CN114405568B - A self-driving microfluidic chip - Google Patents

Abstract

The invention provides a self-driven microfluidic chip for immunofluorescence detection, the main part of which includes a sample injection area for injecting target analytes; an antibody release area for releasing fluorescent substance-antibody complexes for analysis with the target The channel detection area is used to capture the fluorescent substance-antibody-antigen complex formed after the sample passes through the antibody release area, so as to perform fluorescence detection; it is characterized in that, in the front section of the channel detection area, there is a channel that bends up and down as a whole The structure of the channel, that is, the channel fluctuates up and down from the outside, so that when the liquid in the channel passes through the curved structure, it will not only produce horizontal displacement, but also produce up and down displacement, thereby generating a series of vortex flow fields, so that the reactants are fully mixed. , and the time for the fluid to pass can be adjusted by setting the number and length of the curved structures.

Description

A self-driving microfluidic chip

The invention relates to a microfluidic chip, in particular to a self-driving microfluidic chip that can be used for immunofluorescence detection.

Background technique

With the development of immune technology, point of care (POCT), as a new field in laboratory medicine, has attracted more and more attention. It is a new type of method for sampling and analyzing immediately at the patient's site, eliminating the complicated processing procedures of specimens in the laboratory, and quickly obtaining test results, which is of great significance to emergency rescue and other aspects. It has a wide range of applications in blood sugar detection, cardiovascular disease detection, infectious diseases, drug abuse, pregnancy monitoring, etc. The technical principles of POCT mainly include: colloidal gold, dry immunofluorescence, microfluidic immunofluorescence, etc. At present, colloidal gold and dry immunofluorescence have become mainstream technologies in the market, while microfluidics will be the mainstream direction of future development.

Both colloidal gold and dry immunofluorescence detection techniques are based on immunochromatography, and generally require the use of nitrocellulose membranes as the substrate for lateral flow of liquid. Their common advantages are simple and fast operation, high sensitivity and specificity. However, the biggest challenge faced by this immunochromatographic technique is poor repeatability, mainly because a variety of thin and fragile materials need to be pasted together while ensuring the consistency of liquid flow in them. The solution is to completely abandon chromatographic membranes and other thin films and use microfluidic chips.

The microfluidic chip is based on micro-processing, with micro-channel network as the structural feature, without the use of chromatographic media, and its goal is to integrate sampling, pretreatment, reagent addition, mixing, Reaction, separation, detection and other operating units are partially or fully integrated on a chip with micron-scale channels. It is light in size, and the liquid flows directly in the air; it uses a small amount of samples and reagents; it has a fast reaction speed and can be processed in parallel. Due to its simple structure, self-driven microfluidic chip uses capillary force as the driving force source, does not need auxiliary external air pumps and other equipment, and is easy and reliable to use. It has become the mainstream direction of immunofluorescence POCT detection development.

For the microfluidic chip for immunofluorescence detection of antigen detection based on the double-sandwich principle, the main steps of its work are 1. The sample is injected into the sample injection part; 2. The antigen in the sample combines with the fluorescent-antibody complex in the antibody release area to form Fluorescent substance-antibody-antigen complex; 3. In the channel part, the fluorescent substance-antibody-antigen complex formed in the antibody release area is captured by the antibody pre-dotted in the detection area of the channel part, thereby forming a fluorescent substance-antibody in the detection area - Antigen-antibody double-antibody sandwich substance, the fluorescence detection device can detect the fluorescence intensity of this area to calculate the amount of antigen. Every working step in the microfluidic chip has the problem of binding efficiency, and the binding efficiency of each step directly affects the detection results. The binding efficiency is related to factors such as the mixing degree of the sample and the reaction time. At the same time, the number of bubbles and the smoothness of the liquid surface flow will also affect the final detection efficiency.

At present, the self-driven microfluidic chip technology used for clinical immunofluorescence detection is mainly the microfluidic chip technology owned by South Korea Nanon Technology Co., Ltd., and its patent (authorized announcement number: CN 102305867B) discloses a " A chip for analyzing fluids that do not use external energy to move", this chip designs a self-driven microfluidic chip for analysis, which is divided into three parts: pretreatment part, channel part and cleaning part. Realize the content of steps 1 and 2 in the main steps of the work of the above-mentioned microfluidic chip, and the channel part realizes the content of the above-mentioned step 3. Its main feature is that the pretreatment part has a sample injection part and a first buffer part, and the first phase buffer part has a step difference relative to the sample injection part; the pretreatment part also has a sample guide part, which is arranged between the sample injection part and the first buffer part. Between the parts, it is used to break the surface tension of the fluid from the sample injection part to the first buffer part, so as to achieve the purpose of stabilizing the fluid. The main invention of this invention focuses on the period between the injection of the sample and the first buffer part, and the fluid in this part can be relatively stable through related design. However, this invention lacks an effective structural design from the first buffer part to the channel part, so as to ensure that the sample is fully mixed, various substances are fully combined, and less air bubbles and smooth liquid surface flow are ensured.

Therefore, it is necessary to design a new type of self-driven microfluidic chip for immunofluorescence detection, and then all parts of its function are precisely designed to improve the relevant binding efficiency, mixing degree, control the reaction time, reduce air bubbles, Ensure smoothness of liquid surface flow.

Contents of the invention

A self-propelled microfluidic chip for immunofluorescence detection, comprising: a sample injection area for injecting target analytes; an antibody release area for releasing fluorescent substance-antibody complexes to combine with target analytes; channel detection The area is used to capture the fluorescent substance-antibody-antigen complex formed after the sample passes through the antibody release area, so as to perform fluorescence detection; in the front section of the channel detection area, there is a structure that makes the channel bend up and down as a whole, so that the liquid in the channel When passing through the curved structure, not only horizontal displacement, but also up and down displacement will be generated, thereby generating a series of vortex flow fields to fully mix the reactants.

The self-propelled microfluidic chip for immunofluorescence detection is characterized in that the height of the front end of the antibody release area is gradually increased, and the fluid within this channel height range can slow down the flow rate, and the larger channel height can effectively contain Excessive bending of the flow surface reduces the generation of air bubbles in the fluid.

The self-propelled microfluidic chip for immunofluorescence detection is characterized in that there is a vent hole near the central axis of the end of the antibody release area, which is used to reduce the generation of liquid due to height difference and surface tension effect during the flow of the antibody release area. bubbles.

The self-propelled microfluidic chip for immunofluorescence detection is characterized in that the initial height of the front end of the antibody release region ranges from 50 μm to 100 μm, and the end height ranges from 100 μm to 500 μm.

The self-propelled microfluidic chip for immunofluorescence detection is characterized in that the number of curved structures in the front section of the channel detection area can be one or more.

The self-propelled microfluidic chip for immunofluorescence detection is characterized in that the vertical displacement generated by the curved structure in the front section of the channel detection area can be adjusted as required, and the value range can be 100 μm to 500 μm.

The beneficial effects of the present invention are:

1. In the absence of external drive, by designing a structure that bends the channel up and down in the channel detection area, the fluid flows up and down in the channel, and induces a vortex flow field in the channel detection area, thereby enhancing the antigen-antibody binding reaction Efficiency, and effectively avoid the generation of air bubbles, to ensure the diversion of the channel. In addition, this structure increases the volume of the channel, fully ensuring the time required for the combination of antigens and antibodies before entering the detection area.

2. The fluid injected into the antibody release area, and as the height increases, the flow rate slows down, providing sufficient time for antibody release, and fully evolves into a vortex structure flow field in the antibody release area that helps antibody diffusion and mixing , the surface tension provided by the upper and lower walls and side walls of the flow channel plays a key role in the formation of the vortex flow field.

3. Surface tension can cause the liquid flow velocity near the side wall to be faster than that in the middle area, resulting in the formation of an uneven flow surface. The larger channel height of the antibody release area can effectively curb the excessive bending of the flow surface and reduce the generation of air bubbles in the fluid, thereby simplifying the design of the exhaust structure.

4. The vent holes near the central axis at the end of the antibody release area can effectively reduce the possibility of bubbles due to height difference and surface tension effects when the liquid flows from the antibody release area into the channel detection area.

Description of drawings

Fig. 1 is a schematic diagram of the overall structure of the channel of the microfluidic chip of the present invention.

Fig. 2 is a streamline diagram of liquid velocity in the microfluidic chip of the present invention simulated by software.

Fig. 3 is a streamline diagram of liquid velocity in the raised structure in the microfluidic chip of the present invention simulated by software.

Fig. 4 is a schematic diagram of the promotion effect of the raised structure in the microfluidic chip of the present invention on the smoothness of the liquid surface simulated by software.

Detailed ways

Below in conjunction with accompanying drawing illustrate concrete implementation method:

Fig. 1 is a schematic diagram of the overall structure of the flow channel of the microfluidic chip of the present invention, which is divided into four parts as a whole, namely, 110 sample injection area, 120 antibody release area, 130 channel detection area, and 140 front end of waste liquid area. The part displayed as a whole is the flow channel of the microfluidic chip, that is, the sample enters the microfluidic chip from 110, passes through the antibody release area at 120, the channel detection area at 130, and the front end of the waste liquid area at 140. In the figure: 110 is a sample injection area, which is used to inject target analyte, and many micro-columns are distributed on it, which is used to expand the specific surface area and strengthen the mixing of the sample and the pre-set at 110. Part 120 is the antibody release area, wherein 121 is the front end of the antibody release area, which acts as a guide. The initial height of the front end of the antibody release area ranges from 50 μm to 100 μm, and the end height ranges from 100 μm to 500 μm. The liquid in 121 The height is gradually increased and the flow rate is slowed down to provide sufficient time for antibody release. 122 is the fluorescent substance-antibody complex spotting area. In this part of the area, the antigen to be tested can be fully mixed and combined with the fluorescent substance-antibody complex to form a fluorescent substance-antibody-antigen complex complex. In this part, The mixing degree and mixing time of the two substances are very critical factors, which can be achieved through structural design. 123 is an air vent near the central axis of the end of the antibody release area, which is used to reduce the bubbles generated by the height difference and surface tension effect during the flow of the liquid in the antibody release area. 130 is a channel detection area, which is used to capture the fluorescent substance-antibody-antigen complex formed after the sample passes through the antibody release area, so as to perform fluorescence detection. Among them, 131 is a structure in the front section of the channel detection area that makes the channel bend up and down as a whole, that is, the channel is shown as up and down from the outside, so that when the liquid in the channel passes through the curved structure, it will not only generate horizontal displacement, but also produce up and down. Displacement, this structure can be designed into one or more up and down structures according to the relevant properties of the reactants, such as viscosity, etc., and the flow time of the fluid in the channel is also directly related to the number of the structures and the height of the ups and downs of the structures Relatedly, the up and down displacement can be adjusted as required, and the value range can be from 100 μm to 500 μm, so as to adjust the time for the fluid to pass. 140 is the front end of the waste liquid area, and the shape and size of the waste liquid area behind it can be set freely, depending on the amount of liquid to be accommodated.

Fig. 2 is a streamline diagram of liquid velocity in the microfluidic chip of the present invention simulated by software based on the finite element method. The lines in the figure represent the velocity streamlines, which can represent the flow trend of the fluid in the flow channel. The fluid injected into the antibody release area 120, as the height in 121 increases, the flow velocity slows down, providing sufficient time for antibody release, and fully evolves in the area 122 to help the diffusion of the fluorescent substance-antibody complex and its interaction with the target fluid. The up and down symmetrical vortex structure flow field where the sample is mixed. The surface tension provided by the upper and lower walls and side walls of the flow channel plays a key role in the formation of the vortex flow field. At the position of the up and down raised structure of 131, with the symmetry axis of the microfluidic chip as the center of symmetry, two similar vortex flow fields are also generated, thereby further strengthening the interaction between the fluorescent substance-antibody complex and the analyte. Mix and combine.

FIG. 3 is a streamline diagram of liquid velocity in the up and down curved structure 131 in the microfluidic chip of the present invention simulated by software based on the finite element method. In the figure, there are two up and down curved convex structures, which are side views. It can be seen from the figure that a strong vortex flow field is generated inside the raised structure, especially the rising part, which will also strengthen the mixing and binding of the fluorescent substance-antibody complex and the analyte, improving the binding Efficiency, thereby improving detection sensitivity, in practical applications, the raised structure can be designed according to actual needs.

Fig. 4 is a schematic diagram of the speed-promoting effect of the raised structure in the microfluidic chip of the present invention on the smoothness of the liquid surface simulated by software. The streamlines in the figure are the velocity streamlines in the liquid, and at the same time, the frontmost liquid level of the fluid is indicated by arrows. Figure 4(a) is a channel without a raised structure; Figure 4(b) is a channel with two raised structures. As can be seen from the figure, for the microfluidic chip with a raised structure in the present invention, as the height of the liquid increases at part 131, its flow velocity slows down, so that the liquid surface located at the raised part (indicated by the arrow in the figure) part) tends to be perpendicular to the direction of the flow channel, that is, a flat liquid surface is formed, which can effectively avoid the generation of air bubbles, so that there is no need to design an additional exhaust structure in the channel detection area 130 . For the previous simulation of the microfluidic chip without the raised structure, it can be seen that the liquid surface (the part indicated by the arrow in the figure) is curved, and the relatively flat liquid surface is more likely to generate bubbles.

Claims (5)

1. A self-driven microfluidic chip for immunofluorescence detection, comprising: a sample injection area for injecting target analytes; The antibody release area is used to release the fluorescent substance-antibody complex and bind to the target analyte; The height of the front end of the antibody release area is gradually increased. The fluid within this channel height range can slow down the flow velocity. The larger channel height can effectively curb the excessive bending of the flow surface and reduce the generation of air bubbles in the fluid; The channel detection area is used to capture the fluorescent substance-antibody-antigen complex formed after the sample passes through the antibody release area, so as to perform fluorescence detection; In the front section of the channel detection area, there is a structure that makes the channel bend up and down as a whole, so that when the liquid in the channel passes through the curved structure, it will not only generate horizontal displacement, but also produce up and down displacement, thereby generating a series of vortex flow fields. Mix the reactants thoroughly. 2. The self-propelled microfluidic chip that is used for immunofluorescence detection as claimed in claim 1, is characterized in that, there is a vent hole near the central axis of the end of the antibody release area, which is used to reduce the liquid due to height in the flow process of the antibody release area. Bubbles generated by differential and surface tension effects. 3. The self-propelled microfluidic chip for immunofluorescence detection according to claim 1, wherein the initial height of the front end of the antibody release region ranges from 50 μm to 100 μm, and the end height ranges from 100 μm to 500 μm. 4. The self-propelled microfluidic chip for immunofluorescence detection according to claim 1, wherein the number of curved structures in the front section of the channel detection area can be one or more. 5. The self-propelled microfluidic chip for immunofluorescence detection according to claim 1, wherein the up and down displacement generated by the curved structure in the front section of the channel detection area can be adjusted according to needs, and the value range can be 100 μm to 500 μm.