionized water, blow dry with nitrogen after completion, and then place it on a heating table to stand;

(2)涂胶:将正光刻胶涂于单晶硅片表面,并进行甩涂;(2) Glue coating: apply positive photoresist to the surface of the single crystal silicon wafer and spin it;

(3)烘焙:将甩涂后的单晶硅片置于加热台上静置;(3) Baking: put the spin-coated monocrystalline silicon wafer on the heating table and let it stand;

(4)曝光:将光刻掩膜板置于光刻胶表面,进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist for exposure;

(5)显影:对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: develop the exposed single crystal silicon wafer, then clean the surface of the single crystal silicon wafer with deionized water, and dry it with nitrogen;

(6)后烘:将单晶硅片置于加热台上静置,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: the single crystal silicon wafer is placed on a heating table to stand, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于硅基模具上,除去气泡,置于加热台上静置使PDMS完全固化,再将完全固化后的PDMS从硅基模具上揭下,得到PDMS基体,即为微流控芯片的流体通道层;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on a silicon-based mold, remove air bubbles, and place it on a heating table to allow PDMS to completely cure. , and then peel off the fully cured PDMS from the silicon-based mold to obtain the PDMS matrix, which is the fluid channel layer of the microfluidic chip;

(8)清洗玻璃底片:将玻璃底片依次经过无水乙醇和去离子水的冲洗,然后用氮气吹干;(8) Cleaning the glass negative film: Rinse the glass negative film with absolute ethanol and deionized water in sequence, and then dry it with nitrogen;

(9)键合:将PDMS基体和玻璃底片一起放入等离子体机中进行清洗,清洗完毕后将PDMS基体和玻璃底片贴合且保证贴合面不留气泡,然后用重物压实。(9) Bonding: Put the PDMS substrate and the glass negative into the plasma machine for cleaning. After cleaning, the PDMS substrate and the glass negative are bonded together to ensure that no air bubbles remain on the bonding surface, and then compacted with a heavy object.

进一步的,所述微流控芯片的制备方法包括以下步骤:Further, the preparation method of the microfluidic chip includes the following steps:

(1)清洗单晶硅片:首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) Cleaning the single crystal silicon wafer: first rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C Set for 15 minutes;

(2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s;

(3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes;

(4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine;

(5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas;

(6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从硅基模具上揭下,得到微流控芯片的上层流体通道层;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on a silicon-based mold, use a vacuum pump to remove air bubbles, and place it at 60°C for heating under microcomputer temperature control. Stand on the stage for one and a half hours to fully cure the PDMS, and then peel off the fully cured PDMS from the silicon-based mold to obtain the upper fluid channel layer of the microfluidic chip;

(8)清洗玻璃底片:将玻璃底片依次经过,无水乙醇和去离子水的冲洗,然后用氮气吹干;所述玻璃底片即为微流控芯片的玻璃支撑层;(8) Cleaning the glass negative: pass the glass negative in sequence, rinse with absolute ethanol and deionized water, and then dry it with nitrogen; the glass negative is the glass support layer of the microfluidic chip;

(9)键合:将PDMS基体和玻璃底片一起放入等离子体机中进行清洗,清洗完毕后将PDMS基体和玻璃底片在1分钟内贴合且保证贴合面不留气泡,然后用重物压实,键合时间12h,微流控芯片制作完成。(9) Bonding: Put the PDMS substrate and the glass negative into the plasma machine for cleaning. After cleaning, the PDMS substrate and the glass negative are bonded within 1 minute and ensure that no air bubbles remain on the bonding surface, and then use a heavy object. After compaction, the bonding time was 12h, and the microfluidic chip was fabricated.

一种纳米颗粒跨血管输运高通量筛选的微流控芯片,包括流体通道层以及与流体通道层连接的流体渗透层,所述流体通道层的下端面和流体渗透层的上端面接触,所述流体通道层上设置有流体通道,所述流体通道设置在流体通道层的下端面,所述流体通道的一端设置有进样口,所述流体通道的另一端设置有出样口;所述流体渗透层上设置有渗出口,所述流体通道的中部通过间隙通道与渗出口连通。采用本技术方案,载药纳米颗粒溶液经压力施加装置从进样口注入流体通道,载药纳米颗粒溶液在流体通道流通的过程中,一部分载药纳米颗粒溶液从出样口流出,另一部分载药纳米颗粒溶液会通过间隙通道进入渗出口,收集渗出口内的载药纳米颗粒溶液。更换载药纳米颗粒溶液重复上述过程,对比收集溶液中的载药纳米颗粒含量,即可筛选出更适合穿透肿瘤血管壁的载药纳米颗粒。A microfluidic chip for high-throughput screening of nanoparticle transvascular transport, comprising a fluid channel layer and a fluid permeation layer connected with the fluid channel layer, wherein the lower end surface of the fluid channel layer is in contact with the upper end surface of the fluid permeation layer, The fluid channel layer is provided with a fluid channel, the fluid channel is arranged on the lower end face of the fluid channel layer, one end of the fluid channel is provided with a sample inlet, and the other end of the fluid channel is provided with a sample outlet; The fluid permeation layer is provided with a seepage outlet, and the middle part of the fluid channel is communicated with the seepage outlet through a gap channel. Using this technical solution, the drug-loaded nanoparticle solution is injected into the fluid channel from the injection port through the pressure applying device. During the circulation of the drug-loaded nanoparticle solution in the fluid channel, a part of the drug-loaded nanoparticle solution flows out from the sample outlet, and the other part of the drug-loaded nanoparticle solution flows out from the sample outlet. The drug nanoparticle solution will enter the exudate through the interstitial channel, and the drug-loaded nanoparticle solution in the exudate is collected. The above process is repeated by replacing the drug-loaded nanoparticle solution, and the drug-loaded nanoparticles that are more suitable for penetrating the tumor blood vessel wall can be screened by comparing the content of the drug-loaded nanoparticles in the collected solution.

所述微流控芯片采用模塑法进行制备,包括以下步骤:The microfluidic chip is prepared by a molding method, including the following steps:

(1)清洗单晶硅片:用去离子水冲洗,冲洗完毕后用氮气吹干,然后置于加热台上静置;(1) Cleaning the single crystal silicon wafer: rinse with deionized water, blow dry with nitrogen after rinsing, and then place it on a heating table to stand;

(2)涂胶:将正光刻胶涂于单晶硅片表面,并进行甩涂;(2) Glue coating: apply positive photoresist to the surface of the single crystal silicon wafer and spin it;

(3)烘焙:将甩涂后的单晶硅片置于加热台上静置;(3) Baking: put the spin-coated monocrystalline silicon wafer on the heating table and let it stand;

(4)曝光:将光刻掩膜板置于光刻胶表面,进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist for exposure;

(5)显影:对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: develop the exposed single crystal silicon wafer, then clean the surface of the single crystal silicon wafer with deionized water, and dry it with nitrogen;

(6)后烘:将单晶硅片置于加热台上静置,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: the single crystal silicon wafer is placed on a heating table to stand, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于流体通道层的硅基模具上,除去气泡,置于加热台上静置使PDMS完全固化,再将完全固化后的PDMS从流体通道层的硅基模具上揭下,得到流体通道层;然后再以PDMS和固化剂按照体积比15:1的比例混合并且搅拌均匀后,倾倒于流体渗透层的硅基模具上,除去气泡,置于加热台上静置,在PDMS未完全固化时,将其从流体渗透层的硅基模具上揭下,然后迅速与流体通道层对合夹紧;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on the silicon-based mold of the fluid channel layer, remove air bubbles, and place it on a heating table. The PDMS is completely cured, and the fully cured PDMS is removed from the silicon-based mold of the fluid channel layer to obtain the fluid channel layer; then PDMS and the curing agent are mixed in a volume ratio of 15:1 and stirred evenly, Pour it on the silicon-based mold of the fluid-permeable layer, remove air bubbles, and place it on a heating table. When the PDMS is not fully cured, remove it from the silicon-based mold of the fluid-permeable layer, and then quickly align it with the fluid channel layer. close clamping;

(8)热键合:将对合夹紧的流体通道层和流体渗透层置于加热台上静置。(8) Thermal bonding: The fluid channel layer and fluid permeation layer that are clamped together are placed on a heating table and left to stand.

进一步的,所述微流控芯片的制备方法包括以下步骤:Further, the preparation method of the microfluidic chip includes the following steps:

(1)首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) First rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C for 15 minutes;

(2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s;

(3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes;

(4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine;

(5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas;

(6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于流体通道层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从流体通道层的硅基模具上揭下,得到微流控芯片的上层;然后再以PDMS和固化剂按照体积比15:1的比例混合并且搅拌均匀后,倾倒于流体渗透层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置30分钟,在PDMS未完全固化时,将其从流体渗透层的硅基模具上揭下,然后迅速与流体通道层对合夹紧。(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on the silicon-based mold of the fluid channel layer, remove air bubbles with a vacuum pump, and place it at 60°C The microcomputer temperature-controlled heating table was left standing for one and a half hours to fully cure the PDMS, and then the fully cured PDMS was removed from the silicon-based mold of the fluid channel layer to obtain the upper layer of the microfluidic chip; then PDMS and curing agent were used. After mixing in a volume ratio of 15:1 and stirring evenly, pour it onto the silicon-based mold of the fluid permeable layer, remove air bubbles with a vacuum pump, and place it on a microcomputer-controlled heating table at 60°C for 30 minutes. When cured, it was peeled from the silicon-based mold of the fluid permeable layer and then quickly clamped against the fluid channel layer.

(8)热键合:将对合夹紧的流体通道层和流体渗透层置于80℃微电脑控温加热台上静置10小时,微流控芯片制作完成。(8) Thermal bonding: The fluid channel layer and fluid permeation layer that are clamped together are placed on a microcomputer temperature-controlled heating table at 80°C for 10 hours, and the fabrication of the microfluidic chip is completed.

本发明与现有技术相比具有以下有益效果:所述微流控芯片能够用来筛选适合穿透血管壁的载药纳米颗粒,其筛选的准确率高,可以满足现代载药纳米颗粒穿透血管屏障医学研究的需要,整个筛选过程中的耗材只有载药纳米颗粒溶液,所以药物筛选的过程成本低,所述微流控芯片的制备工艺也相对简单,制作成本低。Compared with the prior art, the present invention has the following beneficial effects: the microfluidic chip can be used to screen drug-loaded nanoparticles suitable for penetrating the blood vessel wall, and the screening accuracy is high, which can meet the requirements of modern drug-loaded nanoparticles to penetrate To meet the needs of vascular barrier medical research, the consumables in the entire screening process are only drug-loaded nanoparticle solution, so the cost of the drug screening process is low, and the preparation process of the microfluidic chip is relatively simple and low in production cost.

由此可见,本发明与现有技术相比,具有突出的实质性特点和显著的进步,其实施的有益效果也是显而易见的。It can be seen that, compared with the prior art, the present invention has outstanding substantive features and significant progress, and the beneficial effects of its implementation are also obvious.

附图说明Description of drawings

图1为实施例1中的纳米颗粒跨血管输运高通量筛选的微流控芯片的平面结构示意图;1 is a schematic plan view of a microfluidic chip for high-throughput screening of nanoparticle transvascular transport in Example 1;

图2为实施例1中的纳米颗粒跨血管输运高通量筛选的微流控芯片的立体结构示意图;2 is a schematic three-dimensional structure diagram of a microfluidic chip for high-throughput screening of nanoparticle transvascular transport in Example 1;

图3为图2中A部的放大图;Fig. 3 is the enlarged view of A part in Fig. 2;

图4为实施例2中的纳米颗粒跨血管输运高通量筛选的微流控芯片的结构示意图。FIG. 4 is a schematic structural diagram of a microfluidic chip for high-throughput screening of nanoparticle transvascular transport in Example 2. FIG.

图中:1-进样口,2-流体通道Ⅰ,3-出样口,4-渗出口,5-间隙通道,6-流体通道Ⅱ,7-流体通道层,8-玻璃支撑层,9-流体渗透层,10-流体通道。In the figure: 1-injection port, 2-fluid channel I, 3-sample outlet, 4-exudation port, 5-gap channel, 6-fluid channel II, 7-fluid channel layer, 8-glass support layer, 9 - fluid permeable layer, 10 - fluid channel.

具体实施方式Detailed ways

为能清楚说明本方案的技术特点,下面通过具体实施方式,并结合其附图,对本方案进行阐述。In order to clearly illustrate the technical features of the present solution, the present solution will be described below through specific embodiments and in conjunction with the accompanying drawings.

实施例1Example 1

如图1-3所示,一种纳米颗粒跨血管输运高通量筛选的微流控芯片,其特征是:包括流体通道层7以及与流体通道层7连接的玻璃支撑层8,所述流体通道层7的下端面与玻璃支撑层8的上端面接触,所述流体通道层7内设置有流体通道Ⅰ2和流体通道Ⅱ6,所述流体通道Ⅰ2的一端设置有进样口1,所述流体通道Ⅰ2的另一端设置有出样口3,所述流体通道Ⅰ2的中部通过间隙通道5与流体通道Ⅱ6连通,所述流体通道Ⅱ6的一端设置有渗出口4。载药纳米颗粒溶液经压力施加装置从进样口1注入流体通道Ⅰ2,在经过间隙通道5时,一部分载药纳米颗粒溶液会通过流体通道Ⅰ2从出样口3流出,另一部分载药纳米颗粒溶液通过间隙通道5进入流体通道Ⅱ6并从渗出口4流出,收集渗出口4流出的样品溶液。更换载药纳米颗粒溶液重复上述过程,对比收集样品溶液中的载药纳米颗粒含量,即可筛选出更适合穿透肿瘤血管壁的载药纳米颗粒。所述间隙通道5的尺寸与肿瘤血管壁缺少基膜和粘附蛋白导致其血管壁上出现的孔洞的尺寸相适配。所述出样口3和渗出口4均可通过导管导出。As shown in Figures 1-3, a microfluidic chip for high-throughput screening of nanoparticle transvascular transport is characterized by comprising a fluid channel layer 7 and a glass support layer 8 connected to the fluid channel layer 7. The said The lower end surface of the fluid channel layer 7 is in contact with the upper end surface of the glass support layer 8, the fluid channel layer 7 is provided with a fluid channel I2 and a fluid channel II6, and one end of the fluid channel I2 is provided with a sample inlet 1, and the The other end of the fluid channel I2 is provided with a sample outlet 3, the middle of the fluid channel I2 is communicated with the fluid channel II6 through the gap channel 5, and one end of the fluid channel II6 is provided with a seepage port 4. The drug-loaded nanoparticle solution is injected into the fluid channel I2 from the injection port 1 through the pressure applying device. When passing through the gap channel 5, a part of the drug-loaded nanoparticle solution will flow out from the sample outlet 3 through the fluid channel I2, and another part of the drug-loaded nanoparticle solution will flow out from the sample outlet 3. The solution enters the fluid channel II6 through the gap channel 5 and flows out from the permeation port 4, and the sample solution flowing out of the permeation port 4 is collected. The above process is repeated by replacing the drug-loaded nanoparticle solution, and the drug-loaded nanoparticles that are more suitable for penetrating the tumor blood vessel wall can be screened by comparing the content of the drug-loaded nanoparticles in the collected sample solution. The size of the interstitial channel 5 is adapted to the size of the hole in the vessel wall caused by the lack of basement membrane and adhesion proteins in the tumor vessel wall. Both the sample outlet 3 and the seepage outlet 4 can be led out through a conduit.

所述微流控芯片采用模塑法进行制备,具体方法为:The microfluidic chip is prepared by a molding method, and the specific method is:

(1)清洗单晶硅片:首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) Cleaning the single crystal silicon wafer: first rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C Set for 15 minutes;

(2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s;

(3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes;

(4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine;

(5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas;

(6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从硅基模具上揭下,得到微流控芯片的上层流体通道层;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on a silicon-based mold, use a vacuum pump to remove air bubbles, and place it at 60°C for heating under microcomputer temperature control. Stand on the stage for one and a half hours to fully cure the PDMS, and then peel off the fully cured PDMS from the silicon-based mold to obtain the upper fluid channel layer of the microfluidic chip;

(8)清洗玻璃底片:将玻璃底片依次经过,无水乙醇和去离子水的冲洗,然后用氮气吹干;所述玻璃底片即为微流控芯片的玻璃支撑层;(8) Cleaning the glass negative: pass the glass negative in sequence, rinse with absolute ethanol and deionized water, and then dry it with nitrogen; the glass negative is the glass support layer of the microfluidic chip;

(9)键合:将PDMS基体和玻璃底片一起放入等离子体机中进行清洗,清洗完毕后将PDMS基体和玻璃底片在1分钟内贴合且保证贴合面不留气泡,然后用重物压实,键合时间12h,微流控芯片制作完成。(9) Bonding: Put the PDMS substrate and the glass negative into the plasma machine for cleaning. After cleaning, the PDMS substrate and the glass negative are bonded within 1 minute and ensure that no air bubbles remain on the bonding surface, and then use a heavy object. After compaction, the bonding time was 12h, and the microfluidic chip was fabricated.

所述PDMS为聚二甲基硅氮烷聚合物,PDMS具有良好的生物相容性和光学特性。所述PDMS和固化剂采用美国道康宁SYLGARD184硅橡胶。The PDMS is a polydimethylsilazane polymer, and the PDMS has good biocompatibility and optical properties. The PDMS and curing agent are Dow Corning SYLGARD184 silicone rubber.

实施例2Example 2

一种纳米颗粒跨血管输运高通量筛选的微流控芯片,包括流体通道层7以及与流体通道层连接7的流体渗透层9,所述流体通道层7的下端面和流体渗透层9的上端面接触,所述流体通道层7上设置有流体通道10,所述流体通道10设置在流体通道层7的下端面,所述流体通道10的一端设置有进样口1,所述流体通道10的另一端设置有出样口3;所述流体渗透层9上设置有渗出口4,所述流体通道10的中部通过间隙通道5与渗出口4连通。载药纳米颗粒溶液经压力施加装置从进样口1注入流体通道10,载药纳米颗粒溶液在流体通道10流通的过程中,一部分载药纳米颗粒溶液从出样口3流出,另一部分载药纳米颗粒溶液会通过间隙通道5进入渗出口4,收集渗出口内的载药纳米颗粒溶液。更换载药纳米颗粒溶液重复上述过程,对比收集溶液中的载药纳米颗粒含量,即可筛选出更适合穿透肿瘤血管壁的载药纳米颗粒。所述间隙通道5的尺寸可与肿瘤血管壁缺少基膜和粘附蛋白导致其血管壁上出现的孔洞的尺寸相适配。所述出样口3和渗出口4均可通过导管导出。A microfluidic chip for high-throughput screening of nanoparticle transvascular transport, comprising a fluid channel layer 7 and a fluid permeation layer 9 connected to the fluid channel layer 7, the lower end face of the fluid channel layer 7 and the fluid permeation layer 9 The fluid channel layer 7 is provided with a fluid channel 10, the fluid channel 10 is arranged on the lower end surface of the fluid channel layer 7, and one end of the fluid channel 10 is provided with a sample inlet 1, and the fluid The other end of the channel 10 is provided with a sample outlet 3 ; the fluid permeable layer 9 is provided with a seepage outlet 4 , and the middle of the fluid channel 10 communicates with the seepage outlet 4 through a gap channel 5 . The drug-loaded nanoparticle solution is injected into the fluid channel 10 from the injection port 1 through the pressure applying device. During the flow of the drug-loaded nanoparticle solution in the fluid channel 10, a part of the drug-loaded nanoparticle solution flows out from the sample outlet 3, and the other part is drug-loaded. The nanoparticle solution will enter the infiltration port 4 through the interstitial channel 5, and the drug-loaded nanoparticle solution in the infiltration port will be collected. The above process is repeated by replacing the drug-loaded nanoparticle solution, and the drug-loaded nanoparticles that are more suitable for penetrating the tumor blood vessel wall can be screened by comparing the content of the drug-loaded nanoparticles in the collected solution. The size of the interstitial channel 5 can be adapted to the size of the hole in the vessel wall caused by the lack of basement membrane and adhesion proteins in the tumor vessel wall. Both the sample outlet 3 and the seepage outlet 4 can be led out through a conduit.

所述微流控芯片采用模塑法进行制备,具体方法为:The microfluidic chip is prepared by a molding method, and the specific method is:

(1)首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) First rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C for 15 minutes;

(2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s;

(3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes;

(4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine;

(5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas;

(6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained;

(7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于流体通道层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从流体通道层的硅基模具上揭下,得到微流控芯片的上层;然后再以PDMS和固化剂按照体积比15:1的比例混合并且搅拌均匀后,倾倒于流体渗透层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置30分钟,在PDMS未完全固化时,将其从流体渗透层的硅基模具上揭下,然后迅速与流体通道层对合夹紧。(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on the silicon-based mold of the fluid channel layer, remove air bubbles with a vacuum pump, and place it at 60°C The microcomputer temperature-controlled heating table was left standing for one and a half hours to fully cure the PDMS, and then the fully cured PDMS was removed from the silicon-based mold of the fluid channel layer to obtain the upper layer of the microfluidic chip; then PDMS and curing agent were used. After mixing in a volume ratio of 15:1 and stirring evenly, pour it onto the silicon-based mold of the fluid permeable layer, remove air bubbles with a vacuum pump, and place it on a microcomputer-controlled heating table at 60°C for 30 minutes. When cured, it was peeled from the silicon-based mold of the fluid permeable layer and then quickly clamped against the fluid channel layer.

(8)热键合:将对合夹紧的流体通道层和流体渗透层置于80℃微电脑控温加热台上静置10小时,微流控芯片制作完成。(8) Thermal bonding: The fluid channel layer and fluid permeation layer that are clamped together are placed on a microcomputer temperature-controlled heating table at 80°C for 10 hours, and the fabrication of the microfluidic chip is completed.

所述PDMS为聚二甲基硅氮烷聚合物,PDMS具有良好的生物相容性和光学特性。所述PDMS和固化剂采用美国道康宁SYLGARD184硅橡胶。The PDMS is a polydimethylsilazane polymer, and the PDMS has good biocompatibility and optical properties. The PDMS and curing agent are Dow Corning SYLGARD184 silicone rubber.

本发明中未经描述的技术特征可以通过或采用现有技术实现,在此不再赘述,当然,上述说明并非是对本发明的限制,本发明也并不仅限于上述实施方式,本领域的普通技术人员在本发明的实质范围内所做出的变化、改型、添加或替换,也应属于本发明的保护范围。The technical features that are not described in the present invention can be realized by or using the existing technology, and will not be repeated here. Of course, the above description is not a limitation of the present invention, and the present invention is not limited to the above-mentioned embodiments. Changes, modifications, additions or substitutions made by persons within the essential scope of the present invention shall also belong to the protection scope of the present invention.

Claims (8)

1.一种纳米颗粒跨血管输运高通量筛选的微流控芯片,其特征是:包括流体通道层以及与流体通道层连接的玻璃支撑层,所述流体通道层的下端面与玻璃支撑层的上端面接触,所述流体通道层内设置有流体通道Ⅰ和流体通道Ⅱ,所述流体通道Ⅰ的一端设置有进样口,所述流体通道Ⅰ的另一端设置有出样口,所述流体通道Ⅰ的中部通过间隙通道与流体通道Ⅱ连通,所述流体通道Ⅱ的一端设置有渗出口。1. a kind of microfluidic chip of nanoparticle transvascular transport high-throughput screening, it is characterized in that: comprise fluid channel layer and the glass support layer that is connected with fluid channel layer, the lower end face of described fluid channel layer and glass support layer The upper end surface of the fluid channel layer is in contact with the upper end surface of the fluid channel layer. The fluid channel layer is provided with a fluid channel I and a fluid channel II. One end of the fluid channel I is provided with a sample inlet, and the other end of the fluid channel I is provided with a sample outlet. The middle part of the fluid channel I is communicated with the fluid channel II through a gap channel, and one end of the fluid channel II is provided with a seepage outlet. 2.一种如权利要求1中的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:所述微流控芯片采用模塑法进行制备。2 . A method for preparing a microfluidic chip for high-throughput screening of nanoparticle transvascular transport as claimed in claim 1 , wherein the microfluidic chip is prepared by a molding method. 3 . 3.根据权利要求2所述的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:包括以下步骤:3. the preparation method of the microfluidic chip of nanoparticle transvascular transport high-throughput screening according to claim 2, is characterized in that: comprises the following steps: (1)清洗单晶硅片:用去离子水冲洗,完毕后用氮气吹干,然后置于加热台上静置;(1) Cleaning the single crystal silicon wafer: rinse with deionized water, blow dry with nitrogen after completion, and then place it on a heating table to stand; (2)涂胶:将正光刻胶涂于单晶硅片表面,并进行甩涂;(2) Glue coating: apply positive photoresist to the surface of the single crystal silicon wafer and spin it; (3)烘焙:将甩涂后的单晶硅片置于加热台上静置;(3) Baking: put the spin-coated monocrystalline silicon wafer on the heating table and let it stand; (4)曝光:将光刻掩膜板置于光刻胶表面,进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist for exposure; (5)显影:对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: develop the exposed single crystal silicon wafer, then clean the surface of the single crystal silicon wafer with deionized water, and dry it with nitrogen; (6)后烘:将单晶硅片置于加热台上静置,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: the single crystal silicon wafer is placed on a heating table to stand, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained; (7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于硅基模具上,除去气泡,置于加热台上静置使PDMS完全固化,再将完全固化后的PDMS从硅基模具上揭下,得到PDMS基体,即为微流控芯片的流体通道层;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on a silicon-based mold, remove air bubbles, and place it on a heating table to allow PDMS to completely cure. , and then peel off the fully cured PDMS from the silicon-based mold to obtain the PDMS matrix, which is the fluid channel layer of the microfluidic chip; (8)清洗玻璃底片:将玻璃底片依次经过无水乙醇和去离子水的冲洗,然后用氮气吹干;(8) Cleaning the glass negative film: Rinse the glass negative film with absolute ethanol and deionized water in sequence, and then dry it with nitrogen; (9)键合:将PDMS基体和玻璃底片一起放入等离子体机中进行清洗,清洗完毕后将PDMS基体和玻璃底片贴合且保证贴合面不留气泡,然后用重物压实。(9) Bonding: Put the PDMS substrate and the glass negative into the plasma machine for cleaning. After cleaning, the PDMS substrate and the glass negative are bonded together to ensure that no air bubbles remain on the bonding surface, and then compacted with a heavy object. 4.根据权利要求3所述的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:包括以下步骤:4. the preparation method of the microfluidic chip of nanoparticle transvascular transport high-throughput screening according to claim 3, is characterized in that: comprises the following steps: (1)清洗单晶硅片:首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) Cleaning the single crystal silicon wafer: first rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C Set for 15 minutes; (2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s; (3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes; (4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine; (5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas; (6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained; (7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从硅基模具上揭下,得到微流控芯片的上层流体通道层;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on a silicon-based mold, use a vacuum pump to remove air bubbles, and place it at 60°C for heating under microcomputer temperature control. Stand on the stage for one and a half hours to fully cure the PDMS, and then peel off the fully cured PDMS from the silicon-based mold to obtain the upper fluid channel layer of the microfluidic chip; (8)清洗玻璃底片:将玻璃底片依次经过,无水乙醇和去离子水的冲洗,然后用氮气吹干;(8) Cleaning the glass negative: pass the glass negative in sequence, rinse with absolute ethanol and deionized water, and then dry it with nitrogen; (9)键合:将PDMS基体和玻璃底片一起放入等离子体机中进行清洗,清洗完毕后将PDMS基体和玻璃底片在1分钟内贴合且保证贴合面不留气泡,然后用重物压实,键合时间12h,微流控芯片制作完成。(9) Bonding: Put the PDMS substrate and the glass negative into the plasma machine for cleaning. After cleaning, attach the PDMS substrate and the glass negative within 1 minute and ensure that no air bubbles remain on the bonding surface, and then use a heavy object. After compaction, the bonding time was 12h, and the microfluidic chip was fabricated. 5.一种纳米颗粒跨血管输运高通量筛选的微流控芯片,其特征是:包括流体通道层以及与流体通道层连接的流体渗透层,所述流体通道层的下端面和流体渗透层的上端面接触,所述流体通道层上设置有流体通道,所述流体通道设置在流体通道层的下端面,所述流体通道的一端设置有进样口,所述流体通道的另一端设置有出样口;所述流体渗透层上设置有渗出口,所述流体通道的中部通过间隙通道与渗出口连通。5. a microfluidic chip for high-throughput screening of nanoparticle transvascular transport, is characterized in that: comprising a fluid channel layer and a fluid permeation layer connected with the fluid channel layer, the lower end face of the fluid channel layer and the fluid permeation layer The upper end surface of the layer is in contact, the fluid channel layer is provided with a fluid channel, the fluid channel is arranged on the lower end surface of the fluid channel layer, one end of the fluid channel is provided with a sample inlet, and the other end of the fluid channel is provided with There is a sample outlet; the fluid permeation layer is provided with a seepage outlet, and the middle part of the fluid channel is communicated with the seepage outlet through a gap channel. 6.一种如权利要求5中的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:所述微流控芯片采用模塑法进行制备。6 . A method for preparing a microfluidic chip for high-throughput screening of nanoparticle transvascular transport as claimed in claim 5 , wherein the microfluidic chip is prepared by a molding method. 7 . 7.根据权利要求6所述的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:包括以下步骤:7. the preparation method of the microfluidic chip of nanoparticle transvascular transport high-throughput screening according to claim 6, is characterized in that: comprises the following steps: (1)清洗单晶硅片:用去离子水冲洗,冲洗完毕后用氮气吹干,然后置于加热台上静置;(1) Cleaning the single crystal silicon wafer: rinse with deionized water, blow dry with nitrogen after rinsing, and then place it on a heating table to stand; (2)涂胶:将正光刻胶涂于单晶硅片表面,并进行甩涂;(2) Glue coating: apply positive photoresist to the surface of the single crystal silicon wafer and spin it; (3)烘焙:将甩涂后的单晶硅片置于加热台上静置;(3) Baking: put the spin-coated monocrystalline silicon wafer on the heating table and let it stand; (4)曝光:将光刻掩膜板置于光刻胶表面,进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist for exposure; (5)显影:对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: develop the exposed single crystal silicon wafer, then clean the surface of the single crystal silicon wafer with deionized water, and dry it with nitrogen; (6)后烘:将单晶硅片置于加热台上静置,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: the single crystal silicon wafer is placed on a heating table to stand, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained; (7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于流体通道层的硅基模具上,除去气泡,置于加热台上静置使PDMS完全固化,再将完全固化后的PDMS从流体通道层的硅基模具上揭下,得到流体通道层;然后再以PDMS和固化剂按照体积比15:1的比例混合并且搅拌均匀后,倾倒于流体渗透层的硅基模具上,除去气泡,置于加热台上静置,在PDMS未完全固化时,将其从流体渗透层的硅基模具上揭下,然后迅速与流体通道层对合夹紧;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on the silicon-based mold of the fluid channel layer, remove air bubbles, and place it on a heating table. The PDMS is completely cured, and the fully cured PDMS is removed from the silicon-based mold of the fluid channel layer to obtain the fluid channel layer; then PDMS and the curing agent are mixed in a volume ratio of 15:1 and stirred evenly, Pour it on the silicon-based mold of the fluid-permeable layer, remove air bubbles, and place it on a heating table. When the PDMS is not fully cured, remove it from the silicon-based mold of the fluid-permeable layer, and then quickly align it with the fluid channel layer. close clamping; (8)热键合:将对合夹紧的流体通道层和流体渗透层置于加热台上静置。(8) Thermal bonding: The fluid channel layer and fluid permeation layer that are clamped together are placed on a heating table and left to stand. 8.根据权利要求7所述的纳米颗粒跨血管输运高通量筛选的微流控芯片的制备方法,其特征是:包括以下步骤:8. the preparation method of the microfluidic chip of nanoparticle transvascular transport high-throughput screening according to claim 7, is characterized in that: comprises the following steps: (1)首先用去离子水冲洗,然后经无水乙醇冲洗,再用去离子水进行冲洗,完毕后用氮气吹干,然后置于90℃微电脑控温加热台上静置15分钟;(1) First rinse with deionized water, then rinse with absolute ethanol, then rinse with deionized water, dry with nitrogen after completion, and then place it on a microcomputer temperature-controlled heating table at 90 °C for 15 minutes; (2)涂胶:将正光刻胶涂于单晶硅片表面,利用匀胶机旋涂仪进行甩涂,转速为700r/min,甩涂时间为20s;(2) Gluing: apply positive photoresist on the surface of the single crystal silicon wafer, and use a spin coater of a glue spinner for spin coating, the rotation speed is 700r/min, and the spin coating time is 20s; (3)烘焙:将甩涂后的单晶硅片置于60℃微电脑控温加热台上静置15分钟;(3) Baking: Place the spin-coated monocrystalline silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 15 minutes; (4)曝光:将光刻掩膜板置于光刻胶表面,用曝光机进行曝光;(4) Exposure: place the photolithography mask on the surface of the photoresist and expose it with an exposure machine; (5)显影:利用显影液对曝光后的单晶硅片显影,然后用去离子水清洗单晶硅片的表面,氮气吹干;(5) Development: Use developer to develop the exposed single crystal silicon wafer, then use deionized water to clean the surface of the single crystal silicon wafer, and dry it with nitrogen gas; (6)后烘:将单晶硅片置于60℃微电脑控温加热台上静置40分钟,使硅片表面的光刻胶固化完全,得到硅基模具;(6) Post-baking: place the single crystal silicon wafer on a microcomputer temperature-controlled heating table at 60°C for 40 minutes, so that the photoresist on the surface of the silicon wafer is completely cured, and a silicon-based mold is obtained; (7)构建微流控芯片:先以PDMS和固化剂按照体积比10:1的比例混合并且搅拌均匀后,倾倒于流体通道层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置一个半小时使PDMS完全固化,再将完全固化后的PDMS从流体通道层的硅基模具上揭下,得到微流控芯片的上层;然后再以PDMS和固化剂按照体积比15:1的比例混合并且搅拌均匀后,倾倒于流体渗透层的硅基模具上,用抽真空泵除去气泡,置于60℃微电脑控温加热台上静置30分钟,在PDMS未完全固化时,将其从流体渗透层的硅基模具上揭下,然后迅速与流体通道层对合夹紧;(7) Constructing a microfluidic chip: First mix PDMS and curing agent in a volume ratio of 10:1 and stir evenly, pour it on the silicon-based mold of the fluid channel layer, remove air bubbles with a vacuum pump, and place it at 60°C The microcomputer temperature-controlled heating table was left standing for one and a half hours to fully cure the PDMS, and then the fully cured PDMS was removed from the silicon-based mold of the fluid channel layer to obtain the upper layer of the microfluidic chip; then PDMS and curing agent were used. After mixing in a volume ratio of 15:1 and stirring evenly, pour it onto the silicon-based mold of the fluid permeable layer, remove air bubbles with a vacuum pump, and place it on a microcomputer-controlled heating table at 60°C for 30 minutes. When curing, peel it off the silicon-based mold of the fluid permeable layer, and then quickly clamp it with the fluid channel layer; (8)热键合:将对合夹紧的流体通道层和流体渗透层置于80℃微电脑控温加热台上静置10小时,微流控芯片制作完成。(8) Thermal bonding: The fluid channel layer and fluid permeation layer that are clamped together are placed on a microcomputer temperature-controlled heating table at 80°C for 10 hours, and the fabrication of the microfluidic chip is completed.
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