WO2023093404A1

WO2023093404A1 - Microparticle self-entrainment type microfluidic chip and manufacturing method therefor, and microparticle self-dispersion method

Info

Publication number: WO2023093404A1
Application number: PCT/CN2022/126786
Authority: WO
Inventors: 马波; 刘沣仪; 徐健
Original assignee: 中国科学院青岛生物能源与过程研究所
Priority date: 2021-11-23
Filing date: 2022-10-21
Publication date: 2023-06-01
Also published as: CN114149893A

Abstract

A microparticle self-entrainment type microfluidic chip and a manufacturing method therefor, and a microparticle self-dispersion method. The microparticle self-entrainment type microfluidic chip is provided with a microparticle sample inlet hole and a microparticle sample outlet hole. A microfluidic channel is provided inside the microparticle self-entrainment type microfluidic chip. The microfluidic channel comprises a main channel (2), a side channel (3), and an entrainment structure (4). An upper end of the side channel (3) is in communication connection with the upstream of the main channel (2). The microfluid channel is provided with a microparticle blocking structure (7). A lower end of the side channel (3) is in communication connection with the downstream of the main channel (2). The entrainment structure (4) is located at the position where the lower end of the side channel (3) is in communication connection with the downstream of the main channel (2). The microparticle sample inlet hole is in communication connection with a microparticle phase inlet of the main channel. The microparticle sample outlet hole is in communication connection with a microparticle outlet of the entrainment structure (4). The chip can disperse microparticles which are closely arranged. Separation oil and a driving device are not additionally required. The apparatus is simplified, and the chip can be recycled, so that the cost is reduced. The microparticle self-dispersion method using the chip is suitable for self-entrainment of microparticles of various sizes, and the operation is simple and convenient. The manufacturing method for the chip is high in yield.

Description

微粒自夹流式微流控芯片及其制造方法和微粒自分散方法Particle self-clamping microfluidic chip, manufacturing method thereof, and particle self-dispersing method

技术领域technical field

本发明涉及微流控芯片技术领域，尤其涉及将微粒通过自夹流形成有一定间隔的单微粒顺序流的微粒自夹流式微流控芯片及其制造方法和微粒自分散方法。The invention relates to the technical field of microfluidic chips, in particular to a particle self-encapsulating microfluidic chip in which particles pass through the self-entrapping flow to form a sequential flow of single particles with a certain interval, a manufacturing method thereof, and a particle self-dispersing method.

背景技术Background technique

众所周知，随着现代科学技术的进步,人类对应用领域的研究已经迈入了微观尺度范围,微纳米显微操作技术的研究越来越受到国内外学者的广泛关注。微粒的分离是微流体操纵中的关键技术,广泛应用在生物工程、医药、纳米自组装、化学分析、材料性能评定等各个领域，包括对单细胞、蛋白、纳米材料、液滴等的研究。比如，对单细胞分析可以分析细胞个体间差异包括细胞尺寸、生长速度、化学组成(磷脂、蛋白、代谢物、DNA/RNA)，以及细胞间差异产生的原因、机理。其研究内容涉及到肿瘤生物学、干细胞、微生物学、神经***学和免疫学等领域。基于液滴的微流控芯片技术能够与众多的化学和生物试剂和“电子控制”相互兼容，并且具有良好的可编程和构建性。在大多数应用中，均匀的液滴可以确保恒定、可控和可预测的结果。基于液滴的应用主要集中在液滴操纵、液滴数字PCR(芯片法)，包括dd PCR(微滴式数字芯片法)，以及液滴分选、液滴检测等领域。As we all know, with the advancement of modern science and technology, human research on the application field has entered the micro-scale range, and the research on micro-nano micromanipulation technology has attracted more and more attention from scholars at home and abroad. Particle separation is a key technology in microfluidic manipulation, and it is widely used in various fields such as bioengineering, medicine, nanometer self-assembly, chemical analysis, material performance evaluation, etc., including the study of single cells, proteins, nanomaterials, droplets, etc. For example, single-cell analysis can analyze the differences between individual cells, including cell size, growth rate, chemical composition (phospholipids, proteins, metabolites, DNA/RNA), and the causes and mechanisms of differences between cells. Its research content involves the fields of tumor biology, stem cells, microbiology, neurology and immunology. Droplet-based microfluidic chip technology is compatible with numerous chemical and biological reagents and "electronic control", and has good programmability and constructability. A uniform droplet ensures constant, controllable and predictable results in most applications. Droplet-based applications mainly focus on droplet manipulation, droplet digital PCR (chip method), including ddPCR (droplet digital chip method), droplet sorting, droplet detection and other fields.

随着对基于微流控芯片领域兴趣的增长，更多控制、操纵和功能化单细胞、蛋白、纳米材料液滴等微粒的技术得以开发。其中，微粒的直径可以为微米至纳米量级。利用微流控技术可实现单个细胞、蛋白质、核酸等的封装以及微/纳米颗粒的合成。要对单微粒进行在线操作，首先要将微粒注入芯片。但是目前的微粒再注入技术普遍需要另外添加一路或两路油相作为鞘流液，将紧密排列的微粒分散开来。因此消耗大量夹流液，消耗大量试剂，大幅度增加了实验成本；且夹流液需要蠕动泵等额外的泵驱动设备，导致设备结构复杂，无法做到小型化与便携化，进一步增加实验成本。As interest in the field of microfluidic-based chips grows, more techniques for controlling, manipulating, and functionalizing particles such as single cells, proteins, and nanomaterial droplets are being developed. Wherein, the diameter of the particles may be in the order of micrometers to nanometers. The encapsulation of single cells, proteins, nucleic acids, etc., and the synthesis of micro/nanoparticles can be realized by using microfluidic technology. To perform in-line manipulation of single particles, the particles are first injected into the chip. However, the current particle reinjection technology generally needs to add one or two additional oil phases as the sheath fluid to disperse the closely arranged particles. Therefore, a large amount of entrained liquid and reagents are consumed, which greatly increases the cost of the experiment; and the entrained liquid requires additional pump-driven equipment such as a peristaltic pump, resulting in a complex structure of the equipment, which cannot be miniaturized and portable, and further increases the cost of the experiment. .

发明内容Contents of the invention

针对上述问题，本发明的目的是提供微粒自夹流式微流控芯片及其制造方法和微粒自分散方法，通过所述微粒自夹流式微流控芯片，能够将紧密排列的微粒分散，形成有间隔的单微粒顺序流，无需另外的夹流液与驱动设备，降低了微粒再注入的成本，无需蠕动泵等设备，实现了装置的简化。In response to the above problems, the object of the present invention is to provide a microfluidic chip with self-encapsulation of particles, a manufacturing method thereof, and a method for self-dispersion of particles. Through the microfluidic chip of self-entrapment of particles, closely arranged particles can be dispersed to form a The sequential flow of single particles at intervals does not require additional entrained liquid and driving equipment, which reduces the cost of particle reinjection, and does not require peristaltic pumps and other equipment, which simplifies the device.

为实现上述目的，本发明采取以下技术方案：To achieve the above object, the present invention takes the following technical solutions:

首先，本发明提供微粒自夹流式微流控芯片，所述微粒自夹流式微流控芯片内设置至少一条微流体通道，所述微流体通道包括主通道、至少一段侧通道和夹流结构；First of all, the present invention provides a microfluidic chip with self-entrapment of microparticles, wherein at least one microfluidic channel is provided in the microfluidic chip of self-entrapment microfluidics, and the microfluidic channel includes a main channel, at least one section of side channel and an entrainment structure;

所述主通道上端设置微粒相入口，所述夹流结构下端设置微粒出口；The upper end of the main channel is provided with a particle phase inlet, and the lower end of the entrainment structure is provided with a particle outlet;

所述侧通道上端与所述主通道上游连通连接，所述微流体通道设置阻止微粒进入所述侧通道的微粒阻截结构，所述侧通道下端与所述主通道下游连通连接，所述夹流结构位于所述侧通道下端与所述主通道下游连通连接处；The upper end of the side channel communicates with the main channel upstream, the microfluidic channel is provided with a particle blocking structure that prevents particles from entering the side channel, the lower end of the side channel communicates with the main channel downstream, and the entrainment The structure is located at the connection between the lower end of the side channel and the downstream of the main channel;

所述微粒自夹流式微流控芯片上设置微粒进样孔和微粒出样孔，所述微粒进样孔与所述微粒相入口连通连接，所述微粒出样孔与所述微粒出口连通连接。A microparticle sampling hole and a microparticle sampling hole are provided on the microparticle self-clamping microfluidic chip, the microparticle sampling hole communicates with the microparticle phase inlet, and the microparticle sampling hole communicates with the microparticle outlet .

优选的，所述侧通道与所述主通道连接处高度高于所述侧通道高度构成高度差值；所述微粒阻截结构为设置在所述侧通道与所述主通道连接处的微粒阻拦结构，或为所述侧通道的宽度小于所述液滴直径所构成的尺寸差结构，或为所述高度差值小于所述液滴直径所构成的尺寸差结构，或为其中二者或三者结构的组合。Preferably, the height of the connection between the side channel and the main channel is higher than the height of the side channel to form a height difference; the particle blocking structure is a particle blocking structure arranged at the connection between the side channel and the main channel , or the size difference structure formed by the width of the side channel being smaller than the droplet diameter, or the size difference structure formed by the height difference being smaller than the droplet diameter, or two or three of them combination of structures.

优选的，所述微粒阻截结构为设置在所述侧通道与所述主通道连接处的微粒阻拦结构时，所述微粒阻拦结构为微筛阵列结构。Preferably, when the particle blocking structure is a particle blocking structure arranged at the connection between the side channel and the main channel, the particle blocking structure is a microsieve array structure.

优选的，在所述微粒进样孔处设置气泡阻拦结构。Preferably, a bubble blocking structure is provided at the microparticle sampling hole.

优选的，在所述气泡阻拦结构为微筛阵列结构。Preferably, the bubble blocking structure is a microsieve array structure.

优选的，在所述主通道下游与所述夹流结构之间设置至少一段蛇形通道。Preferably, at least one serpentine channel is provided between the downstream of the main channel and the flow entrainment structure.

优选的，在所述主通道下游设置至少一个的支通道，至少一个所述支通道下游与所述侧通道下端连通连接于所述夹流结构。Preferably, at least one branch channel is provided downstream of the main channel, and the downstream of at least one branch channel communicates with the lower end of the side channel and is connected to the entrainment structure.

优选的，在所述微粒进样孔上游设置正压驱动装置，或在所述微粒出样孔下游设置负压驱动装置。Preferably, a positive pressure driving device is arranged upstream of the particle sampling hole, or a negative pressure driving device is arranged downstream of the particle sampling hole.

其次，本发明提供微粒自夹流式微流控芯片的制造方法，用于制造上述任一项所述的微粒自夹流式微流控芯片，包括以下步骤：Secondly, the present invention provides a method for manufacturing a particle self-entrapment microfluidic chip, which is used to manufacture the particle self-entrapment microfluidic chip described in any one of the above, comprising the following steps:

制备硅胶模板：Prepare the silica template:

通过制图软件设计具有所述主通道的主通道结构和具有所述侧通道的侧通道结构，并进行打印；Designing the main channel structure with the main channel and the side channel structure with the side channel by drawing software, and printing;

将硅片滴上光刻胶后甩胶，并将其覆盖掩膜所述侧通道结构，然后曝光；Dropping the photoresist on the silicon wafer and then throwing the glue, and covering it with the side channel structure of the mask, and then exposing;

将曝光后的硅片再次甩胶后，将其覆盖掩膜所述主通道结构，使得所述主通道结构与所述侧通道结构对准，然后曝光；After the exposed silicon wafer is shaken again, cover and mask the main channel structure so that the main channel structure is aligned with the side channel structure, and then expose;

将未固化部分通过显影液清洗，得到所述硅片模板；Cleaning the uncured part with a developer to obtain the silicon wafer template;

制备具有通道结构的上层PDMS芯片：Prepare the upper layer PDMS chip with channel structure:

将PDMS单体与固化剂混合得到PDMS高聚物；Mix PDMS monomer and curing agent to obtain PDMS polymer;

将所述PDMS高聚物倒在所述硅片模板上，烘干后得到具有所述主通道结构和所述侧通道结构的所述上层PDMS芯片；Pour the PDMS polymer onto the silicon wafer template and dry to obtain the upper PDMS chip with the main channel structure and the side channel structure;

制备所述上层PDMS芯片上的所述微粒进样孔和所述微粒出样孔：Prepare the microparticle sampling hole and the microparticle sampling hole on the upper PDMS chip:

将具有所述通道结构的所述上层PDMS芯片打出所述微粒进样孔和所述微粒出样孔；punching out the microparticle sampling hole and the microparticle sampling hole from the upper PDMS chip having the channel structure;

将待键合的下层芯片采用无刻蚀图案的PDMS光滑基片；Use a PDMS smooth substrate without etching patterns for the underlying chip to be bonded;

制备所述微粒自夹流式微流控芯片：Preparation of the microparticle self-trapping microfluidic chip:

将所述上层PDMS芯片与所述下层芯片等离子体键合制得所述微粒自夹流式微流控芯片，并经过放置后，恢复所述微粒自夹流式微流控芯片的疏水性。Plasma-bonding the upper layer PDMS chip and the lower layer chip to prepare the particle self-entrapment microfluidic chip, and after being placed, the hydrophobicity of the particle self-entrapment microfluidic chip is restored.

再次，本发明提供微粒自分散方法，采用上述任一项所述的微粒自夹流式微流控芯片实现，包括以下步骤：Again, the present invention provides a particle self-dispersing method, which is realized by using the particle self-clamping microfluidic chip described in any one of the above, including the following steps:

接入微粒相的进样端：将所述微粒相的进样端接入所述微粒进样孔；Connecting to the injection port of the particle phase: connecting the sample port of the particle phase to the particle injection hole;

接入压力驱动装置：将所述微粒进样孔接入正压驱动装置，或将所述微粒出样孔接入负压驱动装置；Connecting to the pressure driving device: connecting the particle sampling hole to a positive pressure driving device, or connecting the particle sampling hole to a negative pressure driving device;

分离微粒和流体：驱动所述正压驱动装置或所述负压驱动装置，将所述微粒相的进样端的液滴相通过所述微粒进样孔流入所述微粒自夹流式微流控芯片；所述微粒相在所述主通道下游和所述侧通道下端分别分离出微粒和夹流液；Separation of particles and fluid: drive the positive pressure drive device or the negative pressure drive device, and flow the liquid droplet phase at the injection end of the particle phase into the particle self-pinching microfluidic chip through the particle injection hole ; The particle phase is separated from the particles and the entrainment liquid at the downstream of the main channel and the lower end of the side channel, respectively;

汇集所述微粒和所述夹流液及微粒分散：使得分离出的所述微粒和夹流液在所述夹流结构汇集，通过所述夹流液将所述微粒自分散；Collecting the particles, the entraining liquid, and dispersing the particles: making the separated particles and the entraining liquid gather in the entrainment structure, and self-dispersing the particles through the entraining liquid;

得到自分散后所述微粒：将自分散后的所述微粒经所述微粒出样孔流出。Obtaining the microparticles after self-dispersion: the self-dispersed microparticles flow out through the microparticle sampling hole.

本发明由于采取以上技术方案，其具有以下优点：The present invention has the following advantages due to the adoption of the above technical scheme:

1、本发明提供的微粒自夹流式微流控芯片和微粒自分散方法，适用于各尺寸微粒的自分散，通道高度和宽度可根据微粒直径进行调整；1. The particle self-entrapment microfluidic chip and particle self-dispersion method provided by the present invention are suitable for the self-dispersion of particles of various sizes, and the channel height and width can be adjusted according to the particle diameter;

2、本发明提供的微粒自夹流式微流控芯片和微粒自分散方法，实现了从微粒相中过滤出部分流体作为夹流流体，减少了夹流液消耗，不需额外的泵装置，使得该微粒自夹流式微流控芯片可以做到小型化与便携化；2. The particle self-entrapment microfluidic chip and the particle self-dispersion method provided by the present invention realize the filtration of part of the fluid from the particle phase as the entrainment fluid, which reduces the consumption of entrainment fluid and does not require additional pump devices, making The particle self-clamping microfluidic chip can be miniaturized and portable;

3、本发明提供的微粒自夹流式微流控芯片和微粒自分散方法，使得该芯片可重复利用，降低操作成本；3. The particle self-entrapment microfluidic chip and particle self-dispersion method provided by the present invention make the chip reusable and reduce operating costs;

4、本发明提供的微粒自夹流微流控芯片的制造方法、微粒自分散方法，均操作简便，且其制造方法成品率高，能够保证产品质量。4. The method for manufacturing microfluidic chips with self-encapsulation of microparticles and the method for self-dispersion of microparticles provided by the present invention are both easy to operate, and the manufacturing method has a high yield rate and can ensure product quality.

附图说明Description of drawings

图1为本发明实施例1提供的微粒自夹流式微流控芯片的结构示意图；FIG. 1 is a schematic structural view of the particle self-encapsulating microfluidic chip provided in Example 1 of the present invention;

图2为本发明实施例1的含有蛇形通道及微粒阻拦结构和气泡阻拦结构的结构示意图；Fig. 2 is a schematic structural view of the embodiment 1 of the present invention including a serpentine channel, a particle blocking structure and a bubble blocking structure;

图3为本发明实施例1的微粒阻拦结构和气泡阻拦结构的示意图；Fig. 3 is a schematic diagram of a particle blocking structure and a bubble blocking structure in Example 1 of the present invention;

图4为本发明实施例1的一个侧通道为弯折结构，且在夹流结构处主通道宽度大于侧通道宽度的示意图；Fig. 4 is a schematic diagram of a side channel in Embodiment 1 of the present invention having a bent structure, and the width of the main channel at the entrainment structure is greater than the width of the side channel;

图5为本发明实施例1的两个侧通道为弯折结构，且在夹流结构处主通道宽度大于侧通道宽度的示意图；Fig. 5 is a schematic diagram showing that the two side channels in Embodiment 1 of the present invention are bent structures, and the width of the main channel is greater than the width of the side channels at the entrainment structure;

图6为本发明实施例1的一个侧通道为弯折结构，且在夹流结构处主通道宽度等于侧通道宽度的示意图；Fig. 6 is a schematic diagram of a side channel in Embodiment 1 of the present invention having a bent structure, and the width of the main channel at the entrainment structure is equal to the width of the side channel;

图7为本发明实施例1的两个侧通道为弯折结构的示意图，且在夹流结构处主通道宽度等于侧通道宽度；Fig. 7 is a schematic diagram of the two side channels in Embodiment 1 of the present invention as a bent structure, and the width of the main channel at the entrainment structure is equal to the width of the side channel;

图8为本发明实施例1的一个侧通道为弯折结构，且在夹流结构处主通道宽度小于侧通道宽度的示意图；Fig. 8 is a schematic diagram of a side channel in Embodiment 1 of the present invention having a bent structure, and the width of the main channel at the entrainment structure is smaller than the width of the side channel;

图9为本发明实施例1的两个侧通道为弯折结构，且在夹流结构处主通道宽度大于侧通道宽度的示意图；Fig. 9 is a schematic diagram showing that the two side channels in Embodiment 1 of the present invention are bent structures, and the width of the main channel is greater than the width of the side channel at the entrainment structure;

图10为本发明实施例1的一个侧通道为弧形弯曲结构的示意图；Fig. 10 is a schematic diagram of a side channel in Embodiment 1 of the present invention having an arc-shaped curved structure;

图11为本发明实施例1的两个侧通道为弧形弯曲结构的示意图；Fig. 11 is a schematic diagram of the two side passages in Embodiment 1 of the present invention having an arc-shaped curved structure;

图12为本发明实施例1的一个侧通道为弯折结构、主通道设置两个或多个支通道的示意图；Fig. 12 is a schematic diagram of a side channel in Embodiment 1 of the present invention with a bent structure and two or more branch channels in the main channel;

图13为本发明实施例1的两个侧通道为弯折结构、主通道设置两个或多个支通道的示意图；Fig. 13 is a schematic diagram of the two side passages in Embodiment 1 of the present invention having a bent structure and the main passage being provided with two or more branch passages;

图14为本发明实施例2的微粒自夹流式微流控芯片制造方法的流程图；14 is a flow chart of the method for manufacturing a microparticle self-encapsulating microfluidic chip according to Embodiment 2 of the present invention;

图15为本发明实施例2的主通道结构示意图；Fig. 15 is a schematic diagram of the main channel structure of Embodiment 2 of the present invention;

图16为本发明实施例2的侧通道结构示意图；Fig. 16 is a schematic diagram of the side channel structure of Embodiment 2 of the present invention;

图17为本发明实施例3的微粒自夹流式微流控芯片使用方法的流程图；17 is a flow chart of the method for using the microparticle self-entrapment microfluidic chip according to Example 3 of the present invention;

图18为本发明实施例3的液滴自夹流效果图；Fig. 18 is a diagram showing the effect of droplet self-entrapment in Example 3 of the present invention;

图19为本发明实施例4的微粒自夹流式微流控芯片使用方法的流程图；Fig. 19 is a flow chart of the method for using the particle self-encapsulation microfluidic chip according to Example 4 of the present invention;

图20为本发明实施例4的液滴荧光信号图；Figure 20 is a droplet fluorescence signal diagram of Example 4 of the present invention;

图21为实施例5的微粒自分散方法的流程图；Fig. 21 is the flowchart of the microparticle self-dispersing method of embodiment 5;

图22为实施例6的微粒自分散方法的流程图；Fig. 22 is the flowchart of the microparticle self-dispersing method of embodiment 6;

图23为实施例7的微粒自分散方法的流程图；Fig. 23 is the flowchart of the microparticle self-dispersing method of embodiment 7;

图24为实施例8的微粒自分散方法的流程图；Fig. 24 is the flowchart of the microparticle self-dispersing method of embodiment 8;

图25为实施例9的微粒自分散方法的流程图；Fig. 25 is the flowchart of the microparticle self-dispersing method of embodiment 9;

图26为实施例10的微粒自分散方法的流程图。26 is a flow chart of the microparticle self-dispersion method in Example 10.

图中附图标记：1为微粒进样孔，2为主通道，3为侧通道，4为夹流结构，5为微粒出样孔，6为气泡阻拦结构，7为微粒阻拦结构，8为蛇形通道，A为主通道结构，B为侧通道结构。Reference numerals in the figure: 1 is the microparticle sampling hole, 2 is the main channel, 3 is the side channel, 4 is the entrainment structure, 5 is the microparticle sampling hole, 6 is the air bubble blocking structure, 7 is the particle blocking structure, 8 is the Serpentine channel, A is the main channel structure, B is the side channel structure.

具体实施方式Detailed ways

为使本发明的目的、技术方案和优点更加清楚，下面将结合附图对本发明的技术方案进行清楚、完整地描述。显然，所描述的实施例是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be clearly and completely described below in conjunction with the accompanying drawings. Apparently, the described embodiments are some, but not all, embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.

在本发明的描述中，需要说明的是，术语“上”、“下”、“前”、“后”等指示的方位或位置关系为基于附图所示的方位或位置关系，仅是为了便于描述本发明和简化描述，而不是指示或暗示所指的***或组件必须具有特定的方位、以特定的方位构造和操作，因此不能理解为对本发明的限制。In the description of the present invention, it should be noted that the orientation or positional relationship indicated by the terms "upper", "lower", "front", "rear" etc. is based on the orientation or positional relationship shown in the drawings, and is only for It is convenient to describe the present invention and simplify the description, but not to indicate or imply that the system or component referred to must have a specific orientation, be constructed and operate in a specific orientation, and thus should not be construed as limiting the present invention.

在本发明的描述中，需要说明的是，除非另有明确的规定和限定，术语“装配”、“设置”、“连接”应做广义理解，例如，可以是固定连接，也可以是可拆卸连接，或一体地连接；可以是机械连接，也可以是电连接；可以是直接相连，也可以通过中间媒介间接相连，可以是两个组件内部的连通。对于本领域的普通技术人员而言，可以具体情况理解上述术语在本发明中的具体含义。In the description of the present invention, it should be noted that unless otherwise specified and limited, the terms "assembly", "setting", and "connection" should be understood in a broad sense, for example, it can be a fixed connection or a detachable connection. Connected, or integrally connected; it may be mechanically connected or electrically connected; it may be directly connected or indirectly connected through an intermediary, and it may be the internal communication of two components. Those of ordinary skill in the art can understand the specific meanings of the above terms in the present invention in specific situations.

本发明的目的是提供微粒自夹流式微流控芯片及其制造方法和微粒自分散方法，通过所述微粒自夹流式微流控芯片，能够将紧密排列的微粒分散，形成有一定间隔的单微粒顺序流，无需另外的夹流液与驱动设备，降低了微粒再注入的成本，无需蠕动泵等设备，实现了装置的简化。The object of the present invention is to provide a particle self-entrapping microfluidic chip, a manufacturing method thereof, and a particle self-dispersing method. Through the particle self-entrapping microfluidic chip, closely arranged particles can be dispersed to form a single particle with a certain interval. Sequential flow of particles does not require additional entrainment liquid and driving equipment, which reduces the cost of particle reinjection, and does not require peristaltic pumps and other equipment, which simplifies the device.

下面，结合附图对本发明实施例提供的微粒自夹流式微流控芯片进行详细的说明。In the following, the particle self-encapsulating microfluidic chip provided by the embodiment of the present invention will be described in detail with reference to the accompanying drawings.

实施例1Example 1

如图1所示，本实施例提供的微粒自夹流式微流控芯片，所述微粒自夹流式微流控芯片内设置至少一条微流体通道，微流体通道包括主通道2、至少一段侧通道3和夹流结构4；As shown in Figure 1, the particle self-entrapment microfluidic chip provided in this embodiment is provided with at least one microfluidic channel in the particle self-entrapment microfluidic chip, and the microfluidic channel includes a main channel 2 and at least one section of side channel 3 and entrainment structure 4;

主通道2上端设置微粒相入口，夹流结构4下端设置微粒出口；The upper end of the main channel 2 is provided with a particle phase inlet, and the lower end of the entrainment structure 4 is provided with a particle outlet;

侧通道3上端与主通道2上游连通连接，微流体通道设置阻止微粒进入侧通道3的微粒阻截结构7，侧通道3下端与主通道2下游连通连接，夹流结构4位于侧通道3下端与主通道2下游连通连接处；The upper end of the side channel 3 is connected to the upstream of the main channel 2, the microfluidic channel is provided with a particle blocking structure 7 that prevents particles from entering the side channel 3, the lower end of the side channel 3 is connected to the downstream of the main channel 2, and the entrainment structure 4 is located at the lower end of the side channel 3 and The downstream connection of the main channel 2;

所述微粒自夹流式微流控芯片上还设置微粒进样孔1和微粒出样孔5，微粒进样孔1与主通道2的微粒相入口连通连接，微粒出样孔5与夹流结构4的微粒出口连通连接。The microparticle self-entrapment microfluidic chip is also provided with a particle sampling hole 1 and a particle sampling hole 5, the particle sampling hole 1 is connected to the particle phase inlet of the main channel 2, and the particle sampling hole 5 is connected to the entrainment structure. 4 is communicated with the particle outlet.

本实施例提供的所述微粒自夹流式微流控芯片在使用时，首先将微粒相通过液滴进样孔1进入所述微粒自夹流式微流控芯片的主通道2内，由于主通道2与侧通道3之间设置了微粒阻截结构，微粒相中的微粒只能流入主通道2而无法进入侧通道3，而微粒相中的部分流体进入侧通道3作为夹流液，最后汇集于主通道2下游的夹流结构4处，使得过滤到侧通道3内的夹流液与主通道2内的微粒汇集，进而使得紧密排列的微粒滴在夹流液的作用下，在夹流结构4处自分散。由于无需额外的夹流液与驱动设备，因此降低了微粒再注入的成本，无需蠕动泵等设备，实现了装置的简化。该所述微粒自夹流式微流控芯片可以应用于细胞单包裹、细胞单分散、液滴荧光信号检测、微生物液滴信号检测，也可以应用于液滴酶活筛选、微生物组液滴培养后的流式分选与质谱分析、细胞测序的耦合接口，其适用于细胞分选、液滴操纵、液滴检测、液滴计数、液滴分选、数字核酸扩增等微流控领域；其中，形成微粒自夹流并分选方法还包括对分选的目标微粒进行进一步操作，能够将紧密排列的微粒分散，形成有间隔的单微粒顺序流，其中，间隔可以为微米至米量级，所述操作包含单细胞测序、单细胞形态分析、单细胞培养等。When the microparticle self-encapsulating microfluidic chip provided in this embodiment is in use, firstly, the particle phase enters the main channel 2 of the microparticle self-encapsulating microfluidic chip through the droplet injection hole 1, because the main channel 2 and the side channel 3 is provided with a particle blocking structure, the particles in the particle phase can only flow into the main channel 2 and cannot enter the side channel 3, while part of the fluid in the particle phase enters the side channel 3 as the entrainment liquid, and finally gathers in the At the entrainment structure 4 downstream of the main channel 2, the entrainment liquid filtered into the side channel 3 and the particles in the main channel 2 converge, so that the tightly arranged particles drop under the action of the entrainment liquid, and in the entrainment structure 4 self-dispersing. Since there is no need for additional entrainment liquid and driving equipment, the cost of particle reinjection is reduced, and equipment such as a peristaltic pump is not required, which simplifies the device. The microparticle self-encapsulating microfluidic chip can be applied to cell single encapsulation, cell monodispersion, droplet fluorescence signal detection, microbial droplet signal detection, and can also be applied to droplet enzyme activity screening and microbiome droplet culture. The coupling interface of flow cytometry sorting, mass spectrometry analysis, and cell sequencing is suitable for microfluidic fields such as cell sorting, droplet manipulation, droplet detection, droplet counting, droplet sorting, and digital nucleic acid amplification; among them The method of forming self-entrapment and sorting of particles also includes further operations on the sorted target particles, which can disperse closely arranged particles to form a sequential flow of single particles with intervals, wherein the intervals can be on the order of microns to meters, The operations include single cell sequencing, single cell morphology analysis, single cell culture, etc.

如图3所示，上述实施例中，微粒阻截结构7为设置在侧通道3上端与主通道2上游连通连接处的微粒阻拦结构。微粒阻拦结构阻止微粒相内的微粒流入侧通道3，使得微粒相中的部分流体进入侧通道3，促进微粒相流经侧通道3 上端与主通道2分岔处时，微粒与流体的分离。As shown in FIG. 3 , in the above-mentioned embodiment, the particle blocking structure 7 is a particle blocking structure arranged at the upper end of the side channel 3 and the upstream connection of the main channel 2 . The particle blocking structure prevents the particles in the particle phase from flowing into the side channel 3, so that part of the fluid in the particle phase enters the side channel 3, and promotes the separation of particles and fluid when the particle phase flows through the bifurcation between the upper end of the side channel 3 and the main channel 2.

如图3所示，上述实施例中，微粒阻拦结构可以为微筛阵列结构，但不限于微筛阵列结构。其中，微筛阵列结构的微粒阻拦结构能够进一步提高微粒相流经侧通道3上端与主通道2分岔处时，微粒与夹流液的分离效果和分离效率。如图1所示，上述实施例中，侧通道3与主通道2连接处高度高于侧通道3高度构成高度差值；微粒阻截结构7为侧通道3的宽度小于微粒直径所构成的尺寸差结构，或为高度差值小于微粒直径所构成的尺寸差结构，或为二者结构的组合。As shown in FIG. 3 , in the above embodiment, the particle blocking structure may be a microsieve array structure, but is not limited to the microsieve array structure. Among them, the particle blocking structure of the micro-sieve array structure can further improve the separation effect and separation efficiency of the particles and the entraining liquid when the particle phase flows through the bifurcation between the upper end of the side channel 3 and the main channel 2 . As shown in Figure 1, in the above embodiment, the height of the connection between the side channel 3 and the main channel 2 is higher than the height of the side channel 3 to form a height difference; the particle blocking structure 7 is the size difference formed by the width of the side channel 3 being smaller than the particle diameter The structure, or the size difference structure formed by the height difference value being smaller than the particle diameter, or a combination of the two structures.

即，侧通道3内部为圆形通道时，其内部通道的直径小于微粒直径所构成的尺寸差结构构成微粒阻截结构7；或侧通道3内部为矩形等非圆形通道时，其内部通道的最小宽度小于微粒直径所构成的尺寸差结构构成微粒阻截结构7。或相对于侧通道3与主通道2连接处，即侧通道3与主通道2的分岔处，该连接处的高度高于侧通道3上端入口端的高度，即上述分岔处的高度高于侧通道3上端入口端的高度，该高度差值小于微粒直径的尺寸差结构构成微粒阻截结构7。That is, when the inside of the side channel 3 is a circular channel, the diameter of the internal channel is smaller than the size difference structure formed by the particle diameter to form the particle blocking structure 7; or when the inside of the side channel 3 is a non-circular channel such as a rectangle, the diameter of the internal channel The size difference structure formed by the minimum width being smaller than the particle diameter constitutes the particle blocking structure 7 . Or relative to the connection between the side channel 3 and the main channel 2, that is, the branch of the side channel 3 and the main channel 2, the height of the connection is higher than the height of the upper end of the side channel 3, that is, the height of the above-mentioned branch is higher than The height of the inlet end at the upper end of the side channel 3, the size difference structure whose height difference is smaller than the particle diameter constitutes the particle blocking structure 7.

需要说明的是，主通道2内部为圆形通道时，其内部通道的直径大于微粒直径；或主通道2内部为矩形等非圆形通道时，其内部通道的最小宽度大于微粒直径。或相对于侧通道3与主通道2的连接处，即侧通道3与主通道2的分岔处，主通道2上端入口端的高度高于该连接处的高度，即主通道2上端入口端的高度高于上述分岔处的高度，该高度差值大于微粒直径。It should be noted that when the inside of the main channel 2 is a circular channel, the diameter of the inner channel is larger than the diameter of the particles; or when the inside of the main channel 2 is a non-circular channel such as a rectangle, the minimum width of the inner channel is larger than the diameter of the particles. Or relative to the connection between the side channel 3 and the main channel 2, that is, the bifurcation of the side channel 3 and the main channel 2, the height of the upper entrance of the main channel 2 is higher than the height of the connection, that is, the height of the upper entrance of the main channel 2 Above the height of the bifurcation, the difference in height is greater than the particle diameter.

综上，侧通道3与主通道2连接处高度高于侧通道3高度构成高度差值；微粒阻截结构7为设置在侧通道3与主通道2连接处的微粒阻拦结构7，或为侧通道3的宽度小于微粒直径所构成的尺寸差结构，或为高度差值小于微粒直径所构成的尺寸差结构，或为其中二者或三者结构的组合。使得微粒能够进入通过主通道2，而无法进入通过侧通道3。To sum up, the height of the connection between the side channel 3 and the main channel 2 is higher than the height of the side channel 3 to form a height difference; the particle blocking structure 7 is a particle blocking structure 7 arranged at the connection between the side channel 3 and the main channel 2, or is a side channel The size difference structure formed by the width of 3 being smaller than the particle diameter, or the size difference structure formed by the height difference being smaller than the particle diameter, or a combination of two or three of them. So that particles can enter through the main channel 2, but cannot enter through the side channel 3.

如图1所示，上述实施例中，主通道2上端高度与宽度均大于微粒直径，其目的是降低侧通道3相对于主通道2的流阻，防止微粒进入侧通道3。As shown in FIG. 1 , in the above embodiment, the height and width of the upper end of the main channel 2 are greater than the diameter of the particle, the purpose of which is to reduce the flow resistance of the side channel 3 relative to the main channel 2 and prevent particles from entering the side channel 3 .

上述实施例中，主通道2上端高度或宽度大于侧通道3上端高度或宽度，主通道2上端高度与宽度均大于微粒直径，侧通道3上端高度或宽度小于微粒直径，并且侧通道3上端与主通道2上游连通连接处的微粒阻拦结构7，其目的是更好地防止微粒进入侧通道3。可以进一步的，主通道2下端高度或宽度大于侧通道3下端高度或宽度，即主通道2整体高度或宽度大于侧通道3整体高度或宽度，其作用是促进侧通道3相对于主通道2的流阻的降低，更好地阻止微粒进入侧通道3。In the above embodiment, the height or width of the upper end of the main channel 2 is greater than the height or width of the upper end of the side channel 3, the height and width of the upper end of the main channel 2 are greater than the diameter of the particle, the height or width of the upper end of the side channel 3 is smaller than the diameter of the particle, and the upper end of the side channel 3 is closer to the diameter of the particle. The particle blocking structure 7 at the upstream connection of the main channel 2 is to better prevent particles from entering the side channel 3 . Further, the height or width of the lower end of the main channel 2 is greater than the height or width of the lower end of the side channel 3, that is, the overall height or width of the main channel 2 is greater than the overall height or width of the side channel 3, and its effect is to promote the side channel 3 relative to the main channel 2. The reduction of the flow resistance better prevents particles from entering the side channel 3 .

如图3所示，上述实施例中，微粒进样孔1处设置气泡阻拦结构6(如微筛阵列结构)，气泡阻拦结构6用于防止气泡流入主通道2。As shown in FIG. 3 , in the above embodiment, a bubble blocking structure 6 (such as a micro-sieve array structure) is provided at the microparticle sampling hole 1 , and the bubble blocking structure 6 is used to prevent air bubbles from flowing into the main channel 2 .

上述实施例中，气泡阻拦结构6可以为微筛阵列结构，但不限于阵列结构。其中，微筛阵列结构的气泡阻拦结构6能够提高阻止气泡进入主通道2的效果和效率。In the above embodiments, the bubble blocking structure 6 may be a microsieve array structure, but is not limited to the array structure. Wherein, the bubble blocking structure 6 of the micro-sieve array structure can improve the effect and efficiency of preventing bubbles from entering the main channel 2 .

如图2所示，上述实施例中，主通道2下游与夹流结构4之间设置至少一段蛇形通道8。蛇形通道8能够起到对微粒进行缓流的作用。As shown in FIG. 2 , in the above embodiment, at least one serpentine channel 8 is provided between the downstream of the main channel 2 and the flow structure 4 . The serpentine channel 8 can play the role of slowing the flow of particles.

如图1所示，上述实施例中，为了提高微粒相中微粒与流体的分离效率和分离效果，侧通道3的数量可以为两段或两段以上。此外，为了使得微粒相中的流体更均匀、更充分地与微粒分离，两段或两段以上的侧通道3可以均匀分布地与主通道2连通连接。As shown in FIG. 1 , in the above embodiment, in order to improve the separation efficiency and separation effect between the particles and the fluid in the particle phase, the number of side channels 3 may be two or more. In addition, in order to separate the fluid in the particle phase from the particles more uniformly and fully, two or more sections of side channels 3 can be evenly distributed and connected to the main channel 2 .

上述实施例中，主通道2的下游可以变窄，即主通道2下游的内径尺寸小于其中游的内径尺寸，其目的为方便侧通道3下端与主通道2下游连通连接构成汇集结构，促进侧通道3中的流体作为夹流液将主通道2中的微粒分散。其中，主通道2下游的内径尺寸可以线性减小。In the above-mentioned embodiment, the downstream of the main passage 2 can be narrowed, that is, the inner diameter of the downstream of the main passage 2 is smaller than the inner diameter of the middle reaches. The fluid in channel 3 acts as entrainment fluid to disperse the particles in main channel 2 . Wherein, the inner diameter size downstream of the main channel 2 can be linearly reduced.

上述实施例中，微粒进样孔1作为微粒相进入所述微粒自夹流式微流控芯片的通孔，其上游可以是液滴发生结构，但不限于此种结构。即微粒进样孔1可以与液滴发生结构的液滴出样端连通连接。液滴的油相可以是氟碳油、矿物油、硅油、植物油等；水相可以是纯水，培养液、菌液、反应液、培养过的液滴等。In the above embodiments, the particle injection hole 1 enters the through hole of the particle self-pinching microfluidic chip as the particle phase, and its upstream may be a droplet generating structure, but it is not limited to this structure. That is, the microparticle sampling hole 1 can communicate with the droplet outlet end of the droplet generating structure. The oil phase of the droplet can be fluorocarbon oil, mineral oil, silicone oil, vegetable oil, etc.; the water phase can be pure water, culture solution, bacteria solution, reaction solution, cultured droplet, etc.

上述实施例中，微粒进样孔1作为微粒相进入所述微粒自夹流式微流控芯片的通孔，其上游可以是细胞悬液、微球等，但不限于此流体。细胞悬液、微球等可以是分散于纯水、培养液、反应液中、被培养过的液滴等。In the above embodiments, the microparticle injection hole 1 enters the through hole of the microfluidic chip from the microfluidic chip as the microparticle phase, and its upstream can be cell suspension, microspheres, etc., but it is not limited to this fluid. The cell suspension, microspheres, etc. may be dispersed in pure water, culture solution, reaction solution, cultured droplets, or the like.

如图3所示，上述实施例中，主通道2作为微粒相通道，其与微粒进样孔1连通连接的前端结构可以是阵列结构的气泡阻拦结构6，以防止气泡流入。气泡阻拦结构6上游可以是液滴发生、培养、反应结构，但不局限于此种结构。As shown in FIG. 3 , in the above-mentioned embodiment, the main channel 2 is used as a particle phase channel, and its front-end structure communicating with the particle injection hole 1 can be a bubble blocking structure 6 in an array structure to prevent the inflow of bubbles. The upstream of the bubble blocking structure 6 may be a droplet generating, cultivating, and reacting structure, but it is not limited to this structure.

上述实施例中，侧通道3作为微粒相中的夹流液通道，如前所述，主通道2与侧通道3交界处可以添加阵列结构的微粒阻拦结构，但不限于此种结构。目的是阻止微粒流入侧通道。或者可以相对于微粒的直径，减小侧通道3的内径尺寸，以防止微粒进入侧通道3。In the above embodiment, the side channel 3 is used as the entrainment liquid channel in the particle phase. As mentioned above, a particle blocking structure of an array structure can be added at the junction of the main channel 2 and the side channel 3, but it is not limited to this structure. The purpose is to prevent particles from flowing into the side channels. Alternatively, the size of the inner diameter of the side channel 3 may be reduced relative to the diameter of the particles to prevent the particles from entering the side channel 3 .

如图1所示，上述实施例中，夹流结构4位于主通道2与侧通道3交汇处，在该结构实现通过夹流液将密集排列的微粒自分散。As shown in FIG. 1 , in the above embodiment, the entrainment structure 4 is located at the junction of the main channel 2 and the side channel 3 , and in this structure, the densely arranged particles are self-dispersed through the entrainment liquid.

上述实施例中，微粒出样孔5作为微粒流出所述液滴自分散式微流控芯片的通孔，其下游可以是信号检测结构、分选结构等，但不限于此种结构。即微粒出样孔5可以与信号检测结构、分选结构的进样端连通连接。In the above embodiments, the particle sampling hole 5 is used as a through hole through which the particles flow out from the dispersed microfluidic chip, and its downstream may be a signal detection structure, a sorting structure, etc., but it is not limited to this structure. That is, the microparticle sampling hole 5 can be communicated with the signal detection structure and the sampling end of the sorting structure.

如图13所示，上述实施例中，主通道2下游可以设置两个以上的支通道。至少一个支通道下游与侧通道3下端连通连接。即微流体通道作为样品通道，包括主通道2、侧通道3和夹流结构4，侧通道3从主通道2上游中分出，最终汇集于主通道2下游或汇集于主通道2的支通道。由于夹流结构4位于主通道2和侧通道3汇集处，则当主通道2设置支通道时，夹流结构4位于主通道2的支通道与其侧通道3汇集处。而当主通道2具有两个以上的多个支通道时，每个支通道可以匹配一个或多个侧通道3，则夹流结构4的数量可以为多个。即多个夹流结构4分别位于每个支通道与该支通道的每个侧通道3的汇集处。As shown in FIG. 13 , in the above-mentioned embodiment, more than two branch passages may be provided downstream of the main passage 2 . The downstream of at least one branch channel communicates with the lower end of the side channel 3 . That is, the microfluidic channel is used as a sample channel, including a main channel 2, a side channel 3 and an entrainment structure 4. The side channel 3 is separated from the upstream of the main channel 2, and finally gathers in the downstream of the main channel 2 or in a branch channel of the main channel 2 . Since the entrainment structure 4 is located at the confluence of the main channel 2 and the side channel 3 , when the main channel 2 is provided with a branch channel, the air entrainment structure 4 is located at the confluence of the branch channel of the main channel 2 and the side channel 3 . And when the main channel 2 has more than two branch channels, each branch channel can be matched with one or more side channels 3 , and the number of the flow-entrapping structures 4 can be multiple. That is, a plurality of flow-entrapping structures 4 are respectively located at the confluence of each branch channel and each side channel 3 of the branch channel.

如图1所示，上述样品通道的入口端与微粒进样孔1相连，出口端与微粒出样孔5相连。As shown in FIG. 1 , the inlet end of the above-mentioned sample channel is connected with the microparticle sampling hole 1 , and the outlet end is connected with the microparticle sampling hole 5 .

上述实施例中，所述微粒自夹流式微流控芯片材质可以选用PDMS(聚二甲基硅氧烷)、PMMA(聚甲基丙烯酸甲酯)、石英、硼硅玻璃、氟化钙，但是不限于上述材质。In the above-mentioned embodiment, the material of the particle self-entrapping microfluidic chip can be selected from PDMS (polydimethylsiloxane), PMMA (polymethyl methacrylate), quartz, borosilicate glass, calcium fluoride, but It is not limited to the above materials.

如图3所示，上述实施例中，所述微粒自夹流式微流控芯片的微流体通道的内表面可以为疏水亲油表面。此时，如前所述，主通道2下游与夹流结构4之间设置至少一段蛇形通道8，蛇形通道8对微粒进行缓流，更好地提高夹流液与微粒分离的效果和效率。蛇形通道8可以为“C”字形弯曲通道、“S”字形弯曲通道或“弓”字形弯曲通道。蛇形通道8的弯曲处可以为弧形弯曲结构。As shown in FIG. 3 , in the above-mentioned embodiment, the inner surface of the microfluidic channel of the particle self-entrapping microfluidic chip may be a hydrophobic and lipophilic surface. At this time, as mentioned above, at least one section of serpentine channel 8 is provided between the downstream of the main channel 2 and the entrainment structure 4, and the serpentine channel 8 slows down the flow of particles, so as to better improve the separation effect and efficiency. The serpentine passage 8 may be a "C" curved passage, an "S" curved passage or a "bow" curved passage. The bend of the serpentine passage 8 may be an arc-shaped bend structure.

上述实施例中，微粒进样孔1可以与进样导管接口连接。In the above embodiment, the microparticle sampling hole 1 can be connected with the interface of the sampling catheter.

如图1所示，上述实施例中，侧通道3的内径小于微粒直径。侧通道3的内径小于主通道2的内径。这样能够进一步避免微粒进入侧通道3，保证夹流液与微粒相的有效分离。As shown in FIG. 1 , in the above embodiment, the inner diameter of the side channel 3 is smaller than the particle diameter. The inner diameter of the side channel 3 is smaller than the inner diameter of the main channel 2 . This can further prevent particles from entering the side channel 3 and ensure effective separation of the entraining liquid and the particle phase.

上述实施例中，微粒出样孔5与夹流结构4可以通过通道连通连接，夹流结构4与主通道2可以通过通道连通连接。则侧通道3用于过滤微粒相中的微粒，只允许微粒相中部分流体进入；夹流结构4与通道连通连接所构成的夹流通道，用于将侧通道3的流体汇入，使其作为夹流液，实现分散微粒分散通道内紧密排列的微粒。In the above embodiment, the particle sampling hole 5 and the entrainment structure 4 can be connected through a channel, and the entrainment structure 4 can be connected with the main channel 2 through a channel. Then the side channel 3 is used to filter the particles in the particle phase, and only allows part of the fluid in the particle phase to enter; the entrainment structure 4 is connected to the channel to form an entrainment channel, which is used to bring the fluid in the side channel 3 into it, so that it As an entrainment liquid, the closely arranged particles in the dispersed particle dispersion channel are realized.

上述实施例中，主通道2的微粒相入口可以与液滴发生结构的出样端连通连接。In the above embodiments, the particle phase inlet of the main channel 2 can be communicated with the sample outlet of the droplet generating structure.

上述实施例中，液滴直径可以为50μm，主通道2的高度可以为40～100μm，优选为50μm；侧通道3的高度可以为1～50μm，优选为10μm；夹流结构4与出样孔5之间的通道宽度可以为50～100μm；主通道2最大宽度可以为50～500μm，最小宽度可以为50～100μm；侧通道3最大宽度可以为50～500μm，最小宽度可以为10～100μm。In the above embodiment, the droplet diameter can be 50 μm, the height of the main channel 2 can be 40-100 μm, preferably 50 μm; the height of the side channel 3 can be 1-50 μm, preferably 10 μm; the entrainment structure 4 and the sample outlet The channel width between 5 can be 50-100 μm; the maximum width of main channel 2 can be 50-500 μm, and the minimum width can be 50-100 μm; the maximum width of side channel 3 can be 50-500 μm, and the minimum width can be 10-100 μm.

上述实施例中，微粒进样孔1与液滴发生结构、液滴发生装置或液滴样品进样装置的出样端连通连接，液滴发生结构、液滴发生装置或液滴样品进样装置可以是重力驱动调节进样装置、注射器、蠕动泵、注射泵，但是不限于上述装置。In the above-mentioned embodiments, the particle injection hole 1 is connected to the sample outlet of the droplet generating structure, the droplet generating device, or the droplet sample sampling device, and the droplet generating structure, the droplet generating device, or the droplet sample sampling device It can be a gravity-driven regulating injection device, a syringe, a peristaltic pump, a syringe pump, but is not limited to the above-mentioned devices.

上述实施例中，所述微粒自夹流式微流控芯片可以包含液滴发生结构或装置、细胞或液滴培养结构或装置、细胞或液滴反应结构或装置，液滴发生结构或装置、细胞或液滴培养结构或装置、细胞或液滴反应结构或装置应位于主通道2上游。In the above embodiment, the microparticle self-entrapment microfluidic chip may include a droplet generating structure or device, a cell or droplet culture structure or device, a cell or droplet reaction structure or device, a droplet generating structure or device, a cell Or droplet culture structure or device, cell or droplet reaction structure or device should be located upstream of main channel 2 .

上述实施例中，所述微粒自夹流式微流控芯片可以包含分选结构或装置、检测结构或装置，分选结构或装置、检测结构或装置应位于夹流结构4下游。In the above embodiment, the particle self-entraining microfluidic chip may include a sorting structure or device, a detection structure or device, and the sorting structure or device, detection structure or device should be located downstream of the entrainment structure 4 .

上述实施例中，液滴发生结构或装置可以包含所述微粒自夹流式微流控芯片内的液滴发生结构与所述微粒自夹流式微流控芯片外的液滴发生装置。In the above embodiments, the droplet generating structure or device may include the droplet generating structure inside the particle self-trapping microfluidic chip and the droplet generating device outside the particle self-trapping microfluidic chip.

所述微粒自夹流式微流控芯片内的液滴发生结构包括但不限于T型通道、夹流聚焦、共聚焦结构等。液滴发生后在所述微粒自夹流式微流控芯片内直接与主通道2连通，或通过导管与微粒进样孔1连通。The droplet generation structure of the microparticle self-entrapment microfluidic chip includes but not limited to T-shaped channel, entrainment focus, confocal structure and the like. After the droplet is generated, the particles are directly connected to the main channel 2 from the pinch-flow microfluidic chip, or communicated with the particle injection hole 1 through a conduit.

所述微粒自夹流式微流控芯片外的液滴发生结构或装置包括但不限于采用离心法、振荡法，并通过导管与液滴进样孔1连通连接的液滴发生结构或装置。The droplet generating structure or device outside the microfluidic chip of microparticle self-entrapment includes but not limited to the droplet generating structure or device that adopts centrifugation method and oscillation method, and communicates with the droplet injection hole 1 through a conduit.

所述芯片外液滴样品进样装置选自但不限于：重力驱动调节进样装置、注射器、蠕动泵、注射泵。The off-chip droplet sample sampling device is selected from but not limited to: gravity-driven regulating sampling device, syringe, peristaltic pump, syringe pump.

夹流结构4下游可包括但不限于检测装置、分选装置，即夹流结构4下游的微粒出样孔5可以与不限于检测装置、分选装置等装置的进样端连通连接。其中，检测装置包括但不限于激光激发的荧光检测装置、拉曼检测装置、光学检测装置、荧光检测装置、电导检测装置。The downstream of the entrainment structure 4 may include but not limited to a detection device and a sorting device, that is, the particle sampling hole 5 downstream of the entrainment structure 4 may be connected to a sampling port that is not limited to a detection device, a sorting device, and the like. Wherein, the detection device includes but not limited to a fluorescence detection device excited by a laser, a Raman detection device, an optical detection device, a fluorescence detection device, and a conductivity detection device.

如图4至图13所示，上述实施例中，侧通道3与主通道2上游连通连接的上部可以设置为弯折结构，该弯折结构可以为直角弯折结构、钝角弯折结构或弧形弯折结构。As shown in Figures 4 to 13, in the above embodiments, the upper part of the side channel 3 connected upstream with the main channel 2 can be set as a bent structure, and the bent structure can be a right-angle bent structure, an obtuse angle bent structure or an arc. curved structure.

上述实施例中，侧通道3与主通道2下游连通连接的下部可以设置为弯折结构，该弯折结构可以为钝角弯折结构或弧形弯折结构。In the above-mentioned embodiment, the lower part of the side channel 3 communicating with the main channel 2 downstream can be configured as a bent structure, and the bent structure can be an obtuse angle bent structure or an arc bent structure.

在夹流结构4处，主通道的宽度可以大于、小于或等于侧通道的宽度。At the sandwich structure 4, the width of the main channel may be greater than, smaller than or equal to the width of the side channel.

如图12和图13所示，主通道2可以设置两个以上的多个支通路，其中一个支通路下游与侧通道3下端连通连接，夹流结构4设置在支通路下游与侧通道3下端连通连接处，夹流结构4通过通道与一个微粒出样孔5连通连接；其余支通路直接通过通道与另一个微粒出样孔5连通连接。其中，图4、图6、图8、图10和图12中所示的侧通道3数量为一个，图5、图7、图9、图11和图13中所示的侧通道3数量为两个，且两个侧通道3均匀对称分布，也可以不均匀对称分布。As shown in Figure 12 and Figure 13, the main passage 2 can be provided with more than two branch passages, wherein the downstream of one branch passage is connected to the lower end of the side passage 3, and the entrainment structure 4 is arranged on the downstream of the branch passage and the lower end of the side passage 3 At the connection point, the entrainment structure 4 communicates with a particle sampling hole 5 through a channel; the other branch paths communicate with another particle sampling hole 5 directly through the channel. Wherein, the number of side passages 3 shown in Fig. 4, Fig. 6, Fig. 8, Fig. 10 and Fig. 12 is one, and the number of side passages 3 shown in Fig. 5, Fig. 7, Fig. 9, Fig. 11 and Fig. 13 is Two, and the two side channels 3 are evenly and symmetrically distributed, or unevenly symmetrically distributed.

如图1所示，上述实施例中，侧通道3的下游可以变窄，即侧通道3下游的宽度尺寸小于其中游的宽度尺寸，其目的为方便侧通道3下端与主通道2下游连通连接构成汇集结构，促进侧通道3中的流体作为夹流液将主通道2中的微粒分散。其中，侧通道3下游的宽度尺寸可以线性减小。As shown in Figure 1, in the above-mentioned embodiment, the downstream of the side channel 3 can be narrowed, that is, the width dimension of the downstream side channel 3 is smaller than the width dimension of the middle stream, and its purpose is to facilitate the communication and connection between the lower end of the side channel 3 and the downstream of the main channel 2 A converging structure is formed to promote the fluid in the side channel 3 to disperse the particles in the main channel 2 as entrainment liquid. Wherein, the width dimension downstream of the side channel 3 may decrease linearly.

如图14所示，本发明提供的所述微粒自夹流式微流控芯片的制造方法，包括步骤：制备硅胶模板、制备具有通道结构的上层PDMS芯片、制备上层PDMS芯片上的微粒进样孔1和微粒出样孔5、制备所述微粒自夹流式微流控芯片。其操作简便，成品率高，保证产品质量。As shown in Figure 14, the method for manufacturing the microparticle self-clamping microfluidic chip provided by the present invention includes the steps of: preparing a silica gel template, preparing an upper layer PDMS chip with a channel structure, and preparing a particle injection hole on the upper layer PDMS chip 1 and microparticle sampling hole 5. The microparticles are prepared from the pinched flow microfluidic chip. The operation is simple, the yield is high, and the product quality is guaranteed.

下面，结合附图对本发明实施例提供的所述微粒自夹流式微流控芯片的制造方法进行详细的说明。Hereinafter, the method for manufacturing the microparticle self-encapsulating microfluidic chip provided by the embodiment of the present invention will be described in detail with reference to the accompanying drawings.

实施例2Example 2

如图14所示，所述微粒自夹流式微流控芯片的制造方法，用于制造如实施例1所述的微粒自夹流式微流控芯片，包括以下步骤：As shown in FIG. 14 , the method for manufacturing the particle self-entrapment microfluidic chip is used to manufacture the particle self-entrapment microfluidic chip as described in Example 1, comprising the following steps:

S01.制备硅胶模板：S01. Preparation of silica gel template:

如图15和图16所示，通过制图软件设计具有主通道2的主通道结构A和具有侧通道3的侧通道结构B，并进行打印；As shown in Figure 15 and Figure 16, the main channel structure A with the main channel 2 and the side channel structure B with the side channel 3 are designed by drawing software, and printed;

将硅片滴上光刻胶后甩胶，并将其覆盖掩膜侧通道结构B，然后曝光；Drop the photoresist on the silicon wafer, shake it off, cover it with the mask side channel structure B, and then expose it;

将曝光后的硅片再次甩胶后，将其覆盖掩膜主通道结构A，使得主通道结构A与侧通道结构B对准，然后曝光；After the exposed silicon wafer is shaken again, it is covered with the mask main channel structure A, so that the main channel structure A is aligned with the side channel structure B, and then exposed;

将未固化部分通过显影液清洗，得到硅片模板；Clean the uncured part with a developer to obtain a silicon wafer template;

S02.制备具有通道结构的上层PDMS芯片：S02. Prepare an upper layer PDMS chip with a channel structure:

将PDMS高聚物倒在步骤S01制得的硅片模板上，烘干后得到具有主通道结构A和侧通道结构B的上层PDMS芯片；Pour the PDMS polymer onto the silicon wafer template prepared in step S01, and dry to obtain an upper layer PDMS chip with main channel structure A and side channel structure B;

S03.制备上层PDMS芯片上的微粒进样孔1和微粒出样孔5：S03. Prepare microparticle sampling hole 1 and microparticle sampling hole 5 on the upper PDMS chip:

将步骤S02制得的具有通道结构的上层PDMS芯片打出微粒进样孔1和微粒滴出样孔5；The upper layer PDMS chip with a channel structure prepared in step S02 is punched out with a microparticle sampling hole 1 and a microparticle dripping sampling hole 5;

S04.制备所述微粒自夹流式微流控芯片：S04. Prepare the particle self-clamping microfluidic chip:

将步骤S03制得的上层PDMS芯片与下层芯片等离子体键合制得所述微粒自夹流式微流控芯片，并经过放置后，恢复所述微粒自夹流式微流控芯片的疏水性。Plasma-bonding the upper PDMS chip prepared in step S03 and the lower chip to obtain the particle self-entrapment microfluidic chip, and after placing, restore the hydrophobicity of the particle self-entrapment microfluidic chip.

上述实施例中，在进行步骤S01时，制备硅胶模板中，可以采用CAD制图软件设计具有主通道2的主通道结构A和具有侧通道3的侧通道结构B。In the above embodiment, when performing step S01 , in preparing the silica gel template, CAD drawing software can be used to design the main channel structure A with the main channel 2 and the side channel structure B with the side channel 3 .

上述实施例中，在进行步骤S01时，制备硅胶模板中，可以进行菲林掩膜打印。In the above embodiment, when performing step S01, film mask printing can be performed in preparing the silica gel template.

上述实施例中，在进行步骤S01时，制备硅胶模板中，可以将清洗过的硅片滴上光刻胶进行甩胶。In the above-mentioned embodiment, when performing step S01, in preparing the silica gel template, the cleaned silicon wafer may be dripped with photoresist and then shaken off.

上述实施例中，在进行步骤S01时，制备硅胶模板中，可以将硅片滴上SU-8光刻胶进行甩胶。In the above embodiment, when performing step S01 , in preparing the silica gel template, the silicon wafer may be dripped with SU-8 photoresist to carry out the resist removal.

上述实施例中，在进行步骤S01时，制备硅胶模板中，可以在曝光机下进行曝光。In the above embodiment, when performing step S01, in preparing the silica gel template, the exposure can be performed under an exposure machine.

上述实施例中，在进行步骤S02时，制备具有通道结构的上层PDMS芯片中，可以将PDMS单体与固化剂按比例混匀，该比例可以为质量百分比10:1。In the above embodiment, when performing step S02, in preparing the upper PDMS chip with channel structure, the PDMS monomer and the curing agent can be mixed in proportion, and the ratio can be 10:1 by mass.

上述实施例中，在进行步骤S02时，制备具有通道结构的上层PDMS芯片中，可以将PDMS高聚物倒在硅片模板上，使得PDMS高聚物厚为1～10mm。In the above embodiment, when performing step S02, in preparing the upper layer PDMS chip with channel structure, the PDMS polymer can be poured on the silicon wafer template, so that the thickness of the PDMS polymer is 1-10 mm.

上述实施例中，在进行步骤S03时，制备上层PDMS芯片上的微粒进样孔1 和微粒出样孔5中，将带有通道结构的上层PDMS芯片上用打孔器打出微粒进样孔1和微粒出样孔5。In the above embodiment, when performing step S03, prepare the microparticle sampling hole 1 and the microparticle sampling hole 5 on the upper PDMS chip, and use a puncher to punch out the microparticle sampling hole 1 on the upper PDMS chip with a channel structure. And particle sampling hole 5.

上述实施例中，在进行步骤S03时，制备上层PDMS芯片上的微粒进样孔1和微粒出样孔5中，PDMS光滑基片的厚度为1～10mm。In the above embodiment, when step S03 is performed, the thickness of the PDMS smooth substrate in the microparticle sampling hole 1 and microparticle sampling hole 5 on the upper PDMS chip is prepared to be 1-10 mm.

上述实施例中，在进行步骤S04时，制备所述微粒自夹流式微流控芯片中，将等离子体键合制得所述微粒自夹流式微流控芯片，在70℃的温度条件下，经过放置8～12小时后，恢复其疏水性。In the above-mentioned embodiment, when performing step S04, the particles are prepared in the self-entrapment microfluidic chip, and the particles are prepared by plasma bonding in the self-entrapment microfluidic chip. After standing for 8-12 hours, its hydrophobicity will be restored.

本发明提供的所述微粒自夹流式微流控芯片，可以应用于细胞单包裹、液滴荧光信号检测、微生物液滴信号检测，也可以应用于微生物组液滴培养后的流式分选与质谱分析、细胞测序的耦合接口；也可以用于细胞操纵、微球操纵、液滴操纵、数字核酸扩增、液滴操纵、液滴检测、液滴计数、液滴分选等微流控领域。The microparticle self-entrapment microfluidic chip provided by the present invention can be applied to cell single encapsulation, droplet fluorescence signal detection, microbial droplet signal detection, and can also be applied to flow sorting and microbiome droplet culture. Coupling interface for mass spectrometry and cell sequencing; can also be used in microfluidic fields such as cell manipulation, microsphere manipulation, droplet manipulation, digital nucleic acid amplification, droplet manipulation, droplet detection, droplet counting, and droplet sorting .

下面，结合附图对本发明实施例提供的所述微粒自夹流式微流控芯片的使用方法进行详细的说明。Next, the method of using the microparticle self-entrapment microfluidic chip provided by the embodiment of the present invention will be described in detail with reference to the accompanying drawings.

实施例3Example 3

本实施例以所述微粒自夹流式微流控芯片应用于液滴自夹流为示范，提供通过基于流体力学的再注入式的所述微粒自夹流式微流控芯片实现紧密排列液滴的分散的使用方法。In this embodiment, the application of the microparticle self-entrapment microfluidic chip to the liquid droplet self-entrapment is used as an example, and the method of tightly arranging droplets through the reinjection-based microparticle self-entrapment microfluidic chip based on hydrodynamics is provided. Distributed usage.

如图17所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 17, the self-dispersing method of microparticles is realized by using the microfluidic chip self-clamping microfluidic chip as described in Example 1, including the following steps:

S11.接入液滴相进样端：S11. Connect to the droplet phase injection port:

将液滴相的进样端接入微粒进样孔1；Connect the injection port of the droplet phase to the particle injection hole 1;

S12.接入压力驱动装置：S12. Access to the pressure drive device:

将微粒进样孔1接入正压驱动装置，或将微粒出样孔5接入负压驱动装置；Connect the microparticle sampling hole 1 to the positive pressure driving device, or connect the microparticle sampling hole 5 to the negative pressure driving device;

S13.分离液滴和夹流液：S13. Separation of droplets and entrainment:

驱动步骤S11接入的正压驱动装置或负压驱动装置，将步骤S11接入进样端的液滴相通过微粒进样孔1流入所述微粒自夹流式微流控芯片；液滴相在主通道2下游和侧通道3下端分别分离出液滴和夹流液；Drive the positive pressure driving device or negative pressure driving device connected in step S11, and the liquid droplet phase connected to the sampling end in step S11 flows into the microparticle self-entrapment microfluidic chip through the particle injection hole 1; the liquid droplet phase is in the main The downstream of the channel 2 and the lower end of the side channel 3 separate the liquid droplets and the entrained liquid respectively;

S14.汇集液滴和分离油及液滴自夹流：S14. Collecting droplets and separating oil and droplet self-entrapment:

使得步骤S13中分离出的液滴和夹流液在夹流结构4汇集，通过夹流液将液滴自分散；The droplets separated in step S13 and the entrainment liquid are collected in the entrainment structure 4, and the droplets are self-dispersed by the entrainment liquid;

S15.得到分散后液滴(如图18所示)：S15. Obtain the dispersed droplets (as shown in Figure 18):

将步骤S14分散后的液滴经微粒出样孔5流出。The liquid droplets dispersed in step S14 flow out through the microparticle sampling hole 5 .

上述实施例中，在进行步骤S13时，分离液滴和夹流液中，将步骤S11接入进样端的液滴相通过微粒进样孔1流入所述微粒自夹流式微流控芯片时，液滴相通过气泡拦截结构6将气泡拦截，使得气泡无法进入下游所述微粒自夹流式微流控芯片结构。In the above-mentioned embodiment, when step S13 is performed, the liquid droplet and the entrainment liquid are separated, and the liquid droplet phase that is connected to the sampling end in step S11 flows into the microparticle self-entrapment microfluidic chip through the particle injection hole 1, The liquid droplet phase intercepts the bubbles through the bubble intercepting structure 6, so that the bubbles cannot enter the microfluidic chip structure of the microparticle self-entrapment downstream.

上述实施例中，在进行步骤S13时，分离液滴和夹流液中，将步骤S11接入进样端的液滴相通过微粒进样孔1流入所述微粒自夹流式微流控芯片后，由于主通道2与侧通道3交界处的微粒拦截结构7，或由于侧通道3相对于主通道2的内径小，或由于以上两结构的结合，使得油相进入侧通道3，而液滴无法进入侧通道3，实现液滴相在主通道2下游和侧通道3下端分别分离出液滴和油相。In the above-mentioned embodiment, when step S13 is performed, the liquid droplet and the entrainment liquid are separated, and the liquid droplet phase connected to the injection port in step S11 flows into the particle through the particle injection hole 1 after the particle is self-entrapped in the microfluidic chip, Due to the particle interception structure 7 at the junction of the main channel 2 and the side channel 3, or due to the small inner diameter of the side channel 3 relative to the main channel 2, or due to the combination of the above two structures, the oil phase enters the side channel 3, and the droplets cannot Entering the side channel 3, the liquid droplet phase is separated from the liquid droplet and the oil phase at the downstream of the main channel 2 and the lower end of the side channel 3 respectively.

上述实施例中，在进行步骤S15时，得到分散后液滴后，若一段时间内不会再用该所述微粒自夹流式微流控芯片，则可以将所述微粒自夹流式微流控芯片取下，用无水乙醇将主通道2冲洗干净，放入烘箱烘干后，可以循环再利用。同时也可以用无水乙醇将侧通道3、夹流结构4等的通道清洗干净，即将微流体通道整体清洗干净。In the above embodiment, when step S15 is performed, after the dispersed droplets are obtained, if the microparticles will not be used again for a period of time, the microfluidic chips can be self-supplied. After the chip is removed, the main channel 2 is rinsed with absolute ethanol, and after drying in an oven, it can be recycled. At the same time, the channels such as the side channel 3 and the entrainment structure 4 can also be cleaned with absolute ethanol, that is, the entire microfluidic channel is cleaned.

实施例4Example 4

本实施例以将所述微粒自夹流式微流控芯片作为液滴核酸扩增荧光信号分析统计装置为例，提供将所述微粒自夹流式微流控芯片作为液滴核酸扩增荧光信号分析统计装置的使用方法。In this embodiment, taking the microparticle self-entrapping microfluidic chip as a droplet nucleic acid amplification fluorescent signal analysis and statistics device as an example, it provides an analysis of the particle self-entrapping microfluidic chip as a droplet nucleic acid amplification fluorescence signal analysis. How to use the statistical device.

如图19所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 19, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S21.液滴中核酸扩增：S21. Nucleic acid amplification in droplets:

将含有核酸、酶、缓冲液、引物、荧光染料或靶标等溶液按比例混合，生成液滴，设置相应温度，使目标片段扩增；Mix solutions containing nucleic acids, enzymes, buffers, primers, fluorescent dyes or targets in proportion to generate droplets, and set the corresponding temperature to amplify the target fragments;

其中，将含有核酸、酶、缓冲液、引物、荧光染料或靶标等溶液按比例混合，例如，液滴PCR反应中，supermix比例为：核酸、酶、缓冲液、引物、荧光染料或靶标等溶质与溶液的质量百分比为1：2；Among them, the solutions containing nucleic acids, enzymes, buffers, primers, fluorescent dyes or targets are mixed in proportion. For example, in the droplet PCR reaction, the ratio of supermix is: solutes such as nucleic acids, enzymes, buffers, primers, fluorescent dyes or targets. The mass percentage with the solution is 1:2;

其中，相应温度可以例如，液滴PCR反应中，分为95℃变性、退火和72℃延伸；Wherein, the corresponding temperature can be divided into, for example, 95°C denaturation, annealing and 72°C extension in the droplet PCR reaction;

S22.基于荧光的液滴核酸扩增分析：S22. Fluorescence-based droplet nucleic acid amplification analysis:

使用基于荧光的液滴核酸扩增分析可以对目标片段进行绝对定量，是一种终点计数的核酸定量方式；Absolute quantification of target fragments can be performed using fluorescence-based droplet nucleic acid amplification analysis, which is a nucleic acid quantification method for endpoint counting;

S23.单个液滴的分散与统计：S23. Dispersion and statistics of a single droplet:

利用所述液滴自分散式微流控芯片对收集的液滴进行分散与统计，Using the droplets to disperse and count the collected droplets from the dispersed microfluidic chip,

首先，将如光纤等的荧光信号采集装置置于夹流结构4下游通道处，打开荧光信号采集分析***；Firstly, a fluorescence signal acquisition device such as an optical fiber is placed at the downstream channel of the entrainment structure 4, and the fluorescence signal acquisition and analysis system is turned on;

其次，将微粒进样口1连接含有液滴的软管，软管另一端连接正压驱动泵或注射器等驱动装置，或在微粒出样孔5连接负压驱动泵或注射器等驱动装置；再次，运行驱动装置，使液滴流入所述微粒自夹流式微流控芯片，在夹流结构4中进行分散；Secondly, connect the particle sampling port 1 to a hose containing liquid droplets, and connect the other end of the hose to a drive device such as a positive pressure drive pump or a syringe, or connect a drive device such as a negative pressure drive pump or a syringe to the particle sample outlet 5; again , operating the driving device, so that the droplets flow into the microfluidic chip from the sandwiched flow microfluidic chip, and disperse in the sandwiched flow structure 4;

然后，调整荧光信号采集软件参数，使其收集到液滴荧光信号，并对其进行分析(如图20所示)。Then, adjust the parameters of the fluorescence signal collection software so that the droplet fluorescence signal is collected and analyzed (as shown in FIG. 20 ).

还可以包括步骤S24，对所述微粒自夹流式微流控芯片进行再利用处理：It may also include step S24, reusing the microparticle self-clamping microfluidic chip:

实验结束后，用无水乙醇冲洗所述微粒自夹流式微流控芯片的微流体通道，再将所述微粒自夹流式微流控芯片烘干，以备循环利用。After the experiment, the microfluidic channel of the self-entrapping flow microfluidic chip of the microparticles was rinsed with absolute ethanol, and then the microfluidic channel of the self-entrapping microfluidic chip of the microparticles was dried for recycling.

实施例5Example 5

本实施例以将所述微粒自夹流式微流控芯片作为微生物组液滴信号分析统计装置为例，提供将所述微粒自夹流式微流控芯片作为微生物组液滴信号分析统计装置的使用方法。In this embodiment, the use of the particle self-encapsulation microfluidic chip as a microbiome droplet signal analysis and statistics device is provided as an example, and the use of the particle self-entrapment microfluidic chip as a microbiome droplet signal analysis and statistics device is provided. method.

如图21所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 21, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S31.液滴中微生物液体培养和扩增：S31. Liquid culture and expansion of microorganisms in droplets:

将含有微生物的选择培养基生成液滴，收集到EP(离心管)管中放在培养箱中培养预设时间，使目标微生物大量繁殖，充满液滴；Generate droplets from the selection medium containing microorganisms, collect them into EP (centrifuge tube) tubes and place them in an incubator for a preset period of time, so that the target microorganisms multiply and fill the droplets;

其中，预设时间可以为12小时；Wherein, the preset time may be 12 hours;

S32.基于散射光的无标记的生长表型液滴分选：S32. Label-free sorting of growing phenotype droplets based on scattered light:

使用基于散射光的无标记的生长表型液滴分选***，可以将培养过的液滴进行分选，长满目标微生物的液滴可以被分选出来并收集到；Using a label-free growth phenotype droplet sorting system based on scattered light, the cultured droplets can be sorted, and the droplets full of target microorganisms can be sorted and collected;

S33.单个液滴的分散与分析统计：S33. Dispersion and analysis statistics of a single droplet:

利用所述微粒自夹流式微流控芯片对收集的液滴进行分散，Using the microparticles to disperse the collected droplets from the pinch-flow microfluidic chip,

首先，将液滴信号采集与分选装置置于夹流结构4通道处，打开***；First, place the droplet signal collection and sorting device at channel 4 of the entrainment structure, and open the system;

其次，将微粒进样孔1连接含有菌液的注射器，或用泵从微粒进样孔1泵入菌液；或在微粒出样孔5连接负压驱动泵或注射器等驱动装置；Secondly, connect the microparticle sampling hole 1 to the syringe containing the bacterial solution, or use a pump to pump the bacterial fluid from the microparticle sampling hole 1; or connect the microparticle sampling hole 5 to a driving device such as a negative pressure driven pump or a syringe;

然后，运行驱动装置，使液滴流入所述微粒自夹流式微流控芯片，在夹流结构4中进行分散；Then, operate the driving device to make the droplets flow into the microfluidic chip from the sandwiched flow microfluidic chip, and disperse in the sandwiched flow structure 4;

最后，调整液滴信号采集与分选软件参数，使其收集到液滴信号，并根据信号对其进行统计分析。Finally, adjust the parameters of the droplet signal collection and sorting software to collect the droplet signals, and perform statistical analysis on them.

还可以包括步骤S34，对所述微粒自夹流式微流控芯片进行再利用处理：It may also include step S34, reusing the microparticle self-entrapment microfluidic chip:

实验结束后，用无水乙醇冲洗所述液滴自分散式微流控芯片的微流体通道，再将所述液滴自分散式微流控芯片烘干，以备循环利用。After the experiment, the microfluidic channel of the dispersed microfluidic chip of the droplet was rinsed with absolute ethanol, and then dried for recycling.

实施例6Example 6

本实施例以将所述微粒自夹流式微流控芯片作为微生物组液滴培养后的流式分选与细胞测序的耦合接口为例，提供将所述微粒自夹流式微流控芯片作为微生物组液滴培养后的流式分选与细胞测序的耦合接口的使用方法。In this embodiment, taking the microparticle self-entrapment microfluidic chip as the coupling interface of flow sorting and cell sequencing after microbiome droplet culture as an example, it provides the microparticle self-entrapment microfluidic chip Method for coupling interface of flow cytometry sorting and cell sequencing after group droplet culture.

如图22所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 22, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S41.液滴中微生物液体培养和扩增：S41. Liquid culture and expansion of microorganisms in droplets:

将含有微生物的选择培养基生成液滴，收集到EP管中放在培养箱中培养设定时间，使目标微生物大量繁殖，充满液滴；Generate droplets from the selective medium containing microorganisms, collect them into EP tubes and place them in an incubator for a set period of time, so that the target microorganisms multiply and fill the droplets;

其中，设定时间可以为12小时；Among them, the set time can be 12 hours;

S42.基于散射光的无标记的生长表型液滴分选：S42. Label-free sorting of growing phenotype droplets based on scattered light:

S43.单个液滴的自分散：S43. Self-dispersion of individual droplets:

利用所述液滴自分散式微流控芯片来进行液滴的分散，using the droplet to disperse the droplet from the dispersed microfluidic chip,

首先，将液滴信号采集与分选装置置于夹流结构4的通道处，打开***；First, place the droplet signal collection and sorting device at the channel of the entrainment structure 4, and open the system;

其次，将微粒进样口1连接含有菌液液滴的注射器，或用泵从微粒进样口1泵入菌液进入通道；或在微粒出样孔5连接负压驱动泵或注射器等驱动装置；Secondly, connect the particle inlet 1 to the syringe containing the droplet of bacterial liquid, or use a pump to pump the bacterial liquid from the particle inlet 1 into the channel; or connect the particle outlet 5 to a driving device such as a negative pressure driven pump or a syringe ;

再次，运行驱动装置，使得液滴流入所述微粒自夹流式微流控芯片，在夹流结构4中进行分散；Again, operate the driving device, so that the droplets flow into the microfluidic chip from the sandwiched flow microfluidic chip, and disperse in the sandwiched flow structure 4;

然后，调整液滴信号采集与分选软件参数，使其收集到液滴信号，并根据信号对其进行分选，使所需液滴选择性流入微粒出样孔5。Then, adjust the parameters of the droplet signal collection and sorting software so that the droplet signals are collected and sorted according to the signals, so that the required droplets selectively flow into the microparticle sampling hole 5 .

S44.得到目标微生物的生长状况信息：S44. Obtain the growth status information of the target microorganism:

将微粒出样孔5流出的液滴进行收集，通过测序可以得到目标微生物的多项信息。The droplets flowing out of the microparticle sampling hole 5 are collected, and multiple pieces of information about the target microorganism can be obtained through sequencing.

还可以包括S45.对所述微粒自夹流式微流控芯片进行再利用处理：It may also include S45. Reusing the microparticle self-entrapment microfluidic chip:

实验结束后，用无水乙醇冲洗所述微粒自夹流式微流控芯片的微流体通道，再将所述液滴自分散式微流控芯片烘干，以备循环利用。After the experiment, the microfluidic channel of the particle self-clamping microfluidic chip was rinsed with absolute ethanol, and then the liquid droplet self-distributed microfluidic chip was dried for recycling.

实施例7Example 7

本实施例以将所述微粒自夹流式微流控芯片作为微生物组液滴培养后的流式分选与质谱分析的耦合接口为例，提供将所述微粒自夹流式微流控芯片作为微生物组液滴培养后的流式分选与质谱分析的耦合接口的使用方法。In this embodiment, taking the microparticle self-clamping microfluidic chip as the coupling interface between flow sorting and mass spectrometry analysis after microbiome droplet culture, it provides the microparticle self-clamping microfluidic chip A method using the coupling interface of flow cytometry sorting and mass spectrometry after group droplet culture.

如图23所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 23, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S51.液滴中微生物液体培养和扩增：S51. Liquid culture and expansion of microorganisms in droplets:

S52.基于散射光的无标记的生长表型液滴分选：S52. Label-free sorting of growing phenotype droplets based on scattered light:

S53.单个液滴的自分散：S53. Self-dispersion of individual droplets:

利用所述微粒自夹流式微流控芯片来进行液滴的分散，Using the microparticles to disperse the droplets from the pinch-flow microfluidic chip,

其次，将微粒进样口1连接含有菌液液滴的注射器，或用泵从微粒进样口1泵入菌液进入通道，或在微粒出样孔5连接负压驱动泵或注射器等驱动装置；Secondly, connect the particle inlet 1 to the syringe containing the droplet of bacterial liquid, or use a pump to pump the bacterial liquid from the particle inlet 1 into the channel, or connect the particle outlet 5 to a driving device such as a negative pressure driven pump or a syringe ;

然后，调整液滴信号采集与分选软件参数，使其收集到液滴信号，并根据信号对其进行分选，使所需液滴选择性流入微粒出样孔5；Then, adjust the parameters of the droplet signal collection and sorting software, so that the droplet signal is collected, and sort it according to the signal, so that the required droplet selectively flows into the microparticle sampling hole 5;

S54.质谱分析：S54. Mass spectrometry analysis:

将微粒出样孔5流出的液滴进行收集，做质谱分析。The liquid droplets flowing out of the microparticle sampling hole 5 are collected for mass spectrometry analysis.

还可以包括步骤S55，对所述微粒自夹流式微流控芯片进行再利用处理：It may also include step S55, reusing the microparticle self-entrapment microfluidic chip:

实施例8Example 8

本实施例以将所述微粒自夹流式微流控芯片作为液滴酶活筛选后的与流式分选耦合接口为例，提供将所述微粒自夹流式微流控芯片作为液滴酶活筛选后的与流式分选耦合接口的使用方法。In this example, the microparticle self-entrapment microfluidic chip is used as an interface coupled with flow sorting after the droplet enzyme activity is screened, and the microparticle self-entrapment microfluidic chip is used as a droplet enzyme activity interface. The method of using the filtered interface coupled with flow sorting.

如图24所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 24, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S61.液滴中酶与底物反应：S61. Enzyme and substrate reaction in the droplet:

将含有底物、酶和染料等的溶液生成液滴，使酶、底物和染料充分反应；Generate droplets from a solution containing substrates, enzymes, and dyes to fully react the enzymes, substrates, and dyes;

S62.单个液滴的分散与荧光液滴分选：S62. Dispersion of individual droplets and sorting of fluorescent droplets:

利用所述液滴自分散式微流控芯片对收集的液滴进行分散与荧光检测，Using the droplet self-distributed microfluidic chip to disperse and detect the collected droplets,

其次，将微粒进样口1连接含有液滴的软管，软管另一端连接正压驱动泵或注射器等驱动装置，或在微粒出样孔5连接负压驱动泵或注射器等驱动装置；Secondly, connect the particle inlet 1 to a hose containing liquid droplets, and connect the other end of the hose to a drive device such as a positive pressure drive pump or a syringe, or connect a drive device such as a negative pressure drive pump or a syringe to the particle sample outlet 5;

再次，运行驱动装置，使液滴流入所述微粒自夹流式微流控芯片，在夹流结构4中进行自分散；Again, run the driving device to make the droplets flow into the microfluidic chip of the microparticle self-jamming flow, and perform self-dispersion in the sandwich structure 4;

S63.筛选到含有酶活强的液滴；S63. Screening out droplets containing strong enzyme activity;

将微粒出样孔5流出的液滴进行收集，筛选出能与底物反应的酶。The droplets flowing out of the microparticle sampling hole 5 are collected, and the enzymes capable of reacting with the substrate are screened out.

还可以包括步骤S45，对所述微粒自夹流式微流控芯片进行再利用处理：It may also include step S45, reusing the microparticle self-entrapment microfluidic chip:

实施例9Example 9

本实施例以所述微粒自夹流式微流控芯片应用于细胞自夹流为示范，提供通过基于流体力学的再注入式的所述微粒自夹流式微流控芯片实现紧密排列细胞的分散的使用方法。In this example, the microfluidic chip of the microparticle self-entrapment type is applied to the self-entrapment of cells as an example, and it provides a method to realize the dispersion of closely arranged cells through the reinjection type microfluidic chip of the microparticle self-entrapment type based on hydrodynamics. Instructions.

如图25所示，微粒自分散方法，采用如实施例1中任一项所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 25, the particle self-dispersion method is realized by using the particle self-entrapment microfluidic chip as described in any one of Example 1, comprising the following steps:

S71.接入细胞相进样端：S71. Access to the cell phase injection port:

将细胞相的进样端接入微粒进样孔1；Connect the injection port of the cell phase to the particle injection hole 1;

S72.接入压力驱动装置：S72. Access to the pressure drive device:

S73.分离细胞和夹流液：S73. Separation of cells and entrained fluid:

驱动步骤S71接入的正压驱动装置或负压驱动装置，将步骤S71接入进样端的细胞相通过微粒进样孔1流入所述微粒自夹流式微流控芯片；细胞相在主通道2下游和侧通道3下端分别分离出细胞和夹流液；Drive the positive pressure drive device or negative pressure drive device connected in step S71, and the cell phase connected to the injection end in step S71 flows into the particle self-entrapment microfluidic chip through the particle injection hole 1; the cell phase is in the main channel 2 The downstream and the lower end of the side channel 3 separate the cells and the entrained fluid respectively;

S74.汇集细胞和夹流液及细胞自夹流：S74. Pooled cells and entrainment fluid and cell self-entrainment:

使得步骤S73中分离出的细胞和夹流液在夹流结构4汇集，通过夹流液将细胞自分散；The cells separated in step S73 and the entrainment fluid are collected in the entrainment structure 4, and the cells are self-dispersed by the entrainment fluid;

S75.得到分散后细胞(如图18所示)：S75. Obtain the dispersed cells (as shown in Figure 18):

将步骤S74分散后的细胞经微粒出样孔5流出。The cells dispersed in step S74 flow out through the microparticle sampling hole 5 .

上述实施例中，在进行步骤S73时，分离细胞和夹流液中，将步骤S71接入进样端的细胞相通过微粒进样孔1流入所述微粒自夹流式微流控芯片时，细胞相通过气泡拦截结构6将气泡拦截，使得气泡无法进入下游所述微粒自夹流式微流控芯片结构。In the above embodiment, when step S73 is performed, the cells and the entrainment fluid are separated, and the cell phase that is connected to the injection end in step S71 flows into the microparticles from the entrainment microfluidic chip through the particle injection hole 1, the cell phase Bubbles are intercepted by the bubble interception structure 6, so that the bubbles cannot enter the microfluidic microfluidic chip structure downstream.

上述实施例中，在进行步骤S73时，分离细胞和夹流液中，将步骤S71接入进样端的细胞相通过微粒进样孔1流入所述微粒自夹流式微流控芯片后，由于主通道2与侧通道3交界处的微粒拦截结构7，或由于侧通道3相对于主通道2的内径小，或由于以上两结构的结合，使得细胞周围流体进入侧通道3，而细胞无法进入侧通道3，实现细胞相在主通道2下游和侧通道3下端分别分离出细胞和流体。In the above embodiment, when step S73 is performed, the cells and the entrainment fluid are separated, and the cell phase that is connected to the injection port in step S71 flows into the microparticles from the entrainment microfluidic chip through the particle injection hole 1, due to the main The particle intercepting structure 7 at the junction of the channel 2 and the side channel 3, or because the inner diameter of the side channel 3 is smaller than that of the main channel 2, or because of the combination of the above two structures, the fluid around the cells enters the side channel 3, and the cells cannot enter the side channel. Channel 3 realizes the separation of cells and fluid from the cell phase at the downstream of the main channel 2 and the lower end of the side channel 3, respectively.

上述实施例中，在进行步骤S75时，得到分散后细胞后，若一段时间内不会再用该所述微粒自夹流式微流控芯片，则可以将所述微粒自夹流式微流控芯片取下，用无水乙醇将主通道2冲洗干净，放入烘箱烘干后，可以循环再利用。同时也可以用无水乙醇将侧通道3、夹流结构4等的通道清洗干净，即将微流体通道整体清洗干净。In the above-mentioned embodiment, when step S75 is performed, after the dispersed cells are obtained, if the particles will not be used again for a period of time from the sandwiched flow microfluidic chip, the particles can be self-tipped from the sandwiched flow microfluidic chip Take it off, rinse the main channel 2 with absolute ethanol, put it in an oven for drying, and then it can be recycled. At the same time, the channels such as the side channel 3 and the entrainment structure 4 can also be cleaned with absolute ethanol, that is, the entire microfluidic channel is cleaned.

实施例10Example 10

本实施例以所述微粒自夹流式微流控芯片应用于颗粒自夹流为示范，提供通过基于流体力学的再注入式的所述微粒自夹流式微流控芯片实现紧密排列颗粒的分散的使用方法。In this embodiment, the application of the microparticle self-entrapment microfluidic chip to particle self-entrapment is used as an example, and it is provided to realize the dispersion of closely arranged particles through the reinjection-based microparticle self-entrapment microfluidic chip based on hydrodynamics. Instructions.

如图26所示，微粒自分散方法，采用如实施例1所述的微粒自夹流式微流控芯片实现，包括以下步骤：As shown in Figure 26, the method of particle self-dispersion is realized by using the particle self-clamping microfluidic chip as described in Example 1, including the following steps:

S81.接入颗粒相进样端：S81. Connecting to the injection port of the granular phase:

将颗粒相的进样端接入微粒进样孔1；Connect the injection port of the particle phase to the particle injection hole 1;

S82.接入压力驱动装置：S82. Access to the pressure drive device:

将颗粒进样孔1接入正压驱动装置，或将微粒出样孔5接入负压驱动装置；Connect the particle sampling hole 1 to the positive pressure driving device, or connect the particle sampling hole 5 to the negative pressure driving device;

S83.分离颗粒和夹流液：S83. Separation of particles and entrained fluid:

驱动步骤S81接入的正压驱动装置或负压驱动装置，将步骤S81接入进样端的颗粒相通过微粒进样孔1流入所述微粒自夹流式微流控芯片；颗粒相在主通道2下游和侧通道3下端分别分离出颗粒和夹流液；Drive the positive pressure drive device or negative pressure drive device connected in step S81, and the particle phase connected to the injection end in step S81 flows into the particle self-entrapment microfluidic chip through the particle injection hole 1; the particle phase is in the main channel 2 Particles and entrained liquid are separated from the downstream and the lower end of the side channel 3, respectively;

S84.汇集颗粒和夹流液及颗粒自夹流：S84. Collecting particles and entrained liquid and particle self-entrained flow:

使得步骤S83中分离出的颗粒和夹流液在夹流结构4汇集，通过夹流液将颗粒自分散；The particles separated in step S83 and the entrainment liquid are collected in the entrainment structure 4, and the particles are self-dispersed by the entrainment liquid;

S85.得到分散后颗粒(如图18所示)：S85. Obtain the dispersed particles (as shown in Figure 18):

将步骤S14分散后的颗粒经微粒出样孔5流出。The particles dispersed in step S14 flow out through the particle sampling hole 5 .

上述实施例中，在进行步骤S83时，分离颗粒和夹流液中，将步骤S81接入进样端的颗粒相通过微粒进样孔1流入所述微粒自夹流式微流控芯片时，颗粒相通过气泡拦截结构6将气泡拦截，使得气泡无法进入下游所述微粒自夹流式微流控芯片结构。In the above embodiment, when step S83 is performed, the particles and the entrained liquid are separated, and the particle phase that is connected to the injection port in step S81 flows into the particle through the particle injection hole 1. When the particle self-entrapment microfluidic chip, the particle phase Bubbles are intercepted by the bubble interception structure 6, so that the bubbles cannot enter the microfluidic microfluidic chip structure downstream.

上述实施例中，在进行步骤S83时，分离颗粒和夹流液中，将步骤S81接入进样端的颗粒相通过微粒进样孔1流入所述微粒自夹流式微流控芯片后，由于主通道2与侧通道3交界处的微粒拦截结构7，或由于侧通道3相对于主通道2的内径小，或由于以上两结构的结合，使得颗粒周围流体进入侧通道3，而颗粒无法进入侧通道3，实现颗粒相在主通道2下游和侧通道3下端分别分离出颗粒和流体。In the above-mentioned embodiment, when step S83 is performed, the particles and the entrainment liquid are separated, and the particle phase that is connected to the injection port in step S81 flows into the particles from the entrainment microfluidic chip through the particle injection hole 1, due to the main The particle intercepting structure 7 at the junction of the channel 2 and the side channel 3, or because the inner diameter of the side channel 3 is smaller than that of the main channel 2, or because of the combination of the above two structures, the fluid around the particles enters the side channel 3, and the particles cannot enter the side channel. The channel 3 realizes the separation of the particle phase from the particles and the fluid at the downstream of the main channel 2 and the lower end of the side channel 3 respectively.

上述实施例中，在进行步骤S85时，得到分散后颗粒后，若一段时间内不会再用该所述微粒自夹流式微流控芯片，则可以将所述微粒自夹流式微流控芯片取下，用无水乙醇将主通道2冲洗干净，放入烘箱烘干后，可以循环再利用。同时也可以用无水乙醇将侧通道3、夹流结构4等的通道清洗干净，即将微流体通道整体清洗干净。In the above embodiment, when step S85 is carried out, after the dispersed particles are obtained, if the particles will not be used again for a period of time from the sandwiched flow microfluidic chip, the particles can be self-soldered from the sandwiched flow microfluidic chip Take it off, rinse the main channel 2 with absolute ethanol, put it in an oven for drying, and then it can be recycled. At the same time, the channels such as the side channel 3 and the entrainment structure 4 can also be cleaned with absolute ethanol, that is, the entire microfluidic channel is cleaned.

最后应说明的是：以上实施例仅用以说明本发明的技术方案，而非对其限制；尽管参照前述实施例对本发明进行了详细的说明，本领域的普通技术人员应当理解：其依然可以对前述各实施例所记载的技术方案进行修改，或者对其中部分技术特征进行等同替换；而这些修改或者替换，并不使相应技术方案的本质脱离本发明各实施例技术方案的精神和范围。Finally, it should be noted that: the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: it can still be Modifications are made to the technical solutions described in the foregoing embodiments, or equivalent replacements are made to some of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the various embodiments of the present invention.

Claims

微粒自夹流式微流控芯片，其特征在于，所述微粒自夹流式微流控芯片内设置至少一条微流体通道，所述微流体通道包括主通道、至少一段侧通道和夹流结构；The particle self-entrapment microfluidic chip is characterized in that at least one microfluidic channel is arranged in the particle self-entrapment microfluidic chip, and the microfluidic channel includes a main channel, at least one side channel and an entrainment structure;

所述主通道上端设置微粒相入口，所述夹流结构下端设置微粒出口；The upper end of the main channel is provided with a particle phase inlet, and the lower end of the entrainment structure is provided with a particle outlet;

所述侧通道上端与所述主通道上游连通连接，所述微流体通道设置阻止微粒进入所述侧通道的微粒阻截结构，所述侧通道下端与所述主通道下游连通连接，所述夹流结构位于所述侧通道下端与所述主通道下游连通连接处；The upper end of the side channel communicates with the main channel upstream, the microfluidic channel is provided with a particle blocking structure that prevents particles from entering the side channel, the lower end of the side channel communicates with the main channel downstream, and the entrainment The structure is located at the connection between the lower end of the side channel and the downstream of the main channel;

所述微粒自夹流式微流控芯片上设置微粒进样孔和微粒出样孔，所述微粒进样孔与所述微粒相入口连通连接，所述微粒出样孔与所述微粒出口连通连接。A microparticle sampling hole and a microparticle sampling hole are provided on the microparticle self-clamping microfluidic chip, the microparticle sampling hole communicates with the microparticle phase inlet, and the microparticle sampling hole communicates with the microparticle outlet .
如权利要求1所述的微粒自夹流式微流控芯片，其特征在于，所述侧通道与所述主通道连接处高度高于所述侧通道高度构成高度差值；所述微粒阻截结构为设置在所述侧通道与所述主通道连接处的微粒阻拦结构，或为所述侧通道的宽度小于所述液滴直径所构成的尺寸差结构，或为所述高度差值小于所述液滴直径所构成的尺寸差结构，或为其中二者或三者结构的组合。The particle self-entrapment microfluidic chip according to claim 1, wherein the height of the connection between the side channel and the main channel is higher than the height of the side channel to form a height difference; the particle blocking structure is The particle blocking structure arranged at the junction of the side channel and the main channel is either a size difference structure formed by the width of the side channel being smaller than the diameter of the droplet, or a structure in which the height difference is smaller than the droplet diameter. The size difference structure formed by the diameter of the droplet, or a combination of two or three of them.
如权利要求2所述的微粒自夹流式微流控芯片，其特征在于，所述微粒阻截结构为设置在所述侧通道与所述主通道连接处的微粒阻拦结构时，所述微粒阻拦结构为微筛阵列结构。The particle self-entrapment microfluidic chip according to claim 2, wherein when the particle blocking structure is a particle blocking structure arranged at the connection between the side channel and the main channel, the particle blocking structure It is a microsieve array structure.
如权利要求1所述的微粒自夹流式微流控芯片，其特征在于，在所述微粒进样孔处设置气泡阻拦结构。The particle self-entrapment microfluidic chip according to claim 1, characterized in that a bubble blocking structure is arranged at the particle injection hole.
如权利要求4所述的微粒自夹流式微流控芯片，其特征在于，在所述气泡阻拦结构为微筛阵列结构。The particle self-encapsulating microfluidic chip according to claim 4, wherein the bubble blocking structure is a microsieve array structure.
如权利要求1所述的微粒自夹流式微流控芯片，其特征在于，在所述主通道下游与所述夹流结构之间设置至少一段蛇形通道。The particle self-encapsulating microfluidic chip according to claim 1, wherein at least one serpentine channel is arranged between the downstream of the main channel and the entraining structure.
如权利要求1所述的微粒自夹流式微流控芯片，其特征在于，在所述主通道下游设置至少一个的支通道，至少一个所述支通道下游与所述侧通道下端连通连接于所述夹流结构。The particle self-entrapping microfluidic chip according to claim 1, wherein at least one branch channel is arranged downstream of the main channel, and at least one branch channel downstream communicates with the lower end of the side channel and is connected to the said side channel. The entrainment structure described above.
如权利要求1所述的微粒自夹流式微流控芯片，其特征在于，在所述微粒进样孔上游设置正压驱动装置，或在所述微粒出样孔下游设置负压驱动装置。The particle self-entrapping microfluidic chip according to claim 1, characterized in that a positive pressure driving device is arranged upstream of the particle sampling hole, or a negative pressure driving device is arranged downstream of the particle sampling hole.
一种如权利要求1-8中任一项所述微粒自夹流式微流控芯片的制造方法，其特征在于，包括以下步骤：A method for manufacturing a microparticle self-clamping microfluidic chip according to any one of claims 1-8, characterized in that it comprises the following steps:

制备硅胶模板：Prepare the silica template:

通过制图软件设计具有所述主通道的主通道结构和具有所述侧通道的侧通道结构，并进行打印；Designing the main channel structure with the main channel and the side channel structure with the side channel by drawing software, and printing;

将硅片滴上光刻胶后甩胶，并将其覆盖掩膜所述侧通道结构，然后曝光；Dropping the photoresist on the silicon wafer and then throwing the glue, and covering it with the side channel structure of the mask, and then exposing;

将曝光后的硅片再次甩胶后，将其覆盖掩膜所述主通道结构，使得所述主通道结构与所述侧通道结构对准，然后曝光；After the exposed silicon wafer is shaken again, cover and mask the main channel structure so that the main channel structure is aligned with the side channel structure, and then expose;

将未固化部分通过显影液清洗，得到所述硅片模板；Cleaning the uncured part with a developer to obtain the silicon wafer template;

制备具有通道结构的上层PDMS芯片：Prepare the upper layer PDMS chip with channel structure:

将PDMS单体与固化剂混合得到PDMS高聚物；Mix PDMS monomer and curing agent to obtain PDMS polymer;

将所述PDMS高聚物倒在所述硅片模板上，烘干后得到具有所述主通道结构和所述侧通道结构的所述上层PDMS芯片；Pour the PDMS polymer onto the silicon wafer template and dry to obtain the upper PDMS chip with the main channel structure and the side channel structure;

制备所述上层PDMS芯片上的所述微粒进样孔和所述微粒出样孔：Prepare the microparticle sampling hole and the microparticle sampling hole on the upper PDMS chip:

将具有所述通道结构的所述上层PDMS芯片打出所述微粒进样孔和所述微粒出样孔；punching out the microparticle sampling hole and the microparticle sampling hole from the upper PDMS chip having the channel structure;

将待键合的下层芯片采用无刻蚀图案的PDMS光滑基片；Use a PDMS smooth substrate without etching patterns for the underlying chip to be bonded;

制备所述微粒自夹流式微流控芯片：Preparation of the microparticle self-trapping microfluidic chip:

将所述上层PDMS芯片与所述下层芯片等离子体键合制得所述微粒自夹流式微流控芯片，并经过放置后，恢复所述微粒自夹流式微流控芯片的疏水性。Plasma-bonding the upper layer PDMS chip and the lower layer chip to prepare the particle self-entrapment microfluidic chip, and after being placed, the hydrophobicity of the particle self-entrapment microfluidic chip is restored.
一种采用如权利要求1-8中任一项所述微粒自夹流式微流控芯片实现的微粒自分散方法，其特征在于，包括以下步骤：A self-dispersing method for microparticles realized by a microfluidic chip self-clamping microfluidic chip according to any one of claims 1-8, characterized in that it comprises the following steps:

接入微粒相的进样端：将所述微粒相的进样端接入所述微粒进样孔；Connecting to the injection port of the particle phase: connecting the sample port of the particle phase to the particle injection hole;

接入压力驱动装置：将所述微粒进样孔接入正压驱动装置，或将所述微粒出样孔接入负压驱动装置；Connecting to the pressure driving device: connecting the particle sampling hole to a positive pressure driving device, or connecting the particle sampling hole to a negative pressure driving device;

分离微粒和流体：驱动所述正压驱动装置或所述负压驱动装置，将所述微粒相的进样端的液滴相通过所述微粒进样孔流入所述微粒自夹流式微流控芯片；所述微粒相在所述主通道下游和所述侧通道下端分别分离出微粒和夹流液；Separation of particles and fluid: drive the positive pressure drive device or the negative pressure drive device, and flow the liquid droplet phase at the injection end of the particle phase into the particle self-pinching microfluidic chip through the particle injection hole ; The particle phase is separated from the particles and the entrainment liquid at the downstream of the main channel and the lower end of the side channel, respectively;

汇集所述微粒和所述夹流液及微粒分散：使得分离出的所述微粒和夹流液在所述夹流结构汇集，通过所述夹流液将所述微粒自分散；Collecting the particles, the entraining liquid, and dispersing the particles: making the separated particles and the entraining liquid gather in the entrainment structure, and self-dispersing the particles through the entraining liquid;

得到自分散后所述微粒：将自分散后的所述微粒经所述微粒出样孔流出。Obtaining the microparticles after self-dispersion: the self-dispersed microparticles flow out through the microparticle sampling hole.