A portable sample pretreatment and test strip rapid measurement combined device

Abstract

The utility model discloses a portable sample pretreatment and test paper strip rapid measurement combined device, include: device main part is equipped with sample inlet storehouse, and the top is open and forms first sample inlet hole, purifying storehouse is equipped with below sample inlet storehouse, and is formed by surrounding at least two sieve plates of upper and lower distribution, purifying layer is equipped with in purifying storehouse, first reaction storehouse is linked with purifying storehouse intercommunication, and forms first detection hole, second reaction storehouse is set up with first reaction storehouse each other independence, links with second sample inlet hole and second detection hole intercommunication respectively. The utility model is with antigen-antibody specificity reaction as the basic principle's colloidal gold immune chromatography technique, and the visual of colloidal gold detection and the specificity of immune chromatography detection are integrated, and the utility model discloses a kind of on-site rapid detection for sample, effectively make up the defect of traditional detection method cost higher. It can be widely used in the precise identification and screening of pollutants in the field of food safety supervision departments at all levels.

Description

Technical Field

[0001] This utility model belongs to the field of on-site rapid testing technology for food safety, and in particular relates to a portable sample pretreatment and test strip rapid testing device. Background Technology

[0002] Pesticides, veterinary drugs, and mycotoxins are significant risk factors affecting food quality and safety. Among these, due to factors such as a lack of responsibility among producers and operators, insufficient scientific management of agricultural production, improper food storage conditions, and deteriorating aquaculture environments, excessive levels of pesticides and mycotoxins are commonly found in vegetables and fruits, leading to food safety issues. Furthermore, incidents involving drug residues, such as the "chloramphenicol incident in shrimp," "multiple drug contaminations in turbot," and "excessive drug residues in bullfrogs," frequently occur in aquatic products, posing a potential threat to consumer health. Therefore, it is imperative to strengthen and improve food safety testing capabilities, which is crucial for ensuring the quality and safety of agricultural products and the health of Chinese residents.

[0003] Existing rapid on-site testing technologies for food safety mainly consist of two parts: sample pretreatment and quantitative analysis of analytes. In recent years, globally recognized immunochromatographic test strips have been widely used for rapid on-site screening of risk factors such as pesticide and veterinary drug residues and mycotoxin residues in food due to their advantages of sensitivity, simplicity, and ease of operation, demonstrating significant application prospects and promotional value. The newly promulgated "Agricultural Product Quality and Safety Law of the People's Republic of China" in 2023 stipulates that rapid detection methods such as colloidal gold, recognized by relevant national departments, can be used for random inspections of agricultural product quality and safety, and the test results can serve as evidence for administrative penalties. However, the impact of food sample matrix on the reliability and accuracy of rapid test results has become a bottleneck issue. Firstly, sample pretreatment directly affects the detection limit and reliability of the analytical method. However, due to the wide variety of agricultural products, complex matrices, and the potential for matrix components such as plant pigments and organic acids to interfere with the recognition of immune response elements such as antigens and antibodies, or directly affect the signal intensity of signal markers such as colloidal gold, thus affecting the interpretation of test results. Therefore, there is an urgent need to adopt efficient pretreatment technologies to fully extract the target analytes and to reduce the interference of the sample matrix on the test results through purification and enrichment. However, currently used sample pretreatment methods (such as QuEChERS) are cumbersome, time-consuming, have low automation, poor portability, and high barriers to entry, making it difficult to achieve high-throughput on-site screening of large-scale samples, thus hindering the development of food safety technologies. Furthermore, the large consumption of organic solvents may cause secondary environmental pollution. Secondly, the principle of lateral flow immunochromatography is to use capillary action to allow the added liquid sample to migrate along the direction of the absorbent pad and specifically bind to the stationary phase (antigen or antibody). Then, using colloidal gold, quantum dots, or other nanoparticles as markers, combined with a matching reader or excitation device, the detection results can be visualized and the analyte can be quantitatively analyzed. However, there is still a lack of matching, simple, low-solvent-consumption sample pretreatment technologies that are not limited by large instruments. This restricts the application of colloidal gold and other immunochromatographic test strips in rapid on-site food safety testing.

[0004] Therefore, there is an urgent need to develop portable, easy-to-operate, integrated, and automated sample pretreatment devices and methods to further simplify the testing process, reduce time and labor costs, and improve the level of rapid on-site food safety testing. Utility Model Content

[0005] To overcome the shortcomings of existing technologies, this utility model provides a portable sample pretreatment and rapid test strip combined device, which can automatically complete the purification reaction in the process of rapid on-site food safety testing in a short time. By combining it with colloidal gold immunochromatographic test strips, it can perform rapid sample pretreatment and on-site rapid testing and screening of target analytes for various types of food.

[0006] The technical solution adopted by this utility model to solve its technical problem is: a portable sample pretreatment device, comprising:

[0007] The main body of the device is equipped with a sample inlet chamber, and the top is open to form the first sample inlet port;

[0008] The purification chamber is located below the sample injection chamber and is formed by at least two sieve plates arranged vertically.

[0009] A purification layer is provided inside the purification chamber;

[0010] The first reaction chamber is connected to the purification chamber and has a first detection hole.

[0011] The second reaction chamber is set independently of the first reaction chamber and is connected to the second sample inlet and the second detection port, respectively.

[0012] The first injection port, the first detection port, the second injection port, and the second detection port are located on the side of the main body of the device.

[0013] This invention discloses a portable sample pretreatment device. The sample or extract to be tested is directly injected into the injection chamber. After being filtered through a first sieve plate, the sample or extract enters a purification chamber containing a purification layer. After purification, the sample or extract is filtered again through a second sieve plate. The sample or extract then enters a first reaction chamber through a first passage for reaction. Simultaneously, a blank control is placed in a second injection port and enters a second reaction chamber through a second passage for reaction. The reaction takes approximately 2 minutes. Then, two test strips are placed into the first and second detection ports respectively. After reacting at room temperature for 10 minutes, the colloidal gold immunochromatographic test strips are removed, and the T-C line ratio is measured using a test strip reader to quantitatively analyze the concentration of the analyte in the sample. This portable sample pretreatment device is a convenient, easy-to-use, integrated, and automated sample pretreatment device and method, further simplifying the testing process, reducing time and labor costs, and improving the level of rapid on-site food safety testing.

[0014] Furthermore, the first injection port, the first detection port, the second injection port, and the second detection port are all located on the top surface of the device body. Placing all the ports on the top surface of the device body makes the structure more compact, conforms to user operating habits, facilitates detection operations, and allows for convenient comparison of two sets of samples or sample extracts with colloidal gold immunochromatographic test strips.

[0015] Furthermore, the first reaction chamber and the second reaction chamber are arranged adjacent to each other, and the first detection port and the second...

[0016] The detection wells are arranged adjacent to each other. This layout facilitates the observation of changes in samples or sample extracts in the two reaction chambers without the need to frequently shift the line of sight or adjust the device angle. It allows for simultaneous detection and comparison of test strip results, and enables more intuitive and convenient visual observation to determine the color development of the T and C lines for semi-quantitative screening.

[0017] Furthermore, the bottom of the purification chamber is connected to the first reaction chamber via a first passage; the second reaction chamber is connected to the second sample inlet via a second passage; the second sample inlet has a plug-in section protruding from the side of the device body. There are two sets of connecting channels, consisting of a purification chamber-first passage-first reaction chamber sample introduction system and a second sample inlet-second passage-second reaction chamber sample introduction system. These two sets of connecting channels are independently configured. The purification chamber has a first passage at its bottom, allowing the purified sample to be directly introduced into the reaction chamber for reaction; simultaneously, the second sample inlet is directly connected to the second passage to introduce a blank control into the reaction chamber for reaction. This design avoids sample contamination affecting the detection data, allowing the test sample or sample extract to react simultaneously with the blank control, improving detection efficiency and solving the main problems of cumbersome processes, high contamination risk, and poor portability in traditional pretreatment devices. A plug-in section protruding from the side of the device body is formed in the second injection port. This plug-in section is used to connect the device for placing the blank control. The blank control is injected into the second injection port for reaction without contact with external contamination, which improves the detection efficiency and significantly enhances the reliability of the detection results.

[0018] Furthermore, the main body of the device is entirely transparent. Designing the main body of the device to be transparent allows for timely and quick observation of changes within the reaction chamber, and rapid and effective control of the detection progress. It also allows for direct observation of the sieve plate filtration status to check for blockages or poor flow, enabling timely adjustments. Additionally, it allows for observation of the degree of contamination on the internal surfaces of the device, facilitating internal cleaning and preventing detection errors caused by impurities within the device.

[0019] Furthermore, the purification chamber is formed by a first sieve plate and a second sieve plate arranged vertically.

[0020] A first and second sieve plate are detachably connected to the inner wall of the purification chamber. The inner wall of the purification chamber has an arc-shaped groove, and both the first and second sieve plates have arc-shaped protrusions on their edges. These protrusions engage with clips that enter the arc-shaped grooves, enabling a detachable connection between the first or second sieve plate and the purification chamber. When not in use or after use, the first and second sieve plates can be detached from the purification chamber under external force for individual cleaning and sterilization. They can then be placed back into the purification chamber for the next use. The advantages of this purification chamber design lie in its detachable maintainability and reduced wear and tear.

[0021] Furthermore, the purification layer is formed by densely packed purification agent particles between adjacent sieve plates. The purification agent particles are readily available and have good adsorption effects. As the sample or sample extract passes through the purification agent particles, the solution within the sample or sample extract that affects the data is dissolved and removed, increasing the reliability of the measured data and demonstrating the highly efficient purification capability of the purification layer.

[0022] Furthermore, the sieve plate is a hydrophilic sieve plate; the inner diameter of the sieve holes on the sieve plate is 20 μm. The sieve plate can filter out solid impurities in the sample or sample extract, ensuring the quality of the filtered sample or sample extract, preventing it from blocking subsequent immunochromatographic reactions, and improving the accuracy of the data.

[0023] Furthermore, the syringe includes a filter sieve plate at its outlet, with 50μm inner diameter sieve holes. The filter sieve plate has 50μm sieve holes evenly distributed on its surface. For solid samples, after crushing or homogenizing, there will be a large number of larger particle size solid impurities. In this case, a filter sieve plate needs to be added to the syringe to coarsely filter the sample or sample extract before injecting it into the sample chamber, preventing impurities from directly clogging the sieve holes on the first sieve plate, thus reducing sample filtration efficiency or preventing flow.

[0024] This utility model also discloses a portable device for combining sample pretreatment and rapid test strip detection, including...

[0025] Includes: the aforementioned pretreatment device and colloidal gold immunochromatographic test strip.

[0026] The beneficial effects of this invention are as follows: Colloidal gold immunochromatography, based on the principle of antigen-antibody specific reaction, integrates the visualization advantages of colloidal gold detection with the high specificity, speed, and convenience of immunochromatography. It possesses advantages such as high sensitivity, high throughput, and ease of use, making it suitable for rapid on-site sample detection. This effectively overcomes the shortcomings of traditional detection methods, such as high cost, long processing time, high usage barriers, and poor portability. It can be widely used in the accurate identification and screening of pollutants in food safety regulatory departments at all levels, vegetable production and wholesale bases, agricultural markets, epidemic prevention and control, and environmental protection, and is gradually becoming a personal health self-testing tool in households. Attached Figure Description

[0027] Figure 1 A perspective view of the portable sample pretreatment device and the test strip rapid testing device provided by this utility model.

[0028] Figure 2 A cross-sectional view of the portable sample pretreatment device and the test strip rapid testing device provided by this utility model.

[0029] Figure 3The portable sample pretreatment device and the test strip rapid testing device provided by this utility model are three-dimensional Figure 1 .

[0030] Figure 4 A cross-sectional view of the syringe provided by this utility model.

[0031] Figure 5 The three-dimensional syringe provided by this utility model Figure 2 .

[0032] Figure 6 The three-dimensional sieve plate provided by this utility model Figure 3 .

[0033] Figure 7 The three-dimensional filter screen plate provided by this utility model Figure 4 .

[0034] Figure 8 The images show the detection images of thiamethoxam test strips containing thiamethoxam standard solutions of different concentrations, obtained by the portable sample pretreatment and rapid test strip combined device of this utility model, as well as the standard curves fitted by software.

[0035] Figure 9 This utility model relates to a portable sample pretreatment and rapid test strip combined device for honey sample pretreatment, and obtains detection images of honey matrix solutions containing a series of concentrations of thiamethoxam using test strips, as well as a standard curve of thiamethoxam in the honey matrix fitted by software.

[0036] Figure 10 This utility model relates to a portable sample pretreatment and rapid test strip device for cucumber sample pretreatment, which acquires detection images of cucumber matrix solutions containing a series of concentrations of thiamethoxam using test strips, as well as a standard curve of thiamethoxam in the cucumber matrix fitted by software.

[0037] Figure 11 This utility model relates to a portable sample pretreatment and rapid test strip device for apple juice sample pretreatment, and for acquiring detection images of apple juice matrix solutions labeled with thiamethoxam (5 ng / g) using test strips.

[0038] The components are as follows: 1-Main body of the device; 11-Sample inlet chamber; 111-First sample inlet; 13-First reaction chamber; 131-First detection port; 12-Purification chamber; 121-Arc groove; 14-Second reaction chamber; 141-Second sample inlet; 142-Second detection port; 143-Plug-in section; 151-First passage; 152-Second passage; 16-Top surface; 2-Sieve plate; 21-First sieve plate; 211-Arc protrusion; 22-Second sieve plate; 23-Purifying agent particles; 24-Sieve holes on the sieve plate; 3-Purification layer; 4-Injector; 41-Outlet; 42-Filter sieve plate; 421-Sieve holes on the filter sieve plate. Detailed Implementation

[0039] To enable those skilled in the art to better understand the present invention, the technical solutions in the embodiments of the present invention are clearly and completely described. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort should fall within the scope of protection of the present invention.

[0040] like Figures 1-3 As shown, a portable sample pretreatment device includes an overall transparent device body 1, which is provided with at least a sample injection chamber 11, a purification chamber 12, a purification layer 3, a first reaction chamber 13, and a second reaction chamber 14.

[0041] The top of the sample injection chamber 11 is open to form a first sample injection port 111 for placing the sample into the sample injection chamber 11. Below the sample injection chamber 11 is a purification chamber 12, which is formed by at least two sieve plates 2 arranged vertically and vertically, and the side wall between the two sieve plates 2. In this embodiment, there are two sieve plates 2, including a first sieve plate 21 and a second sieve plate 22. The purification chamber 12 is formed by the first sieve plate 21 and the second sieve plate 22 arranged vertically and horizontally, and the side wall between the first sieve plate 21 and the second sieve plate 22. Of course, in other embodiments, there may be several sieve plates 2. In this case, the same or different purification layers 3 can be set between adjacent sieve plates 2, depending on the actual sample. The sample to be tested or the sample extract can be screened multiple times and graded to filter out unnecessary impurities, so that the final purified sample or extract meets the specified reaction standard and the reaction data is more accurate.

[0042] The first sieve plate 21 and the second sieve plate 22 are detachably connected to the inner wall of the purification chamber 12. Specifically, the inner wall of the purification chamber 12 is provided with an arc-shaped groove 121, and the edges of the first sieve plate 21 and the second sieve plate 22 both form arc-shaped protrusions 211. These arc-shaped protrusions 211 are tightly fitted into the arc-shaped groove 121, realizing the detachable connection between the first sieve plate 21 or the second sieve plate 22 and the purification chamber 12. When not in use or after use, the first sieve plate 21 and the second sieve plate 22 can be detached from the purification chamber 12 under external force, removed separately, cleaned and sterilized, and then placed back into the purification chamber 12 for the next use. Of course, in other embodiments, the detachable structure of the prior art can also be used, and there is no specific limitation. The sample inlet chamber 11 and the purification chamber 12 are connected, and their inner diameters are approximately equal, which is conducive to the disassembly of the sieve plate 2.

[0043] Sieve plate 2 is a hydrophilic sieve plate, meaning that sieve plate 2 is made of a hydrophilic material, such as... Figure 6 As shown, the sieve plate 2 has uniformly distributed sieve holes 24 with an inner diameter of D1, which is 20 μm. This D1 can filter out most of the impurities in the sample or sample extract, ensuring the quality of the filtered sample or sample extract, allowing for a more complete subsequent reaction and improving the accuracy of the data. The sieve plate 2 mainly improves filtration efficiency and filtration quality through precise control of the sieve hole diameter and optimization of the sieve plate material properties.

[0044] The purification layer 3, located inside the purification chamber 12, is formed by densely packed purification agent particles 23 between adjacent sieve plates 2, meaning the purification agent particles 23 fill the entire space between adjacent sieve plates 2. This allows the sample to be tested or its extract to pass through the purification agent particles 23, dissolving and removing solutions within the sample that might affect the data, thus increasing the reliability of the measured data. In this invention, the purification layer 3 is composed of multiple purification agent particles 23. However, the internal filtration and filling material of the purification layer 3 is not limited to purification agent particles 23; other substances capable of filtration and purification within the purification layer 3 are also within the protection scope of this invention.

[0045] The first reaction chamber 13 is connected to the purification chamber 12 via a first passage 151. The first passage 151 extends along the length of the main body 1 of the device, directly introducing the purified sample into the first reaction chamber 13 for reaction. This avoids recontamination after purification and also shortens the reaction time, making the main body 1 more compact, portable, and convenient to use. A first detection hole 131 is provided at the top of the first reaction chamber 13, extending along the height of the main body 1 to its top surface 16. After the reaction is completed, a test strip is directly inserted into the first detection hole 131 to contact the internal liquid and obtain the test result.

[0046] The second reaction chamber 14 is set independently from the first reaction chamber 13, that is, the first reaction chamber 13 and the second reaction chamber 14 are not connected to each other. The second reaction chamber 14 is connected to the second sample inlet 141 through the second channel 152. The second channel 152 extends along the length of the device body 1 to avoid the blank control being contaminated and affecting the detection data. A second detection hole 142 is provided at the top of the second reaction chamber 14. The second detection hole 142 extends along the height of the device body 1 to its top surface 16. After the reaction is completed, the test strip is directly inserted into the second detection hole 142 so that it comes into contact with the liquid inside to obtain the detection result. The first reaction chamber 13 and the second reaction chamber 14 are set independently, and the first sample inlet 111 is directly connected to the purification chamber 12. The bottom of the purification chamber 12 is connected to the first reaction chamber 13 through the first passage 151. Similarly, the second sample inlet 141 is connected to the second reaction chamber 14 through the second passage 152, forming two independent reaction systems. The sample to be tested or sample extract and blank control can be reacted simultaneously through the first sample inlet 111 and the second sample inlet 141, respectively, which improves the detection efficiency and solves the main problems of cumbersome process, high risk of contamination and poor portability in traditional pretreatment devices.

[0047] In order to enable quick and accurate connection of the device for placing blank control, the second injection port 141 is formed with a plug section 143 protruding from the side of the device body 1. Specifically, in this embodiment, the plug section 143 protrudes from the top surface 16 of the device body 1. The plug section 143 is used to connect the device for placing blank control, and inject the blank control into the second injection port 141 for reaction without contact with external contamination, thereby improving the detection efficiency and significantly enhancing the reliability of the detection results.

[0048] The first sample inlet 111, the first detection hole 131, the second sample inlet 141, and the second detection hole 142 are located on the side of the device body 1. This utility model demonstrates one possible arrangement of the openings: the first sample inlet 111, the first detection hole 131, the second sample inlet 141, and the second detection hole 142 are located on the top surface 16 of the device body 1. This arrangement results in a more compact structure, facilitating sample injection and other operations, and conforming to user operating habits. Furthermore, whether the first sample inlet 111, the first detection hole 131, the second sample inlet 141, and the second detection hole 142 are located on other sides of the device body 1, or on other different sides of the device body 1, all fall within the protection scope of this utility model.

[0049] like Figures 2-3As shown, the first reaction chamber 13 and the second reaction chamber 14 are arranged adjacent to each other, and the first detection hole 131 and the second detection hole 142 are arranged adjacent to each other. This layout is conducive to observing the changes of the sample or sample extract in the two reaction chambers without frequently shifting the line of sight or adjusting the device angle. After the test strip detects the result, it is possible to more intuitively observe with the naked eye to judge the color development of the T and C lines for semi-quantitative screening. At the same time, the main body 1 of the device is made of transparent material. The overall transparent main body 1 of the device can observe the changes in the reaction chamber in a timely and quick manner, and quickly and effectively control the progress of the detection state. It can also directly observe whether the filtration state of the sieve plate 2 is blocked or the flow is not smooth, and make timely adjustments. It can also observe the degree of contamination on the internal surface of the device and clean the inside to avoid detection errors caused by impurities inside the device.

[0050] like Figures 4-5 As shown, the aforementioned portable sample pretreatment device and test strip rapid testing device also includes a syringe 4, which is easy to carry and use. To perform initial screening of the sample to be tested, a filter sieve plate 42 is provided at the discharge port 41 of the syringe 4.

[0051] like Figure 7 As shown, the inner diameter of the sieve hole 421 on the filter sieve plate is D2, which is 50μm. The filter sieve plate 42 is evenly distributed and is mainly used for solid sample extracts. After crushing or homogenizing, there will be a lot of impurities. At this time, it is necessary to add the filter sieve plate into the syringe. D2>D1, so that the solid sample extract is coarsely filtered before entering the sample injection chamber 11. This avoids the impurities being too large and directly blocking the sieve hole 24 on the first sieve plate 12, which would reduce the sample filtration efficiency or prevent flow, and reduce the frequency of sieve plate 2 replacement.

[0052] A portable sample pretreatment and rapid test strip combined device includes: a pretreatment device with the above-described structure, and a test strip. In this invention, the test strip is a colloidal gold immunochromatographic test strip, which is manufactured using existing technology. Of course, in other embodiments, the test strip can also be made of other labeling materials, and there are no specific limitations. As a combined device, it can perform detection using any matching card reader, and further details will not be elaborated.

[0053] This utility model discloses a portable sample pretreatment and rapid test strip combined device. First, the main body 1 of the device is placed vertically on a table. The components are then sequentially filled into the purification chamber 12 in the order of second sieve plate 22 – purification layer 3 – first sieve plate 21. The assembly is completed by pressing the piston of the syringe 4 to secure each component. Next, the sample to be tested or sample extract is poured into a clean syringe 4 and coarsely filtered through the filter sieve plate 42 of the syringe 4. The coarsely filtered sample is then directly injected into the sample injection chamber 11 by pushing the syringe. After passing through the first sieve plate 21, the sample enters the purification chamber 12. The purification chamber 12 contains a purification layer 3. After passing through the purification layer 3, the sample is filtered again by the second sieve plate 22 and then flows through the first passage 151 into the first reaction chamber 13 for reaction. Simultaneously, a blank control (i.e., a solution without the test sample) is injected into the second injection well 141. The sample flows through the second passage 152 into the second reaction chamber 14 for reaction, with a waiting time of approximately 2 minutes. Subsequently, two test strips are placed into the first detection well 131 and the second detection well 142, respectively. After reacting at room temperature for 10 minutes, the test strips are removed and the T-C line ratio is measured using a test strip reader to quantitatively analyze the concentration of the analyte in the sample. This utility model provides a convenient sample pretreatment device and a rapid test strip testing device that is easy to carry and operate. It is an integrated, automated sample pretreatment device and method that further simplifies the testing process, reduces time and labor costs, and improves the level of rapid on-site food safety testing.

[0054] This device can automatically complete the purification reaction in the rapid on-site testing process for food safety within 2 minutes, significantly improving the automation and ease of operation of sample pretreatment. The sieve plate and the purification layer are detachable, which helps reduce testing costs. Furthermore, the main body of the device is manufactured using 3D printing, allowing for stable, large-batch production. Its small size, portability, and practicality address the shortcomings of most commonly used pretreatment methods, such as numerous steps, long processing times, and difficulty in achieving high-throughput on-site screening of large-scale samples. When used in conjunction with immunochromatographic test strips, this device completes a series of operations, including sample extraction, addition, purification, and detection, within 15 minutes, enabling rapid sample pretreatment and on-site screening of target analytes for various food types.

[0055] The present invention discloses a method for using a portable sample pretreatment and rapid test strip combined device, comprising the following steps:

[0056] Step 1: Device Design, Fabrication and Assembly

[0057] The main body 1 of the device is 3D printed using transparent resin as the printing material and selective curing technology with photosensitive resin. Internally, it includes a first sample inlet 111, a sample chamber 11, a purification chamber 12, a first detection port 121, a first reaction chamber 13, a second detection port 142, a second reaction chamber 12, and a second sample inlet 141, connected to each reaction chamber via a first passage 151 and a second passage 152. The sieve plate 2 is a hydrophilic sieve plate with a pore size of 20 μm, its shape and size being approximately the same as the first sample inlet 111, and is embedded within the main body 1. The purification chamber 12 is formed by two sieve plates 2 spaced apart. The purification layer 3 is composed of appropriate mass fractions of N-propylethylenediamine, activated carbon, graphitized carbon black, or cross-linked polyvinylpyrrolidone, and is filled within the purification chamber 12 of the main body 1. The device is easy to assemble. Take the main body 1 and place it upright on the table. Fill the purification chamber 12 in the order of second sieve plate 22 - purification layer 3 - first sieve plate 21. Press each component firmly with the piston of syringe 4.

[0058] Step 2: Sample loading and coarse filtration

[0059] For solid sample extracts with a high amount of solid impurities, a filter sieve plate 42 with a pore size of 50 μm is attached to the front of the syringe 4 for coarse filtration. For simple liquid samples with fewer solid impurities, this step can be omitted, and the sample can be directly poured into a clean syringe 4 to the 1 mL mark or the liquid sample can be directly drawn from the syringe 4.

[0060] Step 3: Sample Extraction

[0061] For solid samples, after crushing or homogenizing, weigh 1g of the ground sample into a clean syringe 4, and pour PBS buffer (0.01M, pH 7.4) containing 10% methanol into syringe 4 to 5mL mark, insert the stopcock and shake by hand for 5s to mix evenly; then load the test sample or sample extract and perform coarse filtration as described in step two.

[0062] Step 4: Sample Injection

[0063] Press the plunger of syringe 4 and add 1 mL of the sample to be tested or sample extract into injection chamber 11. At the same time, inject 100 μL of PBS buffer (0.01 M, pH 7.4) containing 10% methanol as a blank control into the second reaction chamber 14 through the second injection well 141. The second detection well 142 is used for the quality control test strip - colloidal gold immunochromatographic test strip.

[0064] Step 5: Purification

[0065] Place the device on the table and wait for the sample or sample extract to pass through the purification chamber 12 and enter the first reaction chamber 13 via the first passage 151. The waiting time is about 2 minutes.

[0066] Step Six: Testing

[0067] Using the previously developed anti-thiamethoxam specific monoclonal antibody and artificial antigen, and colloidal gold as a signal marker, an immunochromatographic test strip was constructed, and a corresponding standard curve was generated. In real sample testing, two of the above colloidal gold test strips were inserted into the first reaction chamber 13 of the first detection well 131 and the second reaction chamber 14 of the second detection well 142, respectively. After reacting at room temperature for 10 minutes, the colloidal gold test strips were removed.

[0068] Step 7: Result Interpretation

[0069] Using a quality control test strip—colloidal gold test strip—the color development of the T and C lines is visually observed for semi-quantitative screening. Based on the constructed sample matrix standard curve, the T-C line ratio is determined using a test strip reader. Quantitative analysis is then performed based on the previously constructed matrix standard curve to calculate the residual level of the target analyte in each sample.

[0070] The present invention will be further described in detail below. Detailed implementation methods and specific operating procedures are given. The embodiments will help to understand the present invention, but the protection scope of the present invention is not limited to the following embodiments.

[0071] In the following embodiments, this invention uses a colloidal gold immunochromatographic test strip targeting thiamethoxam (a neonicotinoid pesticide) as the detection tool. Those skilled in the art will understand that for other target compounds (such as other pesticides, mycotoxins, etc.), corresponding specific immunochromatographic test strips are required, and the labeling material, in addition to colloidal gold, can also be colored latex microspheres, carbon nanotubes, etc. Semi-quantitative screening can be performed by visual interpretation, while quantitative analysis requires a corresponding portable test strip reading device. Furthermore, the type and content of the purification agent and sample extract need to be optimized according to the chemical properties of the analyte.

[0072] Example 1

[0073] This embodiment provides a portable device for combining sample pretreatment and rapid test strip testing, such as... Figure 1 As shown, this device is used to detect thiamethoxam residues in honey, and includes the following steps:

[0074] Step 1: Device Design, Fabrication and Assembly

[0075] A three-dimensional view of the main body 1 of the device is shown below. Figure 1 As shown. A cross-sectional view of the main body 1 of the device is shown below. Figure 2As shown. The main body 1 of the device is 3D printed using transparent resin as the printing material and photosensitive resin selective curing technology. The sieve plate 2 is a hydrophilic sieve plate with a pore size of 20μm, and its shape and size are approximately the same as those of the first sample inlet 111. The purification layer 3 is 20% N-propylethylenediamine by mass. The device is easy to assemble. Take the main body 1 and place it upright on the table. Fill the purification chamber 12 in the order of second sieve plate 22—purification layer 3—first sieve plate 21, and press each component firmly with the piston of syringe 4. The steps involved are described below:

[0076] Step 2: Colloidal gold immunochromatographic test strip detection

[0077] (1) Loading and extraction of the sample to be tested: Take 1g of honey sample, add the sample extraction solution (0.01M, pH 7.4 PBS buffer containing 10% methanol), manually shake for 5s to mix evenly, and transfer the extraction solution to a clean syringe 4.

[0078] (2) Sample injection: Press the plunger of syringe 4 and add 1 mL of diluted sample into injection chamber 11. At the same time, inject 1 mL of 10% methanol-PBS buffer solution into the second reaction chamber 14 through the second injection port 141.

[0079] (3) Purification: Wait for the extract of the sample to be tested to pass through the purification chamber 12 and the first passage 151 in sequence and arrive at the first reaction chamber 13. The waiting time is about 2 minutes.

[0080] (4) Detection: Take two colloidal gold test strips for thiamethoxam and insert them into the first reaction chamber 13 of the first detection well 131 and the second reaction chamber 14 of the second detection well 142, respectively. After reacting at room temperature for 10 minutes, take out the test strips.

[0081] (5) Result interpretation: Using the quality control test strip, the color development of the T and C lines of the sample to be tested is interpreted by visual observation to perform semi-quantitative screening. Honey matrix extract is prepared according to the above pretreatment method and a standard curve is constructed. The ratio of T to C lines is determined by the test strip reader. Quantitative analysis is performed based on the matrix mark to calculate the thiamethoxam residue level in the sample.

[0082] Step 3: Evaluation of matrix effect

[0083] First, a standard curve was constructed for thiamethoxam standard solutions. A series of thiamethoxam standard solutions of varying concentrations were prepared using sample extraction buffer (0.01M, pH 7.4 PBS buffer containing 10% methanol). 100 μL of each solution was added to a 96-well plate. The sample pad end of the colloidal gold immunochromatographic test strip was inserted into the solution, and the reaction was carried out at room temperature for 10 min. Based on the color development of the T line, the lowest visually perceptible concentration was used as the detection limit of the colloidal gold immunochromatographic test strip. Simultaneously, the signal values ​​of the T and C lines of each test strip were recorded using a card reader, and the T / C ratio was calculated. Subsequently, the concentration of thiamethoxam was plotted on the x-axis, and the inhibition rate corresponding to the T / C value was plotted on the y-axis. A four-parameter function was used to fit the equation in Prism8 software, with the half-inhibition rate (IC50) as the threshold. 50 Numerical values ​​are used as sensitivity evaluation indicators. The standard curve and detection graph for the detection of thiamethoxam using a buffer solution constructed in this embodiment are shown below. Figure 8 As shown.

[0084] Take 1g of negative blank honey sample verified by LC-MS / MS, extract the sample using 5mL of PBS containing 10% methanol, weigh 0.2g of purification agent (w / v: 20%) into a 1.5mL clean centrifuge tube, then pipette 1mL of the sample into the centrifuge tube, vortex thoroughly for 2min, and centrifuge at 4℃, 5000g for 5min. After centrifugation, prepare a series of thiamethoxam solutions containing the matrix using the honey extract, and read the signal values ​​of the T and C lines for each concentration using colloidal gold immunochromatographic test strips. Calculate the T / C value and its corresponding inhibition rate, and further construct a thiamethoxam standard curve for the honey matrix.

[0085] The results showed that the fitting R of the honey matrix calibrator was [missing information]. 2 The value is 0.9855. Calculate IC. 50 The value was 0.14 ng / mL. Comparison with the standard curve established using 10% methanol-PBS buffer (blank control) revealed that the detection sensitivity of the honey matrix was similar to that of the blank control (R0). 2 =0.9938, IC 50 (0.11 ng / mL). Therefore, this sample pretreatment method can effectively eliminate the influence of the matrix on the detection method.

[0086] Step 4: Add a recycling test

[0087] Thiamethoxam standard was added to honey samples that had been validated as negative by LC-MS / MS for spiking recovery testing. 1 g of sample was added to a 5 mL centrifuge tube, with spiked concentrations of 1 and 5 ng / g, and three replicates were prepared for each tube. After addition, the samples were allowed to stand at room temperature for 2 hours. Then, sample pretreatment and colloidal gold immunochromatographic assay were performed as described in step two. The T and C signals of each assay strip were recorded using a card reader, and the corresponding T / C ratio and inhibition rate were calculated. The inhibition rate was then substituted into the standard curve of the honey matrix to calculate the thiamethoxam content in the sample, and the spiking recovery rate was calculated.

[0088] The standard curve for the detection of thiamethoxam in honey matrix constructed in this embodiment, and the actual detection sample image are shown below. Figure 9 As shown.

[0089] The results of the recovery tests on honey samples are shown in Table 1. The average recovery rate of thiamethoxam detected by this device was 77.3%–102.3%, with a coefficient of variation of 5.5%–7.9%. According to the pesticide residue test guidelines (NY / T 788-2018), the average recovery rate of this sample pretreatment method meets the requirements for quantitative analysis.

[0090] Table 1 shows the detection results of honey-added samples based on a portable sample pretreatment and rapid test strip device.

[0091]

[0092] Example 2

[0093] This embodiment provides a portable sample pretreatment and rapid test strip device for detecting thiamethoxam residues in cucumbers. It includes the following steps:

[0094] Step 1: Device Design, Fabrication and Assembly

[0095] The main body 1 of the device is 3D printed using transparent resin as the printing material and selective curing technology with photosensitive resin. The sieve plate 2 is a hydrophilic sieve plate with a pore size of 20 μm, and its shape and size are approximately the same as those of the first sample inlet 111. The purification layer 3 is 20% N-propylethylenediamine by mass. The device is easy to assemble. Take the main body 1 and place it upright on the table. Fill the purification chamber 12 in the order of second sieve plate 22 - purification layer 3 - first sieve plate 21, and press each component firmly with the piston of syringe 4. The steps involved are described below:

[0096] Step 2: Colloidal gold immunochromatographic test strip detection

[0097] (1) Sample dilution: Weigh 1g of homogenized cucumber sample, add 5mL of sample extraction solution (0.01M, pH 7.4 PBS buffer containing 10% methanol), and manually shake for 5s to mix evenly.

[0098] (2) Loading and coarse filtration of the sample or sample extract: Fill the 5mL clean syringe 4 with the hydrophilic filter sieve plate 42 and pour in an appropriate volume of cucumber sample extract.

[0099] (3) Sample injection: Press the plunger of syringe 4 and add 1 mL of the coarsely filtered sample extract into the injection chamber 11. At the same time, inject 1 mL of 10% methanol-PBS buffer solution into the second reaction chamber 14 through the second injection port 141.

[0100] (4) Purification: Wait for the extract of the sample to be tested to pass through the purification chamber 12 and the first passage 151 in sequence and arrive at the first reaction chamber 13. The waiting time is about 2 minutes.

[0101] (5) Detection: Take two colloidal gold immunochromatographic test strips for thiamethoxam and insert them into the first reaction chamber 13 of the first detection well 131 and the second reaction chamber 14 of the second sample injection well 141 respectively. After reacting at room temperature for 10 minutes, take out the test strips.

[0102] (6) Result interpretation: Using the quality control test strip, the color development of the T and C lines of the sample to be tested is interpreted by visual observation to perform semi-quantitative screening. Cucumber matrix extract is prepared according to the above pretreatment method and a standard curve is constructed. The ratio of T to C lines is determined by the test strip reader. Quantitative analysis is performed based on the matrix mark to calculate the thiamethoxam residue level in the sample.

[0103] Step 3: Evaluation of matrix effect

[0104] Cucumber samples that tested negative by LC-MS / MS were pretreated according to the above procedure. The sample extract was used to prepare a series of concentrations of thiamethoxam, with 100 μL of each concentration added to a 96-well plate. One end of the sample pad of the colloidal gold immunochromatographic test strip was inserted into the solution, and the reaction was carried out at room temperature for 10 min. The signal values ​​of the T and C lines of each test strip were recorded using a card reader. The T / C ratio at each concentration was calculated using a blank control matrix as a control. Subsequently, the concentration of thiamethoxam was plotted on the x-axis, and the inhibition rate corresponding to the T / C value was plotted on the y-axis. A four-parameter function fitting equation was used in Prism8 software, with the half-inhibition concentration (IC50) as the modulus. 50 Numerical values ​​are used as sensitivity evaluation indicators.

[0105] The results showed that the fitting R of the cucumber matrix markings was [missing information]. 2 The value is 0.9901. Calculate IC. 50The value was 0.10 ng / mL. Comparison with the standard curve established using 10% methanol-PBS buffer (blank control) revealed that the detection sensitivity of cucumber matrix and blank matrix was similar (IC50). 50 (0.11 ng / mL). Therefore, this sample pretreatment method can effectively eliminate the influence of cucumber matrix on the detection method.

[0106] Step 4: Add a recycling test

[0107] Thiamethoxam standard was added to cucumber samples that had been validated as negative by LC-MS / MS for a spiking recovery test. 1 g of sample was added to a 5 mL centrifuge tube, with spiked concentrations of 1 and 5 ng / g, and three replicates were prepared for each tube. After addition, the samples were allowed to stand at room temperature for 2 hours. Then, sample pretreatment and colloidal gold immunochromatographic assay were performed as described in step two. The T and C signals of each assay strip were recorded using a card reader, and the corresponding T / C ratio and inhibition rate were calculated. The inhibition rate was then substituted into the standard curve of the cucumber matrix to calculate the thiamethoxam content in the sample, and the recovery rate was calculated.

[0108] The standard curve for the detection of thiamethoxam in cucumber matrix constructed in this embodiment, and the actual detection image are shown below. Figure 10 As shown.

[0109] The results of the recovery test on cucumber samples are shown in Table 2. The average recovery rate of thiamethoxam detected by this device was 90.9%–107.4%, with a coefficient of variation of 3.6%–7.6%. According to the pesticide residue test guidelines (NY / T 788-2018), the average recovery rate of this sample pretreatment method meets the requirements for quantitative analysis.

[0110] Table 2 shows the detection results of cucumber samples using a portable sample pretreatment and rapid test strip device.

[0111]

[0112] Example 3

[0113] This embodiment provides a portable sample pretreatment and rapid test strip device for semi-quantitative screening of thiamethoxam residues in apple juice.

[0114] Step 1: Device Design, Fabrication and Assembly

[0115] The main body 1 of the device is 3D printed using transparent resin as the printing material and selective curing technology with photosensitive resin. The sieve plate 2 is a hydrophilic sieve plate with a pore size of 20 μm, and its shape and size are approximately the same as those of the first sample inlet 111. The purification layer 3 is 20% N-propylethylenediamine by mass. The device is easy to assemble. Take the main body 1 and place it upright on the table. Fill the purification chamber 12 in the order of second sieve plate 22 - purification layer 3 - first sieve plate 21, and press each component firmly with the piston of syringe 4. The steps involved are described below:

[0116] Step 2: Colloidal gold immunochromatographic test strip detection

[0117] (1) Loading and coarse filtration of the sample or sample extract: Fill the 5mL clean syringe 4 with the hydrophilic filter sieve plate 42 and pour in an appropriate volume of uniformly mixed fruit juice sample.

[0118] (2) Sample injection: Press the plunger of syringe 4 and add 1 mL of sample into injection chamber 11. At the same time, inject 1 mL of PBS buffer into the second reaction chamber 14 through the second injection port 141.

[0119] (3) Purification: Wait for the extract of the sample to be tested to pass through the purification chamber 12 and the first passage 151 in sequence and arrive at the first reaction chamber 13. The waiting time is about 2 minutes.

[0120] (4) Detection: Take two colloidal gold immunochromatographic test strips for thiamethoxam and insert them into the first reaction chamber 13 of the first detection well 131 and the second reaction chamber 14 of the second sample injection well 141 respectively. After reacting at room temperature for 10 minutes, take out the colloidal gold immunochromatographic test strips.

[0121] (5) Result interpretation: Combined with the quality control test strip, the color development of the T and C lines of the sample to be tested is interpreted by visual observation in order to carry out semi-quantitative screening.

[0122] Step 3: Evaluation of matrix effect

[0123] First, a series of thiamethoxam standard solutions of various concentrations were prepared using PBS buffer (0.01M, pH=7.4). 100 μL of each solution was added to a 96-well plate. The sample pad end of the colloidal gold immunochromatographic test strip was inserted into the solution, and the reaction was carried out at room temperature for 10 min. Using PBS as a control, the lowest concentration at which the T line disappeared, as judged by visual inspection, was taken as the limit of detection (LOD) of the colloidal gold immunochromatographic test strip. The results showed that the LOD was 5 ng / mL. Then, apple juice samples that were negative for thiamethoxam by LC-MS / MS were pretreated. The specific procedures were as described in step two. The color development of the T and C lines on the test strip was observed visually, and the effect of the apple juice sample on the detection effect of the colloidal gold immunochromatographic test strip in well 142 was analyzed to evaluate the purification effect of the device on the apple juice sample matrix.

[0124] Step 4: Add a recycling test

[0125] Thiamethoxam standard was added to apple juice samples that were negative by LC-MS / MS for a spiking recovery test. 1 mL of sample was transferred to a 5 mL centrifuge tube, and the spiked concentrations were set at 1 and 5 ng / g, respectively, based on the visual disappearance concentration of the test strip, with three replicates. After standing at room temperature for 2 hours, sample pretreatment and colloidal gold immunochromatographic test strip detection were performed as described in step two. The reliability and accuracy of this portable sample pretreatment and rapid test strip coupling device were analyzed by visually observing the T-line development and combining it with the colloidal gold immunochromatographic test strip in the second detection well (142).

[0126] The matrix purification effect and the addition and recovery results in this embodiment are as follows: Figure 11 As shown.

[0127] The results showed that using a portable sample pretreatment and rapid test strip device, with 20% N-propylethylenediamine as the filler, eliminated the interference of apple juice samples on the color development of the colloidal gold immunochromatographic test strip, making the T and C lines similar to those of the colloidal gold immunochromatographic test strip in the second detection well 142 using a buffer solution. Furthermore, for apple juice samples spiked at 5 ng / g, the portable sample pretreatment and rapid test strip device showed complete disappearance of the T line, consistent with the visual detection limit using thiamethoxam standard solution, indicating that this sample pretreatment method meets the requirements for semi-quantitative analysis.

[0128] The above-described embodiments only illustrate three implementation methods of this application, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent application. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this utility model, and these modifications and improvements should also be considered within the scope of protection of this utility model. Therefore, the scope of protection of this patent application should be determined by the appended claims. All components not explicitly stated in these embodiments can be implemented using existing technology.

Claims

1. A portable sample pretreatment device, characterized in that, Includes a device body (1), which has at least the following components: The top of the sample inlet chamber (11) is open to form the first sample inlet hole (111). The purification chamber (12) is located below the sample injection chamber (11) and is formed by at least two sieve plates (2) arranged vertically. A purification layer (3) is provided inside the purification chamber (12); The first reaction chamber (13) is connected to the purification chamber (12) and has a first detection hole (131). The second reaction chamber (14) is set independently from the first reaction chamber (13) and is connected to the second sample inlet (141) and the second detection port (142) respectively; The first sample inlet (111), the first detection hole (131), the second sample inlet (141), and the second detection hole (142) are located on the side of the main body (1) of the device.

2. The portable sample pretreatment device according to claim 1, characterized in that: The first sample inlet (111), the first detection hole (131), the second sample inlet (141), and the second detection hole (142) are located on the top surface (16) of the main body (1) of the device.

3. The portable sample pretreatment device according to claim 1, characterized in that: The first reaction chamber (13) and the second reaction chamber (14) are arranged adjacent to each other, and the first detection hole (131) and the second detection hole (142) are arranged adjacent to each other.

4. The portable sample pretreatment device according to claim 3, characterized in that: The bottom of the purification chamber (12) is connected to the first reaction chamber (13) through the first passage (151); the second reaction chamber (14) is connected to the second injection port (141) through the second passage (152); the second injection port (141) has a plug section (143) protruding from the side of the main body (1) of the device.

5. The portable sample pretreatment device according to claim 1, characterized in that: The main body of the device (1) is transparent.

6. The portable sample pretreatment device according to claim 1, characterized in that: The purification chamber (12) is formed by a first sieve plate (21) and a second sieve plate (22) arranged vertically. The first sieve plate (21) and the second sieve plate (22) are detachably connected to the inner wall of the purification chamber (12).

7. The portable sample pretreatment device according to claim 1, characterized in that: The purification layer (3) is formed by purification agent particles (23) densely distributed between adjacent sieve plates (2).

8. The portable sample pretreatment device according to claim 1, characterized in that: The sieve plate (2) is a hydrophilic sieve plate; the inner diameter of the sieve holes (24) on the sieve plate (2) is 20 μm.

9. The portable sample pretreatment device according to claim 1, characterized in that: It also includes a syringe (4), whose outlet (41) is provided with a filter screen plate (42), the inner diameter of the sieve holes (421) on the filter screen plate is 50μm.

10. A portable device for sample pretreatment and rapid test strip measurement, characterized in that: It includes the pretreatment apparatus as described in any one of claims 1-9, and the colloidal gold immunochromatographic test strip.

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