Paper-based microfluidic chips, microfluidic detection systems, liquid detection methods, and their applications

Abstract

The present invention discloses a paper-based microfluidic chip, a microfluidic detection system, a liquid detection method, and their uses. The chip includes a paper substrate layer (2), and a detection cell (21) is provided on the paper substrate layer (2). The detection cell (21) has a first contact angle region (211) and a second contact angle region (212) therein, and the contact angle of the liquid in the detection cell (21) at the first contact angle region (211) is larger than the contact angle at the second contact angle region (212). Based on the mechanism of chromaticity transfer on the paper chip, the surface tension of the liquid is adjusted and controlled to provide a driving force toward the inside of the droplet, thereby causing the droplet to shrink from the first contact angle region (211) to the second contact angle region (212) with a smaller contact angle, and the chromaticity is transferred to the second contact angle region (212) and concentrated, thereby improving the chromaticity and uniformity per unit area under the premise that the concentration of the object to be measured is the same, improving the detection sensitivity, lowering the detection limit, and widening the spot collection range, thereby improving the handling of spot collection and the detection reproducibility.

Description

Detailed Description of the Invention

[0001] [Cross-Reference to Related Applications] The present application claims the rights and interests of Chinese patent applications 202211379392.8, 202222939918.5, 202222939919.X, and 202211379390.9 filed on November 4, 2022, and the content of the said applications is incorporated herein by reference. [Technical Field]

[0002] The present invention relates to microfluidic detection technology, and specifically to a paper-based microfluidic chip. Based on this, the present invention also relates to a microfluidic detection system comprising the paper-based microfluidic chip, and a liquid detection method using the paper-based microfluidic chip. Furthermore, the present invention also relates to uses of the paper-based microfluidic chip, the microfluidic detection system, and the liquid detection method. [Background Art]

[0003] Microfluidic technology has the advantages of fast mass and heat transfer, high analysis efficiency, low reagent consumption, low analysis cost, environmental friendliness and easy integration, and is also easily adaptable to small portable detection and analysis instruments, so it is expected to have promising application prospects in the fields of water quality detection, environmental detection, food and medical treatment, etc. While microfluidic technology has brought a new development direction to detection and analysis instruments such as portable water quality detectors, problems such as additional fluid control requirements in microfluidic technology and the increased volume of detection instruments have brought new challenges to the portability of instruments.

[0004] Paper-based microfluidic chips can effectively address these challenges in terms of fluid control. These paper-based microfluidic chips, known as "paper chips," are microfluidic analysis technology platforms that achieve self-propulsion by relying on capillary force. Compared to microfluidic chips using other substrates, paper chips are characterized by their low cost and portability, making them promising for fields such as water quality detection, environmental detection, and food and medical applications. With the continuous development and advancement of smartphone imaging technology and software capabilities, it is now possible to capture images of the detection cells of paper chips using mobile phones, enabling chromaticity identification and colorimetric analysis through embedded software. This eliminates the need for additional signal analysis equipment, and the combination of paper chips and mobile phone imaging analysis further enhances the portability of detection devices.

[0005] However, due to limitations in reagent capacity, non-uniformity of paper materials, and issues such as insufficient color development, reduced uniformity, and decreased reproducibility caused by capillary action (e.g., the coffee ring effect), detection using paper chips suffers from low sensitivity, a low detection limit, and insufficient accuracy and reproducibility, which are common problems that limit the development of paper chip detection technology.

[0006] To improve the sensitivity and accuracy of paper chip detection, researchers have devised various methods, such as designing bidirectional liquid inlet channels, pre-positioning reagents on both sides of the detection cell, and ensuring that reagents are driven by the target liquid into the detection cell from both sides, thereby reducing chromaticity diffusion to the edges of the detection cell and improving chromaticity uniformity of the detection cell. Another example is utilizing the concentration effect of the coffee ring effect, where the detection limit of the target substance can be lowered by extracting color from the coffee ring generated by the color reaction. Furthermore, conventional techniques have proposed improving detection sensitivity by utilizing the adsorption effect of noble metal nanoparticles or carbon quantum dots, and supporting these adsorbed substances on paper chips to concentrate the target substance. However, the above methods still have the following shortcomings: Improving the uniformity of color development allows for a uniform distribution of chromaticity, but it does not improve detection sensitivity or lower the detection limit. When quantifying by the concentration effect of the coffee ring, the randomness and variability of coffee ring formation make it impossible to fix the position from which the color is extracted. Moreover, because the position from which the color is extracted differs, there is a large difference in chromaticity, and as a result, the accuracy, reproducibility, and handling of detection cannot be ensured. [Summary of the Invention] [Problems the invention aims to solve]

[0007] The object of the present invention is to provide a paper-based microfluidic chip, a microfluidic detection system, and a liquid detection method in order to solve the problems that exist in conventional colorimetric quantification paper chips, where the detection limit and detection sensitivity cannot be met due to insufficient chromaticity and non-uniform chromaticity dispersion, resulting in low detection accuracy and reproducibility. The paper-based microfluidic chip and liquid detection method have high detection accuracy and sensitivity, can effectively lower the detection limit for liquid detection, and have good handling and reproducibility. [Means for solving the problem]

[0008] To achieve the above objective, one aspect of the present invention provides a paper-based microfluidic chip comprising a paper substrate layer, wherein a detection cell is provided on the paper substrate layer, the detection cell having a first contact angle region and a second contact angle region inside, and configured such that the contact angle of the liquid in the detection cell in the first contact angle region is greater than its contact angle in the second contact angle region.

[0009] A second aspect of the present invention provides a microfluidic detection system including the above-described paper-based microfluidic chip, the microfluidic detection system further includes a control unit for adjusting and controlling one or more of the ambient temperature, airflow velocity, humidity, and vacuum level in the region where the detection cell is located.

[0010] A third aspect of the present invention provides a liquid detection method comprising the steps of: introducing a liquid to be measured into a detection cell of a paper-based microfluidic chip; allowing the paper-based microfluidic chip to stand for a predetermined time; and performing chromaticity identification and / or colorimetric analysis on a predetermined area within the detection cell in the third aspect.

[0011] A fourth aspect of the present invention provides the use of the above-mentioned paper-based microfluidic chip, microfluidic detection system, or liquid detection method in water quality detection, environmental detection, and food and medical applications. [Effects of the invention]

[0012] According to the above technical solution, the present invention provides a first contact angle region and a second contact angle region with different contact angles within the detection cell, based on a chromaticity movement mechanism on a paper chip. By adjusting and controlling the surface tension of the liquid, a driving force is generated toward the inside of the droplet, thereby causing it to contract from the first contact angle region to the second contact angle region with a smaller contact angle, and the chromaticity is moved to the second contact angle region and concentrated. As a result, assuming that the concentration of the object to be measured is the same, the chromaticity and uniformity per unit area are improved, the detection sensitivity is improved, the detection limit is lowered, and the point sampling range is widened, improving the handling of point sampling and the reproducibility of detection. [Brief explanation of the drawing]

[0013] Figure 1 is a schematic diagram of a paper-based microfluidic chip according to a preferred embodiment of the present invention. Figure 2 shows the detection effect of a paper-based microfluidic chip according to a preferred embodiment of the present invention. Figure 3 is a schematic diagram of the gradient distribution of each contact angle region of the detection cell of a paper-based microfluidic chip according to a preferred embodiment of the present invention. Figure 4 is a schematic diagram of a disassembled paper-based microfluidic chip according to another preferred embodiment of the present invention. Figure 5 is a scatter plot showing the relationship between the chromaticity enhancement effect and the pore size of the ventilation holes. Figure 6 is a schematic diagram of a paper-based microfluidic chip according to another preferred embodiment of the present invention. Figure 7 shows the detection effect of the paper-based microfluidic chip in Figure 6. Figure 8 compares the chromaticity enhancement effects of various paper-based microfluidic chips. Figure 9 is a schematic diagram of a paper-based microfluidic chip according to another preferred embodiment of the present invention. Figure 10 is a distribution diagram of detection cells in a paper-based microfluidic chip according to another preferred embodiment of the present invention. Figure 11 is a scatter plot showing the relationship between the chromaticity enhancement effect and detection time at various ambient temperatures. Figure 12 is a schematic diagram of the disassembled paper-based microfluidic chip according to Embodiment 1 of the present invention. Figure 13 is a schematic diagram of the chromaticity enhancement effect of the paper-based microfluidic chip in Figure 12. Figure 14 is a schematic diagram of the structure of a paper-based microfluidic chip according to Embodiment 2 of the present invention. Figure 15 is a schematic diagram of the chromaticity enhancement effect of the paper-based microfluidic chip in Figure 14. Figures 16 and 17 show quantitative curves of the paper-based microfluidic chip in Example 3 of the present invention, respectively. [Modes for carrying out the invention]

[0014] Hereinafter, embodiments for carrying out the invention of the present invention will be described in detail with reference to the drawings. It should be understood that the embodiments for carrying out the invention described herein are for illustrative and interpretive purposes only and do not limit the invention.

[0015] In the present invention, the endpoints of the disclosed ranges and any values ​​should be understood to include values ​​close to these ranges or values, rather than being limited to precise ranges or values. In the case of numerical ranges, one or more new numerical ranges can be obtained by combining the endpoint values ​​of each range, the endpoint values ​​of each range and individual dot values, and the individual dot values, and these numerical ranges are deemed to be specifically disclosed herein.

[0016] As shown in Figure 1, a first aspect of the present invention provides a paper-based microfluidic chip comprising a paper substrate layer 2, which may be made of paper-like material such as filter paper or cellulose filtration membrane, and which functions, for example, as a microfluidic analysis technology platform for liquid detection, and has advantages such as lower cost, no need for external drive units, and superior biocompatibility and portability compared to conventional microfluidic chips. The paper-based microfluidic chip may have one or more paper substrate layers 2, and the thickness of the paper substrate layer 2 is not particularly limited. In each embodiment shown in the drawings of the present invention, only the case where there is one paper substrate layer 2 is shown in order to clearly illustrate the relevant structure (see Figures 4 and 9).

[0017] A detection cell 21 is formed in the paper substrate layer 2 of the paper-based microfluidic chip, and a liquid to be detected (e.g., wastewater, food or pharmaceutical solution) may be introduced into the detection cell 21 in order to detect / analyze information such as the components and concentrations in the liquid by means of chromaticity identification, colorimetric analysis, etc., as described below. In the present invention, as shown in Figure 1, the detection cell 21 has a first contact angle region 211 and a second contact angle region 212 inside, and the contact angle of the liquid to be detected introduced into the detection cell 21 is greater in the first contact angle region 211 than in the second contact angle region 212.

[0018] Therefore, when the liquid to be detected enters the detection cell 21, the surface tension of the liquid to be detected is adjusted and controlled by each contact angle region within the detection cell 21, generating a driving force toward the inside of the droplet. This causes the liquid to contract from the first contact angle region 211 to the second contact angle region 212, which has a smaller contact angle, and the chromaticity is shifted to the second contact angle region and concentrated. By providing regions with different contact angles, the paper-based microfluidic chip of the present invention can improve chromaticity and uniformity per unit area, improve detection sensitivity, lower the detection limit, and widen the point sampling range, thereby improving the handling of point sampling and detection reproducibility, assuming that the concentration of the substance to be measured is the same.

[0019] It is important to understand that the contact angles in each region within the detection cell 21 of the liquid to be detected mainly refer to the contact angles of these regions with respect to water or hydrophilic liquids. Therefore, although the paper-based microfluidic chip of the present invention itself does not contain the liquid to be detected, the same liquid to be detected will have different wettability with respect to different contact angle regions of the detection cell 21, resulting in different contact angles. For example, the water contact angle of the first contact angle region 211 may be set to be greater than 90°, thereby making it difficult for water or hydrophilic liquids to wet that region. On the other hand, the water contact angle of the second contact angle region 212 may be set to less than 30°. As a result, when the liquid to be detected enters the detection cell 21, the contact angle of the first contact angle region 211 is large, so a driving force is generated toward the second contact angle region 212 during the evaporation process of the liquid to be detected in the detection cell 21. The droplets contract toward the second contact angle region 212, and as the evaporation of water in the liquid to be detected progresses, the chromaticity moves toward the second contact angle region 212 due to this contraction process. Therefore, the chromaticity accumulates in the second contact angle region 212 within the detection cell 21, forming a spot with high chromaticity, which facilitates chromaticity identification and colorimetric analysis by a portable detection device (a smartphone with the software installed). Figure 2 shows the detection effect diagram of the paper-based microfluidic chip of the present invention, where the first contact angle region 211 surrounds the second contact angle region 212 and has a water contact angle of 91°, and the second contact angle region 212 is a circle with a diameter of 2 mm. Therefore, by providing regions with different contact angles, the chromaticity enhancement effect within the detection cell can be improved, detection sensitivity can be increased, the detection limit can be lowered, and the point sampling range can be expanded, improving the handling of point sampling and detection reproducibility, thus facilitating chromaticity identification and colorimetric analysis.

[0020] In the present invention, the first contact angle region 211 and the second contact angle region 212 may be formed within the detection cell 21 in various ways. For example, the first contact angle region 211 may be formed by laying, depositing, or impregnating a hydrophobic material in the portion of the detection cell 21 near the outer edge, and / or the second contact angle region 212 may be formed by laying, depositing, or impregnating a hydrophilic material in the central portion of the detection cell 21. Specifically, a first contact angle region 211 with a large contact angle can be obtained by laying a polytetrafluoroethylene film in the portion of the detection cell 21 near the outer edge, or by depositing or impregnating a hydrophobic material such as a silanating reagent or a fluorine-containing material on the surface of that portion, thereby separating the solution to be detected from the substrate of the paper substrate layer or hydrophobically modifying the substrate of the paper substrate layer. In the central part of the detection cell 21, a hydrophilic second contact angle region 212 is formed by the paper substrate layer itself, such as filter paper or a cellulose filtration membrane, without any treatment. Alternatively, the surface of this portion is subjected to plasma treatment, and a bovine serum albumin (BSA) solution or the like is applied to reduce the contact angle through surface modification, thereby forming a second contact angle region 212 with a small contact angle. In a preferred embodiment, the contact angle of the entire detection cell 21 is increased by laying a polytetrafluoroethylene film inside the detection cell 21, or by depositing or impregnating a hydrophobic material such as a silanating reagent or a fluorine-containing material, thereby forming a hydrophobic layer with a relatively large contact angle. Subsequently, in the central part of the detection cell 21, a hydrophilic material (for example, laying filter paper or applying BSA) is laid, deposited, or impregnated above the hydrophobic layer to form a second contact angle region 212 with a relatively small contact angle in the central part of the detection cell 21. In this case, the other parts of the detection cell 21 become the first contact angle region 211.

[0021] In the detection cell 21 of the paper-based microfluidic chip according to the present invention, the first contact angle region 211 may be configured such that the water contact angle is larger than 60°, preferably larger than 90°, more preferably larger than 120°, and the second contact angle region 212 may be configured such that the water contact angle is less than 30°, and further close to 0°. Thereby, as evaporation proceeds, the solution in the detection cell 21 gathers in the second contact angle region 212, so that chromaticity is concentrated in the second contact angle region 212 and a spot is formed. In this way, a low-concentration liquid to be detected can be effectively detected, the detection sensitivity is improved, and the detection limit can be lowered.

[0022] As described above, in the present invention, by providing regions with different contact angles in the detection cell 21, chromaticity is concentrated in a specific region, whereby the detection point sampling position can be determined as needed, and this sampling position depends on the position of the second contact angle region 212 within the detection cell 21. In the illustrated preferred embodiment, the detection cell 21 is formed in a circular shape with a diameter of 2 mm to 8 mm, and the second contact angle region 212 is located at the center position of the detection cell 21, whereby the liquid to be detected that has entered the detection cell 21 can uniformly gather at the center from all directions, which contributes to improving detection accuracy and precision. In an alternative embodiment, the second contact angle region 212 may be located at other positions of the detection cell 21, for example, other central portions offset from the center position, while the first contact angle region 211 surrounds the second contact angle region 212. Alternatively, the detection cell 21 may be formed as a regular polygon with a circumcircle diameter of 2 mm to 8 mm, and the second contact angle region 212 may be arranged at the center position of the regular polygon.

[0023] Based on the movement mechanism of chromaticity within the detection cell 21, the size of the spot formed by chromaticity enrichment and the degree of enrichment greatly depend on the size of the second contact angle region 212. That is, when the second contact angle region 212 is small, the degree of chromaticity enrichment is high and the formed spot is small, which is more conducive to the detection of low-concentration solutions. For this reason, the size of the detection cell 21 may be determined according to the size of the second contact angle region 212 such that the proportion of the area of the second contact angle region 212 in the detection cell 21 is 50% or less, preferably 30% or less. In the case of a general detection cell (a circular shape with a diameter of 2 mm to 8 mm, or a regular polygon with a circumcircle diameter of 2 mm to 8 mm), the second contact angle region 212 may be configured as a circular region with a diameter of 0.5 mm to 5 mm (preferably 1 mm to 3 mm), or a regular polygonal region with a circumcircle diameter of 0.5 mm to 5 mm (preferably 1 mm to 3 mm).

[0024] As shown in Fig. 1 and Fig. 3, in order to enhance the effects of chromaticity enrichment and uniform distribution, in the paper-based microfluidic chip according to the present invention, multi-stage contact angle regions may be further provided in the detection cell 21, whereby the contact angle increases stepwise from the center to the periphery of the detection cell 21, forming a plurality of contact angle regions distributed in a gradient. Specifically, the first contact angle region 211 may include a first gradient region 2111 close to the outer edge of the detection cell 21 and a second gradient region 2112 close to the second contact angle region 212, wherein the contact angle of the second gradient region 2112 is smaller than that of the first gradient region 2111. Therefore, during the evaporation process of the liquid, the driving force gradually decreases along the direction from the edge of the detection cell 21 toward the second contact angle region 212, whereby the acceleration of the liquid moving toward the second contact angle region 212 gradually decreases, which contributes to the uniform distribution of chromaticity in the second contact angle region 212. Fig. 3 is a schematic diagram showing the gradient distribution of each contact angle region in the detection cell. It can be understood that the first contact angle region 211 including the illustrated two-stage gradient region is merely an example, and in the paper-based microfluidic chip according to the present invention, the detection cell 21 may be provided with a plurality of gradient regions.

[0025] In the paper-based microfluidic chip according to the present invention, the detection cell 21 may be configured to be open to the outside, and volatile components in the liquid to be detected may be directly evaporated through the upper opening of the detection cell 21 until the chromaticity is concentrated in the second contact angle region 212. Based on this, the present invention can further improve the chromaticity concentration effect within the detection cell by controlling the direction and speed of chromaticity movement by volatilizing the liquid to be detected only in a specific region, and this will be described in detail below.

[0026] As shown in Figure 4, a paper-based microfluidic chip according to another preferred embodiment of the present invention includes the above-mentioned paper substrate layer 2 on which a detection cell (not shown) is provided, and an upper layer 3 and a lower layer 1 provided above and below the paper substrate layer 2, respectively. Of these, the lower layer 1 may cover the lower side (first side) of the paper substrate layer 2 through adhesive or the like, and the upper layer 3 may cover the upper side (second side) of the paper substrate layer 2 through adhesive or the like. The upper layer 3 and the lower layer 1 may cover all or part of the detection cell, forming an impermeable and impermeable portion. For example, the upper layer 3 and the lower layer 1 as a whole may be manufactured from an impermeable and impermeable material, and the portion covering the detection cell may be manufactured from an impermeable and impermeable material. Specifically, the non-permeable and non-air permeable material may be polyvinyl chloride, polyethylene, polypropylene, polystyrene, silicone, polytetrafluoroethylene, etc. The upper layer 3 and lower layer 1, which are made of a permeable material, may be subjected to hydrophobic treatment, such as by applying wax to the filter paper and heating it to impregnate the wax into the filter paper, or by impregnating the filter paper in a plastic solution dissolved in an organic solvent and drying it. The lower layer 1 and upper layer 3 may or may not be in contact with each other, and their areas and shapes do not have to be the same. The area may be smaller than the area of ​​the paper substrate layer 2, but it is necessary to cover the entire detection cell. The lower layer 1 and upper layer 3 may be made of transparent or opaque material, but transparent material is preferred.

[0027] Ventilation holes 31 are formed in the portion of the upper layer 3 that covers the detection cell, thereby allowing the liquid 4 inside the detection cell to evaporate in a specific area through the ventilation holes 31. This guides the liquid 4 inside the detection cell so that it accumulates at the location of the ventilation holes 31 during the evaporation process, concentrating the chromaticity at that location. Specifically, the upper layer 3 covers the edge portion of the detection cell, preventing evaporation of the liquid 4 from the edge of the detection cell. The ventilation holes 31 of the upper layer 3 connect the detection cell to the outside space, so that the liquid inside the detection cell can only evaporate to the outside through the ventilation holes 31. Therefore, during the evaporation process, the evaporation rate of water is faster near the ventilation holes 31 inside the detection cell, and water at the edge is replenished near the ventilation holes 31. In this way, the chromaticity moves to that location, and a spot with concentrated chromaticity is formed.

[0028] In this case, the position of the ventilation holes 31 relative to the detection cell may be set such that the position of the ventilation holes 31 corresponds to the position of the second contact angle region 212 within the detection cell (on the same vertical line), thereby further enhancing the chromaticity enrichment effect within the detection cell, lowering the detection limit, and improving detection sensitivity.

[0029] Therefore, the vent hole 31 may have the same shape and dimensions as the second contact angle region 212, and both its area and vent diameter may be smaller than those of the second contact angle region 212. For example, the vent hole 31 may be a regular polygon or a circle with a diameter or circumscribed circle diameter of 0.5 mm to 5 mm, preferably 1 mm to 3 mm. The vent hole 31 may be located above the center of the detection cell to uniformly deepen the chromaticity during the volatilization process of the liquid in the detection cell.

[0030] Furthermore, the size of the vent hole 31 significantly affects the chromaticity enrichment effect. As shown in Figure 5, within a predetermined range (before chromaticity saturates), the smaller the vent hole diameter, the higher the chromaticity of the ultimately formed spot. For example, if the vent hole diameter is 3 mm, the enriched chromaticity (distance) is only about 120, while if the vent hole diameter is 1.5 mm, the enriched chromaticity (distance) reaches 200 or more. Therefore, in order to achieve an even higher chromaticity enrichment effect, the diameter of the vent hole 31 or the circumscribed circle diameter may be set to 1 mm to 3 mm, but this may result in a longer enrichment time. For this reason, the evaporation of the liquid may be accelerated by adjusting and controlling the ambient temperature, humidity, and vacuum level in the area where the detection cell is located, which will be described in detail below.

[0031] Figure 6 shows an improved embodiment of the paper-based microfluidic chip of the present invention, which provides a chip structure with beneficial effects. Specifically, in the paper-based microfluidic chip, the paper substrate layer 2 is provided with a liquid storage cell 24 arranged around the detection cell 21, so that if the evaporation rate of the liquid in the detection cell 21 is too fast and all the chromatic particles cannot move to the vicinity of the vent holes 31, a colorless liquid can be added to the liquid storage cell 24 to replenish the liquid that should evaporate in the detection cell 21, allowing the chromatic particles to continue moving to the vicinity of the vent holes 31. In the liquid supply channel, one or more openable and closable liquid supply holes may be provided in the portion of the upper layer 3 that covers the liquid storage cell 204, and the liquid supply holes may be opened when liquid replenishment is needed and closed after liquid replenishment is complete. In actual detection, the colorless liquid added to the liquid storage cell 24 may be water or a mixture of different solvents, but the added liquid is required to be able to dissolve / carry the chromatic particles and not interact with other parts of the chip. The detection effect of a paper-based microfluidic chip with an added liquid storage cell 24 is shown in Figure 7. Figure 8 shows a comparison of the chromaticity enrichment effect of paper-based microfluidic chips with and without a liquid storage cell 24. As can be seen from this figure, by adding replenishment liquid to the liquid storage cell 24, the degree of chromaticity enrichment near the vent holes can be significantly improved. Under the same conditions, the chip without a liquid storage cell (chip 2) enriches its chromaticity from 48.2 to 190.3, while the chip with a liquid storage cell (chip 1) enriches its chromaticity from 49.4 to 257.4.

[0032] In some embodiments of the present invention, the liquid to be detected may be added directly into the detection cell 21 from above, for example, the ventilation holes 31 may function as sample addition holes. In some other embodiments, the paper substrate layer 2 may have a sample addition area 22 at a location other than the detection cell 21, and the sample addition area 22 and the detection cell 21 may be connected via a diffusion channel 23. In this case, the upper layer 3 may have a sample addition hole 32 at a location corresponding to the sample addition area 22, as shown in Figure 9. Thereafter, the liquid to be detected may be injected into the sample addition area 22 via the sample addition hole 32, enter the detection cell 21 via the diffusion channel 23 by a self-driving action, and then proceed to the subsequent chromaticity enrichment and detection process. In some embodiments of the present invention, as shown in Figure 10, there may be multiple detection cells 21, the number of which may be determined according to the number of samples to be detected and / or the number of parameters to be detected. Preferably, each detection cell 21 is at the same distance from the sample addition area 22, that is, it is preferable that the lengths of each diffusion channel 23 are equal. In the detection cell 21, seven sample addition areas 22 are provided surrounding the center. Therefore, when it is necessary to perform parallel detection or repeated testing of multiple samples, detection can be completed in one go using the tip, thereby improving the detection rate and further reducing errors between parallel tests.

[0033] To facilitate detection, a color-developing reagent may be pre-placed in the detection cell 21 or the diffusion channel 23. This color-developing reagent can cause a clear color to develop in the liquid within the detection cell 21 through a chemical reaction or the like, thereby facilitating chromaticity identification and colorimetric analysis.

[0034] A second aspect of the present invention provides a microfluidic detection system including the above-described paper-based microfluidic chip, the microfluidic detection system including related equipment used in combination with the above-described paper-based microfluidic chip, such as a chip holder and a camera.

[0035] In particular, in paper-based microfluidic chips with ventilation holes, the microfluidic detection system may include an adjustment and control unit for adjusting and controlling one or more of the ambient temperature, airflow velocity, humidity, and vacuum level in the area where the detection cell 21 is located. By adjusting these environmental elements, the evaporation rate of the liquid in the detection cell 21 is controlled, and the chromatic enrichment effect is improved. For example, a heating plate may be provided to heat the ambient temperature in the area where the detection cell 21 is located, and the heating plate maintains the ambient temperature in the area where the detection cell 21 is located within a predetermined temperature range between 35°C and 45°C. Figure 11 shows the relationship between chromatic distance and detection time at ambient temperatures of 25°C, 35°C, and 45°C, where the final degree of enrichment is almost identical when the chromatic enrichment rate at ambient temperatures of 35°C and 45°C is much higher than at 25°C. Therefore, the detection efficiency is effectively increased by providing an appropriate adjustment and control unit.

[0036] The adjustment and control unit may be configured to adjust and control other environmental factors besides ambient temperature, for example, it may include a venting device to increase the airflow velocity around the vent 31 or reduce the humidity near the vent 31 by releasing pressurized gas above the detection cell 21 or replacing the air above the detection cell 21. Alternatively, the adjustment and control unit may include a vacuum oven, in which the paper-based microfluidic chip is placed in this vacuum oven and left for a predetermined time to accelerate chromatic intensification during the detection process.

[0037] A third aspect of the present invention provides a liquid detection method using a paper-based microfluidic chip, comprising the steps of: S1 introducing a liquid to be measured into a detection cell 21 of a paper-based microfluidic chip; S2 allowing the paper-based microfluidic chip to stand for a predetermined time; and S3 performing chromaticity identification and / or colorimetric analysis on a predetermined region within the detection cell 21. The predetermined region is a chromaticity-enhancing region within the detection cell 21.

[0038] As described above, in order to increase the rate of liquid evaporation and improve the chromaticity enhancement effect, in step S2 above, one or more of the ambient temperature, humidity, and vacuum level of the region where the detection cell 21 is located can be adjusted and controlled. For example, the ambient temperature of the region where the detection cell 21 is located can be adjusted and controlled to a predetermined temperature range maintained between 25°C and 60°C.

[0039] A fourth aspect of the present invention provides the use of the above-described paper-based microfluidic chip, microfluidic detection system, or liquid detection method in water quality detection, environmental detection, and food and medical applications. For example, the above-described paper-based microfluidic chip, microfluidic detection system, and liquid detection method can be used for detecting the content of nickel, chromium, phosphates, etc., in water, or for measuring various indicators in biomedicine, or for standard measurement of various substances in food.

[0040] The paper-based microfluidic chip, microfluidic detection system, and liquid detection method according to the present invention can improve quantitative accuracy and lower the detection limit. Compared to conventional technology, the present invention: 1) Increases the intensity of the chromaticity of the detection cell in colorimetric quantification, further increasing the intensity to the point where the color is not visible, thereby lowering the detection limit of the analyte. 2) By adjusting and controlling the chromaticity, the uniformity of the distribution is improved while improving the chromaticity on the paper chip, the point sampling position is increased, handling is improved, and detection accuracy and reproducibility are further improved. 3) It can be realized through a simple method of combining material surface modification through 3D structural design, so there is no need to add additional complex equipment, thereby ensuring the portability of paper chip detection. 4) It is applicable to most paper chips in colorimetric quantification, and has high practicality and versatility.

[0041] The present invention will be described in detail below with reference to examples, but the contact angle is measured by the OCA200 fully automatic single-fiber contact angle measuring device. [Example 1]

[0042] Figure 12 shows a paper-based microfluidic chip according to the embodiment, which comprises a lower layer 1, a paper substrate layer 2, and a concentration layer 25. The paper substrate layer 2 comprises six detection cells 21 arranged in an annular array and a blank control detection cell in the center, and the concentration layer 25 is a concentration carrier added to each detection cell 21, the concentration carrier being in the central part (including but not limited to the center) of the detection cell 21. The detection cells 21 are made of a material with a large contact angle (e.g., filter paper modified with a fluorinated silane reagent or a plastic sheet with its own large contact angle), and the concentration carrier is made of a material with a small contact angle (e.g., filter paper, cellulose filtration membrane, or other material that has undergone plasma treatment or surface modification), the concentration carrier is smaller in size than the detection cell 21 and forms a second contact angle region with a small contact angle, and is generally preferably circular or a regular polygon with a circumscribed circle diameter of 1 mm to 3 mm. The rest of the detection cell 21 becomes a first contact angle region with a large contact angle. The enriched carrier, detection cell, and lower layer are adhered to each other by adhesion or other means.

[0043] The colored solutions after the reaction (e.g., a 0.1% dye solution, or a solution of the target substance at each concentration mixed with a specific reagent) were dropped onto detection cells arranged in a circular array, with the central detection cell serving as a blank control. The chromaticity enhancement effect is shown in Figure 13. After enhancing the chromaticity under specific conditions (e.g., 35°C, 10 min), the detection cells were imaged under natural light conditions, and the RGB values ​​of the photographs were analyzed to calculate the chromaticity distance D. The chromaticity of the 0.1% red dye solution after enhancement is shown in the table below.

[0044] [Table 1] [Example 2]

[0045] In this embodiment, a paper chip with a three-layer structure is used as a base chip with enhanced chromaticity, and an aqueous solution to which a dye has been added is used as a sample. This demonstrates that the present invention can enhance the chromaticity of all color-based solutions, and that chromaticity enhancement occurs in any small area that is in communication with air and does not need to be coaxial with the sample addition hole.

[0046] As shown in Figure 14, the paper chip structure consists of a paper substrate layer, fabricated by a cutting method, covered with a transparent, non-permeable film on both the upper and lower sides. The upper film (upper layer) has ventilation holes at positions corresponding to the detection cells. The sample injection holes are located at the same distance from each detection cell. A dye solution is injected through the sample injection holes, and this dye solution is distributed along hydrophilic diffusion channels, reaching each detection cell 1-7. When the paper chips are left in a natural environment (ambient temperature 26°C, humidity 70%), after a predetermined time (10-60 min), the chromaticity is concentrated in the ventilation hole regions of the detection cells, and as time progresses, the contrast between the chromaticity of the concentrated regions and the chromaticity of other regions becomes more pronounced. The RGB values ​​were read for the chromaticity of multiple regions within different detection cells and within the same detection cell, and the results are shown in the following table and Figure 15.

[0047] [Table 2]

[0048] [Table 3]

[0049] As can be seen from each of the tables above, the R, G, and B values ​​are similar between different detection cells and between different regions of the same detection cell. Furthermore, the standard deviations of the R, G, and B values ​​are all less than 3% between different detection cells and between different regions of the same detection cell. This suggests that the chromaticity proposed in this invention is enhanced, and that the uniformity and reproducibility of the liquid detection method are excellent. [Example 3]

[0050] This embodiment illustrates the reduction in the detection limit of nickel in water due to a chromatic enhancement structure.

[0051] In the paper-based microfluidic chip used in this embodiment, the lower layer is a transparent, non-permeable membrane, the paper substrate layer is hydrophobically modified filter paper (the sample addition area, diffusion channel, and detection cell retain hydrophilicity, while the remaining area is hydrophobically modified, and the detection cell is pre-filled with a composite reagent mainly composed of dimethylglyoxime that can undergo a specific color reaction with nickel), and the upper layer is a transparent, non-permeable membrane provided with a sample addition hole communicating with the sample addition area and a ventilation hole communicating with the detection cell. The diameter of the detection cell is 4 mm, and the diameter of the ventilation hole is 2 mm.

[0052] During detection, a nickel-containing water sample is injected through the sample injection well. The sample flows along the diffusion channel into the detection cell, where it reacts with the composite reagent to produce a pink substance. After the chip is left for a predetermined time, the water vapor evaporates along the upper vents, and the color-developing components diffuse and accumulate in the chromaticity-enhancing region due to the action of evaporation, causing the color to gradually deepen. After the reaction is complete, the chromaticity-enhancing region of the paper chip is imaged, and chromaticity information from this region is extracted using MATLAB for quantitative calculations.

[0053] As can be seen from Figures 16 and 17, ordinary paper chips have a high detection limit for nickel in water, and as the chromaticity is increased, the detection limit for nickel in water decreases to 0.1 mg / L, indicating a significant effect of reducing the detection limit. When standard curves were fitted to nickel solutions within the linear range, the linearity of the resulting fitted lines improved very significantly. [Example 4]

[0054] This embodiment illustrates the reduction in the detection limit of chromium in water due to a chromatic enhancement structure.

[0055] In the paper-based microfluidic chip used in this embodiment, the lower layer is a transparent, non-permeable membrane, the intermediate layer is hydrophobically modified filter paper (the sample addition area, diffusion channel, and detection cell retain hydrophilicity, while the remaining area is hydrophobically modified, and the detection cell is pre-filled with a composite reagent mainly composed of diphenylcarbazide that can undergo a specific color reaction with chromium), and the upper layer is a transparent, non-permeable membrane provided with a sample addition hole communicating with the sample addition area and a ventilation hole communicating with the detection cell. The diameter of the detection cell is 4 mm, and the diameter of the ventilation hole is 2.5 mm.

[0056] During detection, when a chromium-containing water sample is injected from the sample addition area, the sample flows along the diffusion channel into the detection cell, reacts with the composite reagent to produce a pink substance. After leaving the tip for a predetermined time, the water vapor evaporates along the upper vents, and the color-developing components diffuse and accumulate in the chromaticity-enhancing area due to the action of evaporation, causing the color to gradually deepen.

[0057] As a result, before color intensification, the detection limit of chromium in a normal paper chip is 0.05 mg / L, while in the paper chip according to this embodiment, the color development is enhanced and the detection limit drops to 0.03 mg / L. [Example 5]

[0058] This example illustrates the reduction in the detection limit of nitrite in water due to a chromatic enhancement structure.

[0059] In the paper-based microfluidic chip used in this embodiment, the lower layer is a transparent, non-permeable membrane, the paper substrate layer is hydrophobically modified filter paper (the sample addition area, diffusion channel, and detection cell retain hydrophilicity, while the remaining area is hydrophobically modified, and the detection cell is pre-filled with Griess reagent which can specifically react with nitrite), and the upper layer is a transparent, non-permeable membrane with a sample addition hole communicating with the sample addition area and a ventilation hole communicating with the detection cell. The diameter of the detection cell is 5 mm, and the diameter of the ventilation hole is 2 mm.

[0060] During detection, when a chromium-containing water sample is injected from the sample addition area, the sample flows along the diffusion channel into the detection cell, and the combined reagent reacts to produce a pink substance. After leaving the tip for a predetermined time, the water vapor evaporates along the upper vents, and the color-developing components diffuse and accumulate in the chromaticity-enhancing area due to the action of evaporation, causing the color to gradually deepen. As a result, before the color intensification, the detection limit for chromium in a normal paper chip is 0.1 mg / L, while in the paper chip according to this embodiment, the color development is enhanced and the detection limit drops to 0.05 mg / L.

[0061] Preferred embodiments of the present invention have been described in detail above with reference to the drawings, but the present invention is not limited thereto. Within the scope of the technical concept of the present invention, several simple modifications are possible, including combining individual specific technical features in any appropriate manner. To avoid unnecessary redundancy, various combinations of the present invention will not be described separately. However, these simple modifications and combinations should also be considered within the disclosure of the present invention and all fall within the scope of protection of the present invention. [Brief explanation of the drawing]

[0062] [Figure 1] This is a schematic diagram of a paper-based microfluidic chip according to a preferred embodiment of the present invention. [Figure 2] This is a diagram illustrating the detection effect of a paper-based microfluidic chip according to a preferred embodiment of the present invention. [Figure 3] This is a schematic diagram of the gradient distribution in each contact angle region of a detection cell of a paper-based microfluidic chip according to a preferred embodiment of the present invention. [Figure 4] This is a schematic diagram of a disassembled paper-based microfluidic chip according to another preferred embodiment of the present invention. [Figure 5] This is a scatter plot showing the relationship between the chromaticity enhancement effect and the pore size of the ventilation holes. [Figure 6] This is a schematic diagram of a paper-based microfluidic chip according to another preferred embodiment of the present invention. [Figure 7]Figure 6 shows the detection effect of a paper-based microfluidic chip. [Figure 8] This figure compares the chromatic enhancement effects of various paper-based microfluidic chips. [Figure 9] This is a schematic diagram of a paper-based microfluidic chip according to another preferred embodiment of the present invention. [Figure 10] This is a distribution diagram of detection cells in a paper-based microfluidic chip according to another preferred embodiment of the present invention. [Figure 11] This is a scatter plot showing the relationship between the chromaticity enhancement effect and detection time at various ambient temperatures. [Figure 12] This is a schematic diagram of the disassembled paper-based microfluidic chip according to Embodiment 1 of the present invention. [Figure 13] Figure 12 is a schematic diagram of the chromaticity enhancement effect of paper-based microfluidic chips. [Figure 14] This is a schematic diagram of the structure of a paper-based microfluidic chip according to Embodiment 2 of the present invention. [Figure 15] Figure 14 is a schematic diagram of the chromaticity enhancement effect of paper-based microfluidic chips. [Figure 16] This is a quantitative curve of a paper-based microfluidic chip in Example 3 of the present invention. [Figure 17] This is a quantitative curve of a paper-based microfluidic chip in Example 3 of the present invention.

Claims

1. Paper-based microfluidic chip, The paper-based microfluidic chip includes a paper substrate layer (2), the paper substrate layer (2) is provided with a detection cell (21), the detection cell (21) has a first contact angle region (211) and a second contact angle region (212) inside, and is configured such that the contact angle of the liquid inside the detection cell (21) in the first contact angle region (211) is greater than its contact angle in the second contact angle region (212). The paper-based microfluidic chip includes a lower layer (1) provided on the first side of the paper substrate layer (2) and an upper layer (3) provided on the second side of the paper substrate layer (2) opposite to the first side, wherein the lower layer (1) and the upper layer (3) have a water-impermeable and air-permeable portion that covers the detection cell (21), and ventilation holes (31) are formed in the portion of the upper layer (3) that covers the detection cell (21). The paper substrate layer (2) is further provided with a liquid storage cell (24) arranged around the detection cell (21) so that liquid can be supplied to the detection cell (21), and one or more openable and closable liquid supply holes are provided in the portion of the upper layer (3) that covers the liquid storage cell (24). Paper-based microfluidic chips.

2. The paper-based microfluidic chip according to claim 1, wherein the detection cell (21) is provided with a first contact angle region (211) which is laid, deposited, or impregnated with a hydrophobic material, and / or the detection cell (21) is provided with a second contact angle region (212) which is laid, deposited, or impregnated with a hydrophilic material.

3. The paper-based microfluidic chip according to claim 1, wherein a hydrophobic layer is formed within the detection cell (21) by laying, depositing, or impregnating a hydrophobic material within the detection cell (21), and in the central portion of the detection cell (21), a second contact angle region (212) and the first contact angle region (211) surrounding the second contact angle region (212) are formed by laying, depositing, or impregnating a hydrophilic material above the hydrophobic layer.

4. The paper-based microfluidic chip according to claim 1, wherein the second contact angle region (212) is located in the central part of the detection cell (21), and the first contact angle region (211) surrounds the second contact angle region (212).

5. The paper-based microfluidic chip according to claim 4, wherein the detection cell (21) is formed in the shape of a circle with a diameter of 2 mm to 8 mm or a regular polygon with a circumscribed diameter of 2 mm to 8 mm, and the second contact angle region (212) is located at the center of the detection cell (21) and / or the second contact angle region (212) is a circular region with a diameter of 0.5 mm to 5 mm or a regular polygonal region with a circumscribed diameter of 0.5 mm to 5 mm.

6. The paper-based microfluidic chip according to claim 4, wherein the first contact angle region (211) is configured such that the contact angle of the liquid in the detection cell (21) in the first contact angle region (211) increases as you move from the second contact angle region (212) toward the edge of the detection cell (21).

7. The paper-based microfluidic chip according to claim 1, wherein the water contact angle of the first contact angle region (211) is greater than 60°, preferably greater than 90°, and more preferably greater than 120°.

8. The paper-based microfluidic chip according to claim 1, wherein the ventilation holes (31) are regular polygons or circular, and both their area and hole diameter are smaller than those of the second contact angle region (212), and preferably the diameter of the ventilation holes (31) or the circumscribed circle diameter is 0.5 mm to 5 mm, preferably 1 mm to 3 mm.

9. The paper-based microfluidic chip according to claim 1, wherein the position of the ventilation hole (31) in the upper layer (3) corresponds to the position of the second contact angle region (212) in the detection cell (21).

10. The paper substrate layer (2) is further provided with a sample addition region (22) and a diffusion channel (23) connecting the sample addition region (22) and the detection cell (21), The paper-based microfluidic chip according to claim 1, wherein the upper layer (3) is provided with a sample addition hole (32) at a position corresponding to the sample addition area (22).

11. The paper-based microfluidic chip according to claim 10, wherein a color-developing reagent is pre-disposed in the detection cell (21) and / or the diffusion channel (23).

12. A microfluidic detection system comprising a paper-based microfluidic chip according to any one of claims 1 to 11.

13. The microfluidic detection system according to claim 12, further comprising an adjustment and control unit for adjusting and controlling one or more of the ambient temperature, airflow velocity, humidity, and vacuum level in the region where the detection cell (21) is located.

14. A liquid detection method, Step S1 involves introducing the liquid to be measured into the detection cell (21) of the paper-based microfluidic chip according to any one of claims 1 to 11, Step S2 involves leaving the paper-based microfluidic chip to stand for a predetermined time, A liquid detection method comprising step S3 of performing chromaticity identification and / or colorimetric analysis on a predetermined area within the detection cell (21).

15. The liquid detection method according to claim 14, wherein in step S2, the ambient temperature of the region where the detection cell (21) is located is set to between 25°C and 60°C.

16. The liquid detection method according to claim 14, wherein in step S2, one or more of the ambient temperature, humidity, and vacuum level of the region in which the detection cell (21) is located are adjusted and controlled using an adjustment and control unit.

17. Use of a paper-based microfluidic chip according to any one of claims 1 to 11 in water quality detection, environmental detection, and food and medical applications.

18. Use of the microfluidic detection system according to claim 12 in water quality detection, environmental detection, and food and medical applications.

19. Use of the liquid detection method according to claim 14 in water quality detection, environmental detection, and food and medical applications.

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