CN111721761A - Enzyme inhibition rapid test card and independent sample introduction card suitable for detecting pesticide residue in organic solvent extraction sample and using method thereof - Google Patents
Enzyme inhibition rapid test card and independent sample introduction card suitable for detecting pesticide residue in organic solvent extraction sample and using method thereof Download PDFInfo
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Abstract
The invention provides an enzyme inhibition rapid test card, an independent sample introduction card and a use method thereof, wherein the enzyme inhibition rapid test card is suitable for detecting pesticide residues in organic solvent extraction samples and comprises a sample introduction card, an enzyme region card and a substrate card, and a sample introduction region and a super-inert edge which completely surrounds the sample introduction region are arranged on the sample introduction card. The independent sample introduction card can be matched with a conventional enzyme inhibition rapid test card for use, and the conventional enzyme inhibition rapid test card is formed by connecting an enzyme area card and a substrate card. The enzyme inhibition rapid test card provided by the invention is additionally provided with a sample injection area card; the online volatilization of the organic solvent in the sample injection area is accelerated by utilizing the overlarge specific surface area of a paper material, after the organic solvent is volatilized, the enzyme inhibition and color development reaction are sequentially developed in the modes of folding, laminating and the like, the dual requirements of the combination of the organic phase pretreatment and the enzyme inhibition method of the sample and the zero contact of the organic solvent and the immobilized enzyme are met, and the detection sensitivity and the accuracy of the enzyme inhibition rapid-determination card are improved.
Description
Technical Field
The invention relates to an enzyme inhibition rapid test card for directly detecting organophosphorus and carbamate pesticide residues suitable for extraction pretreatment of an organic solvent sample, an independent sample introduction card capable of being matched with a conventional pesticide residue enzyme inhibition rapid test card for use and a use method thereof, and belongs to the field of laboratory analysis tools.
Background
The enzyme inhibition method is the most mature and widely used method for quickly detecting pesticide residue so far, and mainly utilizes the inhibition effect of pesticide on esterase activity to specifically detect organophosphorus and carbamate pesticides. The enzyme inhibition rapid detection card is used for detecting the pesticide residue on the paper carrier by an enzyme inhibition method, and the advantages of short detection time, low cost and the like are shown by utilizing the characteristics of cheap and easily available paper materials, strong biocompatibility and the like, so that the enzyme inhibition rapid detection card is widely applied to the field of rapid detection of pesticide residue in China. During detection, if the sample does not contain organophosphorus or carbamate pesticide residues, the enzyme activity is not inhibited, the substrate can be hydrolyzed normally to generate a certain amount of products, and certain signal changes such as color, fluorescence and the like are presented on the surface of the paper carrier; when the tested sample contains the pesticide, the pesticide components damage the enzyme activity to a certain degree, so that the substrate cannot be normally catalyzed and hydrolyzed, the generation amount of hydrolysate is reduced, and the corresponding signal change is reduced. By utilizing the principle, the enzyme inhibition rapid test card can rapidly and effectively carry out visual screening on a large number of samples.
At present, the conventional enzyme inhibition rapid test card mainly consists of two parts, namely an enzyme area and a substrate area. When in detection, firstly, a sample is dripped into an enzyme area to carry out enzyme inhibition reaction; after the reaction is sufficient, the test strip is folded in half to enable the enzyme area to be fully contacted with the substrate area, so that the enzyme color reaction is developed; and finally, comparing the chromogenic results of the rhizoma bletillae sample, and judging the pesticide residue level in the sample. Since most organic solvents have an inhibiting effect on enzyme activity, in order to avoid additional interference of organic phase solutions on immobilized enzymes, conventional enzyme inhibition tacheometers can only use aqueous phase solutions such as phosphate buffer solutions and the like as extraction solvents in the sample pretreatment process. However, most organophosphorus and carbamate pesticides are poorly water soluble and readily soluble in organic solvents. Therefore, the extraction efficiency of the pretreatment process dominated by the aqueous phase is too low. The prior literature discloses that the recovery rate of water extraction for organophosphorus and carbamate pesticide residues is mostly below 30 percent, even a lot of the recovery rate is below 10 percent. Such a low extraction rate obviously affects the accuracy and reliability of the detection result to a great extent. Even if the subsequent detection is accurate, the detection result cannot completely reflect the pesticide residue content of the sample under the condition that the extraction of the pretreatment is incomplete. Thus, the utility of conventional enzyme inhibition tacheometers is largely limited by the extraction solvent.
Therefore, it is necessary to design a new enzyme inhibition rapid test card product compatible with the pretreatment of the organic solvent sample to improve the detection performance of the existing enzyme inhibition rapid test card in practical use by breaking through the configuration mode of the traditional rapid test card.
Disclosure of Invention
The invention aims to provide an enzyme inhibition rapid test card compatible with organic phase sample pretreatment, which is compatible with an aqueous phase solvent and an organic phase solvent, and is mainly suitable for the organic phase solvent. When an organic solvent is used as a sample pretreatment extraction solvent, the enzyme inhibition rapid test card can avoid the organic solvent in a sample solution from causing extra interference on the activity of the immobilized enzyme, improve the detection sensitivity and ensure the accuracy of a detection result.
In addition, the invention also provides a sample introduction mode and a matching device which can be matched with the existing enzyme inhibition rapid-determination card for use, so that the existing enzyme inhibition rapid-determination card can directly detect the sample solution extracted by the organic solvent, and the detection performance of the enzyme inhibition rapid-determination card is improved.
In order to solve the technical problems, the invention adopts the technical scheme that:
an enzyme inhibition rapid test card suitable for detecting pesticide residues in organic solvent extraction samples is composed of a sample introduction card, an enzyme region card and a substrate card, wherein a sample introduction region and a super-inert edge surrounding and completely surrounding the sample introduction region are arranged on the sample introduction card, and an enzyme region and a hydrophobic edge surrounding and completely surrounding the enzyme region are arranged on the enzyme region card; providing a substrate area on said substrate card and a hydrophobic edge surrounding said substrate area completely;
the sample feeding area, the enzyme area and the substrate area are made of test paper or other porous sheet or plate materials (such as thick paper sheets, thick paper boards and the like) which can resist organic solvents, and a test paper substrate is preferably adopted. Wherein the enzyme area is fixed with an enzyme reagent on the test paper substrate, and the substrate area is fixed with a chromogenic substrate on the test paper substrate; the enzymatic reagent is typically cholinesterase; the chromogenic substrate is typically indophenol acetate or indole acetate.
In the invention, the pesticide residue of the organic solvent extraction sample mainly refers to organophosphorus and carbamate pesticide residues, and the organophosphorus and carbamate pesticides are poor in water solubility and easy to dissolve in an organic solvent, so that the extraction efficiency can be improved only by extracting the sample with the organic solvent, the recovery rate can reach more than 90%, and the accuracy and reliability of detection are ensured.
The hydrophobic edge can be formed by the known technology such as wax spraying printing or the like or the method or the material such as plastic film covering or the like, and the super-inert edge can be obtained by special material treatment;
the sample inlet card is surrounded by a super-inert edge which completely surrounds a sample inlet area, the super-inert refers to hydrophobic ester, and the sample inlet of the card is an organic phase solution, so the edge boundary of the sample inlet area must be neither hydrophilic nor infiltrated by an organic solvent, namely the hydrophobic ester, the edge boundary is generally made of a fluorinated material, and the fluorinated material is neither hydrophilic nor lipophilic and can be better used as the sample inlet range boundary of an organic sample. The fluorinated material may be a commonly used one such as teflon or the like.
The sample introduction card, the enzyme region card and the substrate card can be connected with each other or can be connected with each other, and the sample introduction card, the enzyme region card and the substrate card are three independent cards when not connected with each other. Can be aligned, overlapped and attached to each other for use.
The sample introduction card, the enzyme area card and the substrate card can be connected together to form a whole, so that the use is convenient.
Furthermore, the sample introduction card, the enzyme area card and the substrate card can be connected together in various ways, and after the sample introduction card, the enzyme area card and the substrate card are connected into a whole, the sample introduction area and the enzyme area, and the enzyme area and the substrate area can be completely overlapped together in a folding, aligning and other ways to be comprehensively and effectively attached and contacted; preferably, the enzyme zone card is placed between the sample card and the substrate card.
Furthermore, it is preferable that the sample introduction card, the enzyme region card and the substrate card are linearly arranged in this order and connected into a long strip shape.
Or the sample introduction card, the enzyme region card and the substrate card are connected into an L shape, preferably, the enzyme region card is arranged at the corner of the L, and the sample introduction card and the substrate card are respectively positioned at two adjacent edges of the L.
The enzyme area and the substrate area can be respectively fixed on the gasket by the enzyme or the substrate reagent and then connected with the test paper substrate, or can be directly fixed by separating corresponding areas on the test paper substrate. The immobilization of the enzyme reagent and the chromogenic substrate is well known in the art.
The configuration and the area of the sample introduction area, the enzyme area and the substrate area have no specific requirements, and can be changed according to the experimental needs.
Preferably, the sample injection region, the enzyme region and the substrate region are circular, and further preferably, the area of the enzyme region is smaller than or equal to that of the sample injection region, and the area of the enzyme region is smaller than or equal to that of the substrate region, so that the sample injection region can completely cover the enzyme region when the enzyme region and the sample injection region are overlapped, attached and contacted; when the enzyme region and the substrate region are superposed, the substrate region can completely cover the enzyme region.
The sample inlet area can be additionally provided with a sample filtering function according to experimental needs, the sample inlet area is connected with the sample filtering area through a channel, and the online filtering function of solid impurities is realized by utilizing the technologies such as a microfluidic paper chip and the like.
The device can cover the surface of the prepared speed measuring card with a protective film according to the storage requirement, and the protective film is removed when the device is used.
The invention also provides application of the enzyme inhibition rapid detection card suitable for detecting the pesticide residue in the organic solvent extraction sample in detecting the organophosphorus and carbamate pesticide residue in the organic solvent extraction sample, and specifically the application method comprises the following steps:
(1) dripping an organic phase extracting solution of a sample to be detected in a sample injection area, placing the sample injection area at room temperature, and observing that the sample injection area is completely dried after an organic solvent is completely volatilized, wherein the time is about 3-10 minutes;
(2) dripping phosphate buffer solution into the enzyme area to keep the enzyme area wet; overlapping and attaching the sample introduction card and the enzyme region card in a folding or aligning mode, completely attaching and contacting the sample introduction region and the enzyme region to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(3) after the incubation is completed, separating the sample introduction card from the enzyme region card, removing the contact between the sample introduction region and the enzyme region, overlapping and attaching the enzyme region card and the substrate card in a folding or aligning way and the like, completely attaching and contacting the enzyme region and the substrate region to promote enzyme color development reaction, reacting for 3min at 37 ℃, then separating the enzyme region card from the substrate card, and observing the enzyme region to obtain a sample color development image; according to the experiment requirement, a proper amount of phosphate buffer solution is dripped into the substrate area to keep the color reaction in a wet state;
(4) taking phosphate buffer solution as a control solution, carrying out simultaneous detection with the sample according to the same steps of the steps (1), (2) and (3), and observing an enzyme area to obtain a control detection graph;
(5) comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues. The color development result can also be acquired by a camera, a scanner and other equipment, and data is processed by utilizing software such as images, so that the pesticide residue level in the sample can be quantitatively analyzed.
The phosphate buffer is typically 100mM phosphate buffer at pH 8.0.
Further, as mentioned above, the sample introduction card, the enzyme region card and the substrate card may be connected with each other or with each other, and when the enzyme region card and the substrate card are connected with each other, they are the conventional enzyme inhibition rapid test card, and they can be purchased in the market.
Therefore, the invention also provides an enzyme inhibition rapid test card suitable for detecting pesticide residues in organic solvent extraction samples, which consists of a sample introduction card and a conventional enzyme inhibition rapid test card, wherein the sample introduction card is provided with a sample introduction area and an ultra-inert edge surrounding and completely surrounding the sample introduction area; the conventional enzyme inhibition rapid test card is formed by connecting an enzyme area card and a substrate card, wherein the enzyme area card is provided with an enzyme area and a hydrophobic edge which completely surrounds the enzyme area; a substrate area and a hydrophobic edge surrounding the substrate area completely are disposed on the substrate card. The conventional enzyme inhibition speed test card can be purchased in the market.
In the enzyme inhibition rapid test card combination system consisting of the sample card and the conventional commercially available enzyme inhibition rapid test card, the sample card can also be independently used as an independent sample card product matched with the conventional enzyme inhibition rapid test card for rapid test of pesticide residue enzyme inhibition method of organic sample solution. The significance of the separate sample introduction card is that the cost for repeatedly purchasing conventional enzyme inhibition speed-measuring cards can be saved for users who already purchase the conventional enzyme inhibition speed-measuring cards on the market. However, in practical use, the independent sample introduction card still needs to be matched with a conventional enzyme inhibition rapid test card for use, and the independent sample introduction card cannot be used independently. The independent sample introduction card and the conventional enzyme inhibition rapid test card can be used together to realize direct rapid test of sample solution extracted by organic solvent, which can not be realized by the conventional enzyme inhibition rapid test card.
The invention also provides a technical scheme that:
an independent sample introduction card used with a conventional enzyme inhibition rapid test card for detecting pesticide residues of organic solvent extraction samples is characterized in that a sample introduction area and an ultra-inert edge surrounding the sample introduction area completely are arranged on the independent sample introduction card, and test paper or other porous sheet or plate-shaped materials capable of tolerating organic solvents are used as base materials for the sample introduction area.
The independent sample introduction card is matched with a conventional enzyme inhibition rapid test card for use to form an enzyme inhibition rapid test card system for extracting pesticide residues of a sample by using an organic solvent, the conventional enzyme inhibition rapid test card is formed by connecting an enzyme area card and a substrate card, and an enzyme area and a hydrophobic edge which completely surrounds the enzyme area are arranged on the enzyme area card; a substrate area and a hydrophobic edge surrounding the substrate area completely are disposed on the substrate card. The conventional enzyme inhibition speed test card can be purchased in the market.
On the independent sample feeding card, the super-inert edge which completely surrounds the sample feeding area is surrounded, the super-inert refers to hydrophobic ester, and the sample feeding of the card is organic phase solution, so the edge boundary of the sample feeding area must not be hydrophilic and cannot be infiltrated by organic solvent, namely the hydrophobic ester, the edge boundary is generally made of fluorinated materials, and the fluorinated materials are not hydrophilic and lipophilic and can be better used as the sample feeding range boundary of organic samples. The fluorinated material may be a commonly used one such as teflon or the like.
The shape and the area of the independent sample introduction card have no specific requirements and can be set according to the requirements of the conventional enzyme inhibition rapid test card product required to be matched; the principle of the shape design of the sample card is that the sample area and the enzyme area of the rapid-measuring card can be aligned and jointed effectively.
The shape and the size of the sample loading area on the independent sample introduction card are set according to the shape and the size of an enzyme area of an enzyme inhibition rapid-determination card product which needs to be matched with the sample loading area, and the shape of the sample loading area is preferably circular according to the shape of a rapid-determination card on the market; furthermore, for convenience of operation, the diameter of the sample injection region is preferably larger than or equal to that of the enzyme region, so as to ensure that the enzyme region can be completely covered by the sample injection region when the enzyme region and the sample injection region are overlapped, attached and contacted;
the sample inlet area can be additionally provided with a sample filtering function according to the experimental requirement, the sample inlet area is connected with the sample filtering area through a micro-fluidic chip processing technology preparation channel, a sample solution flows through the sample filtering area to reach the sample inlet area, and the online filtering function of solid impurities is realized in the process; further, the sample filtration section may be previously fixed with an adsorbent for adsorbing and removing a specific impurity component in the sample solution. The technology required for preparing the microfluidic channel and the filtering area is known technology, wax-spraying printing and Teflon solution selective coating method are preferred for preparation, and the specific processing method does not influence the use effect.
In addition, the back of the sample injection area of the independent sample injection card can be fixed with an adsorbent in a physical adsorption mode and the like, and the adsorbent is used for online purification and filtration of specific impurity components in the sample. When in use, the sample solution is directly dripped on the surface of the adsorbent on the back of the sample introduction area, and the sample introduction area on the front of the sample introduction card is directly attached to the enzyme area of the conventional enzyme inhibition rapid test card during detection.
According to the preservation requirement, the surface of the prepared independent sample introduction card can be covered with a protective film, and the protective film is removed when the card is used.
The invention also provides a using method of the independent sample introduction card which is matched with the conventional enzyme inhibition rapid test card and used for detecting the pesticide residue of the organic solvent extraction sample. The using method is substantially the same as that of the enzyme inhibition rapid test card suitable for detecting the pesticide residue in the organic solvent extraction sample. Specifically, the method comprises the following steps:
(1) dripping an organic phase extracting solution of a sample to be detected in a sample introduction area of an independent sample introduction card, placing the card at room temperature, and observing that the sample introduction area is completely dried after an organic solvent is completely volatilized, wherein the time is about 3-10 minutes;
(2) adding phosphate buffer solution with pH 8.0 and 100mM to the enzyme area of a conventional enzyme inhibition rapid test card purchased in advance, and keeping the enzyme area wet;
(3) completely covering the sample introduction area of the independent sample introduction card on the enzyme area of the rapid-determination card in an alignment and lamination manner, enabling the sample introduction area to be completely laminated and contacted with the enzyme area so as to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(4) after the incubation is completed, the independent sample introduction card and the enzyme area card are separated, the contact between the sample introduction area and the enzyme area is removed, the enzyme area card and the substrate card are overlapped and attached according to the instruction conditions of the enzyme inhibition rapid test card, and the enzymatic reaction and result judgment are carried out.
Further, in the step (4), the enzymatic reaction and the determination of the result may be performed as follows:
completely attaching and contacting the enzyme area and the substrate area, promoting enzyme color development reaction, reacting for 3min at 37 ℃, separating the enzyme area card from the substrate card, and observing the enzyme area to obtain a sample color development image; according to the experiment requirement, a proper amount of phosphate buffer solution is dripped into the substrate area to keep the color reaction in a wet state;
using a phosphate buffer solution of 100mM and pH 8.0 as a control solution, performing detection simultaneously with the sample in the same manner as in steps (1), (2), (3), and (4), and observing the enzyme region to obtain a control detection map;
comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues. The color development result can also be acquired by a camera, a scanner and other equipment, and data is processed by utilizing software such as images, so that the pesticide residue level in the sample can be quantitatively analyzed.
The invention has the following beneficial effects:
(1) the enzyme inhibition rapid test card for detecting pesticide residue provided by the invention breaks through the configuration of the conventional enzyme inhibition rapid test card and the sample adding mode, and the sample area card is newly increased in the rapid test card system, which is not existed in the configuration and the using method of the conventional enzyme inhibition rapid test card. In the prior art, a conventional enzyme inhibition rapid test card only contains an enzyme area and a substrate area, and a sample is directly added into the enzyme area, so that a sample solution extracted by an organic solvent cannot be directly detected. The water phase extraction sample has poor extraction efficiency and cannot ensure complete extraction; if the organic phase is used for extracting a sample and the sample is directly added into an enzyme area for detection, the detection result is inaccurate, and a false positive result appears.
The invention creatively adds the sample introduction card, utilizes the overlarge specific surface area of a paper material, can accelerate the online volatilization of an organic solvent in a sample introduction area after sample introduction, sequentially develops enzyme inhibition and color development reaction in a folding, laminating and other modes after the organic solvent is volatilized, and meets the dual requirements of the combination of organic phase pretreatment and an enzyme inhibition method and the zero contact of the organic solvent and an immobilized enzyme of a sample, thereby improving the detection sensitivity and the accuracy of the enzyme inhibition rapid-determination card.
(2) The invention also provides an independent sample introduction card which can be independently used, and the independent sample introduction card is matched with the conventional enzyme inhibition rapid test card for use, so that a user who purchases the conventional enzyme inhibition rapid test card can directly purchase and configure the independent sample introduction card, the cost of repeatedly purchasing an enzyme area card and a substrate card is avoided, the advantages of the independent sample introduction card are as described in the specification, the pretreatment effect of a sample is obviously improved, and meanwhile, the additional damage and infection of an extraction solvent to the enzyme activity on the rapid test card are avoided, so that the effect of integrally improving the sensitivity and the accuracy of the rapid test method is achieved.
The invention has reasonable design, clear subareas, flexible and simple structure, low manufacturing difficulty and low cost, and is suitable for large-scale mass production; the kit has the advantages of convenient use, simple and convenient operation, short detection time, low cost, easy popularization and use, and suitability for high-sensitivity, quick and accurate screening of large-batch samples in the food circulation process.
Drawings
FIG. 1 is a schematic strip structure of the enzyme inhibition rapid test card of the present invention, wherein A is a schematic structure and B is a schematic material diagram in FIG. 1.
FIG. 2 is a schematic diagram showing the L-shaped structure of the enzyme inhibition rapid test card of the present invention, wherein A and B in FIG. 2 are two connection modes.
FIG. 3 is a schematic diagram of a stand-alone card-like structure of the enzyme inhibition speed measurement card of the present invention.
FIG. 4 is a flow chart of the use of the enzyme inhibition tach of the present invention.
FIG. 5 is a graph showing the effect of the detection of the conventional enzyme inhibition rapid test card (A, C, E) and the present invention (B, D, F) on an aqueous phase solution and an organic phase solution.
FIG. 6 is a schematic diagram of the structure of the independent sample card of the present invention.
FIG. 7 is a flow chart of the use of the independent sample card of the present invention in combination with a conventional enzyme inhibition rapid test card.
FIG. 8 is a graph showing the effect of the enzyme inhibition card (D, E, F) on the detection of aqueous and organic phase solutions, when a conventional enzyme inhibition card (A, B, C) is used in combination with a separate sample card.
Detailed Description
The enzyme inhibition rapid test card and the independent sample card of the present invention are further described with reference to the following specific examples and the accompanying drawings, but the scope of the present invention is not limited thereto.
Example 1 enzyme inhibition Rapid test card with independent sample introduction region
As shown in figures 1, 2 and 3, the enzyme inhibition rapid test card with independent sample introduction regions consists of a sample introduction card 1, an enzyme region card 2 and a substrate card 3, wherein the sample introduction card 1 is provided with a hydrophilic sample introduction region 1-1 and an ultra-inert edge 1-2 which completely surrounds the sample introduction region 1-1, and the enzyme region card 2 is provided with an enzyme region 2-1 and a hydrophobic edge 2-2 which completely surrounds the enzyme region 2-1; the substrate card 3 is provided with a substrate area 3-1 and a hydrophobic rim 3-2 surrounding the substrate area 3-1 completely.
The sample introduction region 1-1, the enzyme region 2-1 and the substrate region 3-1 can use test paper, cardboard or cardboard as a base material, preferably a test paper base is adopted, wherein the enzyme region 2-1 fixes an enzyme reagent on the test paper base, and the substrate region 3-1 fixes a substrate on the test paper base; the enzymatic reagent is typically cholinesterase; the substrate is typically indophenol acetate or indole acetate.
The hydrophobic edge can be formed by known techniques such as wax-jet printing or by methods such as area-selective plastic film covering; the super-inert edge is formed by hydrophobic ester material, and fluorinated material such as Teflon can be used.
The sample introduction card 1, the enzyme region card 2 and the substrate card 3 can be connected with each other or not connected with each other or connected with any two of them, and when the sample introduction card 1, the enzyme region card 2 and the substrate card 3 are not connected with each other, the three cards are three independent cards, as shown in fig. 3.
When the enzyme area card 2 and the substrate card 3 are connected, the enzyme area card is an existing conventional enzyme inhibition rapid test card, and can be purchased and obtained on the market, as shown in the right diagram of the A diagram of fig. 7, at this time, the sample introduction card 1 can be used as an independent card (the left diagram of the A diagram of fig. 7) to be matched with the conventional enzyme inhibition rapid test card for use.
Preferably, the sample introduction card 1, the enzyme region card 2 and the substrate card 3 are connected together to form a whole, so that the use is convenient.
Furthermore, the sample introduction card 1, the enzyme region card 2 and the substrate card 3 can be connected together in various ways, and the sample introduction region 1-1 and the enzyme region 2-1, and the enzyme region 2-1 and the substrate region 3-1 can be completely aligned and overlapped in folding, aligning and other ways to be comprehensively and effectively attached and contacted; preferably, the enzyme zone card 2 is placed between the sample card 1 and the substrate card 3.
Furthermore, it is preferable that the sample card 1, the enzyme region card 2, and the substrate card 3 are sequentially connected in a strip shape. FIG. 1 is a schematic view of the strip-shaped structure of the enzyme inhibition rapid test card of the present invention,
or the sample introduction card 1, the enzyme region card 2 and the substrate card 3 are connected into an L shape, preferably, the enzyme region card 3 is arranged at the corner of the L, and the sample introduction card 1 and the substrate card 2 are respectively arranged at two adjacent edges of the L. In fig. 2, a and B are two L-shaped connecting modes.
The enzyme area and the substrate area can be respectively fixed on the gasket by the enzyme reagent and the substrate reagent and then connected with the test paper substrate, or can be directly fixed by separating corresponding areas on the test paper substrate. The immobilization of both enzymatic and substrate reagents is well known to those skilled in the art.
Preferably, the sample introduction region 1-1, the enzyme region 2-1 and the substrate region 3-1 are circular, and further preferably, the area of the enzyme region 2-1 is smaller than or equal to that of the sample introduction region 1-1, and the area of the enzyme region 2-1 is smaller than or equal to that of the substrate region 3-1, so as to ensure that the sample introduction region 1-1 can completely cover the enzyme region 2-1 when the enzyme region 2-1 and the sample introduction region 1-1 are in contact; when the enzyme region 2-1 and the substrate region 3-1 are bonded, the substrate region 3-1 can completely cover the enzyme region 2-1.
The sample inlet area can be additionally provided with a sample filtering function according to experimental needs, the sample inlet area is connected with the sample filtering area through a channel, and the online filtering function of solid impurities is realized by utilizing the technologies such as a microfluidic paper chip and the like.
Example 2
The enzyme inhibition rapid test card with the independent sample injection region can be used for detecting organophosphorus and carbamate pesticide components in an organic phase solution, and the specific operation is as follows:
(1) referring to a diagram A in FIG. 4, dropping an organic phase extracting solution of a sample to be detected in a sample injection region, placing the sample injection region at room temperature, and observing that the sample injection region is completely dried after an organic solvent is completely volatilized, wherein the time is about 3-10 minutes;
(2) referring to panel B of fig. 4, 100mM phosphate buffer at pH 8.0 was added dropwise to the enzyme region to keep the enzyme region wet; overlapping and attaching the sample introduction card and the enzyme region card in a folding or aligning mode, completely attaching and contacting the sample introduction region and the enzyme region to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(3) referring to the graph C in FIG. 4, after incubation is completed, separating the sample introduction card from the enzyme region card, removing the contact between the sample introduction region and the enzyme region, overlapping and attaching the enzyme region card and the substrate card in a folding or aligning manner, so that the enzyme region and the substrate region are completely attached and contacted to promote enzyme color development reaction, reacting for 3min at 37 ℃, then separating the enzyme region card from the substrate card, and observing the enzyme region to obtain a sample color development image; according to the experiment requirement, a proper amount of phosphate buffer solution is dripped into the substrate area to keep the color reaction in a wet state;
(4) using a phosphate buffer solution of 100mM and pH 8.0 as a control solution, performing detection simultaneously with the sample in the same manner as in steps (1), (2), and (3), and observing the enzyme region to obtain a control map;
(5) comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues. The color development result can also be acquired by a camera, a scanner and other equipment, and data is processed by utilizing software such as images, so that the pesticide residue level in the sample can be quantitatively analyzed.
The conventional enzyme inhibition rapid test card and the enzyme inhibition rapid test card are compared and tested, and the test method comprises the following steps:
conventional enzyme inhibition tacheometers contain only an enzyme region and a substrate region and are tested as follows:
40 mu L of phosphate buffer (100mM, pH:8.0) and a solution containing 0.05mg/L carbofuran in methanol are respectively dripped into an enzyme area of 3 conventional enzyme inhibition rapid test cards, the enzyme area and a substrate area are completely overlapped, attached and contacted in a folding mode for enzyme color development reaction at 37 ℃ for 3min, the enzyme area and the substrate area are separated, the color of the enzyme area is observed, the results after the color development reaction are respectively shown as A, C, E in figure 5, a graph A is phosphate buffer, namely a control product, which is negative, and a graph C and a graph E are respectively the solution containing 0.05mg/L carbofuran in methanol. Compared with the A picture, C, E is lightened in color, and the detection result shows positive, which indicates that the conventional enzyme inhibition rapid detection card is easily interfered by organic solvent, the pesticide concentration level in the organic phase solution cannot be effectively distinguished, false positive result is easily generated in the detection process, and the accuracy of the detection result is difficult to guarantee.
The detection method of the novel enzyme inhibition rapid test card comprises the following steps:
dripping 40 μ L of phosphate buffer (100mM, pH 8.0) and 0.05mg/L carbofuran in methanol into the sample injection region of 3 sheets of the novel enzyme inhibition rapid-determination card, placing at room temperature, dripping 40 μ L of phosphate buffer (100mM, pH 8.0) into the enzyme region after water and organic solvent are completely volatilized, and keeping the enzyme region wet; the method comprises the steps of completely attaching and contacting a sample introduction area and an enzyme area in a folding mode to promote enzyme inhibition reaction, incubating for 10min at 37 ℃, removing the contact between the sample introduction area and the enzyme area after the incubation is completed, completely overlapping and attaching and contacting the enzyme area and a substrate area in a folding mode to promote enzyme chromogenic reaction, reacting for 3min at 37 ℃, separating the enzyme area and the substrate area, observing the color of the enzyme area, wherein the result after the chromogenic reaction is respectively shown as B, D, F in figure 5, a figure B is phosphate buffer, namely a reference substance, and is negative, and a figure D and a figure F are respectively methanol and a solution containing 0.05mg/L of carbofuran in the methanol. Compared with a control product of a picture B, a picture D is unchanged and is negative, a picture F is obviously lightened in color, and the result shows that the detection result of the rapid test card on the aqueous phase solution and the organic phase solution which do not contain pesticide is negative, and the detection result on 0.05mg/L carbofuran in methanol is positive, so that the rapid test card is not limited by the property of the detection solution, can effectively screen the pesticide residue in the solution while being compatible with the organic phase solution to be detected, and provides a good way for improving the detection sensitivity and the detection result accuracy of the enzyme inhibition rapid test card.
EXAMPLE 3 independent sample card used with conventional enzyme inhibition Rapid test card
The sample card 1 in example 1 can be used as an independent sample card, as an independent product, and used with a conventional enzyme inhibition rapid test card (composed of an enzyme region card 2 and a substrate card 3). Because the conventional enzyme inhibition rapid test card is a mature commodity on the market, the invention provides the independent sample introduction card which can be used by users who purchase the conventional enzyme inhibition rapid test card, and avoids the cost of repeatedly purchasing the enzyme area card 2 and the substrate card 3.
As shown in FIG. 6, the independent sample card 1 of the present invention is provided with a sample introduction area 1-1 and an ultra-inert edge 1-2 surrounding the sample introduction area 1-1 completely.
The sample introduction zone may be a test paper, a thick paper sheet or a cardboard as a base material.
The preparation of the ultra-inert rim can be carried out using different known methods, and the invention provides the following preparation examples:
(1) designing the shape and size of a sample injection area on a computer in advance, and drawing a corresponding design graph; preferably, the shape of the sample injection area is circular; further, the diameter of the sample injection area is preferably larger than or equal to the diameter of the enzyme tablet of the matched rapid test card, so that the sample injection area can completely cover the enzyme tablet when the sample injection area is attached to the enzyme tablet in use.
(2) Printing the pattern of the sample injection area drawn in the step (1) on a paper substrate by using a wax-spraying printer;
(3) and (4) putting the printed graph into a 110 ℃ oven, taking out after about 10min, and cooling in the air. At this point, it is found that the printed wax line pattern has completely penetrated the paper substrate.
(4) And (3) dripping water solution into the sample injection region by adopting a region selective protection method to protect the sample injection region, completely covering and modifying the unprotected region by using a Teflon solution, integrally cleaning the sample injection card by using purified water, and drying to obtain the sample injection card with the edge of the super-inert sample injection region.
Further, physical adsorption and other modes can be used, an adsorbent is fixed on the back surface of the sample injection region prepared in the step (4) and is used for online purification and filtration of specific impurity components in the sample, an example is shown in an independent sample injection card 1B in fig. 6, wherein 1B is the front surface of the sample injection card, and the front surface is directly attached to an enzyme sheet when in use; 1B2 is the back of the sample card, the adsorbent material is fixed in the area, and when in use, the sample solution is directly dripped on the surface of the adsorbent.
Further, more complex sample processing functions may be integrated on the sample card by means of wax printing or the like using known techniques such as microfluidic paper chip processing, for example as shown in areas 1-3 of 1C in fig. 6.
Example 4 Rapid detection of pesticide residue Using independent sample card and conventional enzyme inhibition Rapid test card
The independent sample introduction card of the embodiment 3 is matched with a conventional enzyme inhibition rapid test card to be used for rapid test of pesticide residues in a sample solution extracted by an organic solvent by an enzyme inhibition method, and organophosphorus and carbamate pesticide residues in the sample extracting solution are detected.
The conventional enzyme inhibition rapid test card is composed of an enzyme region card 2 and a substrate card 3 which are connected, as shown in the right diagram of A in figure 7. This is a commercially available product. The conventional enzyme inhibition tacheometry card used in the examples of the present invention was purchased from Guangzhou oasis Biotechnology Ltd. The enzyme area card 2 is provided with an enzyme area 2-1 and a hydrophobic edge 2-2 which completely surrounds the enzyme area; the substrate card 3 is provided with a substrate area 3-1 and a hydrophobic rim 3-2 surrounding the substrate area completely.
The independent sample introduction card is matched with a conventional enzyme inhibition rapid test card for use, and the specific operation for detecting organophosphorus and carbamate pesticide residues in the organic extracting solution of the sample to be detected is as follows:
(1) referring to a diagram A in FIG. 7, dropping an organic phase extracting solution of a sample to be detected in a sample injection area of an independent sample injection card, placing the card at room temperature, and observing that the sample injection area is completely dried generally after an organic solvent is completely volatilized, wherein the time is about 3-10 minutes;
(2) referring to panel B of fig. 7, a phosphate buffer solution of 100mM and pH 8.0 was added dropwise to the enzyme region of the conventional enzyme inhibition rapid test card to keep the enzyme region wet; overlapping and attaching the independent sample introduction card and the enzyme region card in an alignment mode, completely attaching and contacting the sample introduction region and the enzyme region to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(3) referring to the graph C in FIG. 7, after incubation, the independent sample introduction card and the enzyme region card are separated, the contact between the sample introduction region and the enzyme region is released, the enzyme region card and the substrate card are overlapped and attached according to the instruction conditions of the conventional enzyme inhibition rapid-determination card, and enzymatic reaction and result judgment are carried out.
Specifically, the method comprises the following steps: overlapping and attaching the enzyme sheet and the substrate card in a folding alignment mode and the like, enabling the enzyme area and the substrate area to be completely attached and contacted to promote enzyme color development reaction, reacting for 3min at 37 ℃, then separating the enzyme area card from the substrate card, and observing the enzyme area to obtain a sample color development image; according to the experiment requirement, a proper amount of phosphate buffer solution is dripped into the substrate area to keep the color reaction in a wet state;
taking a blank organic solvent used for extracting the sample as a control solution, carrying out simultaneous detection with the sample according to the same steps of the steps (1), (2) and (3), and observing an enzyme area to obtain a control detection graph;
comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues. The color development result can also be acquired by a camera, a scanner and other equipment, and data is processed by utilizing software such as images, so that the pesticide residue level in the sample can be quantitatively analyzed.
When the sample card integrated with the purification function is used, only in the first step of sample injection, a sample is directly dripped into the adsorbent area, and the sample flows through the adsorbent area and finally reaches the sample injection area; after the solvent in the sample injection area is volatilized, the sample injection area is attached to the enzyme area which is wetted in advance, and the subsequent operation steps are the same as the steps (2) and (3).
The conventional enzyme inhibition rapid test card and the sample injection card are compared and tested with the conventional enzyme inhibition rapid test card, and the test method comprises the following steps:
conventional enzyme inhibition tacheometers contain only an enzyme region and a substrate region and are tested as follows:
40 mu L of phosphate buffer (100mM, pH:8.0), pure methanol and methanol solution containing 0.05mg/L carbofuran are respectively dripped into an enzyme plate area of 3 conventional enzyme inhibition rapid test cards, the mixture is incubated for 10min at 37 ℃, then the enzyme area and a substrate area are completely overlapped, attached and contacted in a folding mode to carry out enzyme color development reaction, the reaction is carried out for 3min at 37 ℃, the enzyme plate and the substrate area are separated, the color of the enzyme area is observed, the result after the color development reaction is respectively shown as A, B, C in figure 8, a picture is phosphate buffer, namely a reference substance, which is negative, and B pictures and C pictures are respectively methanol and solution containing 0.05mg/L carbofuran in the methanol. Compared with the A picture, B, C is light in color, and the detection result shows positive, which indicates that the conventional enzyme inhibition rapid-determination card is easily interfered by organic solvent, the pesticide concentration level in organic phase solution can not be effectively distinguished, false positive result is easily generated in the detection process, and the accuracy of the detection result is difficult to guarantee.
The invention develops an independent sample introduction card to be matched with a conventional enzyme inhibition rapid test card for use, and the test method comprises the following steps:
dripping 40 μ L of phosphate buffer (100mM, pH 8.0), pure methanol, and methanol solution containing 0.05mg/L carbofuran into the sample injection region of 3 sample cards, standing at room temperature, dripping 40 μ L of phosphate buffer (100mM, pH 8.0) into the enzyme region after water and organic solvent are completely volatilized, and keeping the enzyme region wet; aligning the sample injection area and the enzyme area to make the sample injection area and the enzyme area completely contact with each other to promote enzyme inhibition reaction, incubating for 10min at 37 ℃, removing the contact between the sample injection area and the enzyme area after the incubation is completed, completely overlapping and contacting the enzyme area and the substrate area in a folding mode to promote enzyme chromogenic reaction, reacting for 3min at 37 ℃, separating the enzyme area and the substrate area, observing the color of the enzyme area, wherein the result after the chromogenic reaction is respectively shown as D, E, F in figure 8, a D picture is phosphate buffer solution, namely a control product, and is negative, and an E picture and an F picture are respectively methanol and solution containing 0.05mg/L carbofuran in the methanol. Compared with a D picture reference substance, the E picture is unchanged and negative, the F picture is obviously lightened in color, and the result shows that when the quick test card is matched with an independent sample introduction card for use, the quick test card is negative in the detection results of the water phase solution and the organic phase solution which do not contain pesticide, and positive in the detection result of 0.05mg/L carbofuran in methanol, so that the use of the sample introduction card is not limited by the properties of the detection solution, the pesticide residue in the solution can be effectively screened while the organic phase solution to be detected is compatible, and a good way is provided for improving the detection sensitivity and the detection result accuracy of the enzyme inhibition quick test card.
Claims (16)
1. An enzyme inhibition rapid test card suitable for detecting pesticide residues in organic solvent extraction samples is composed of a sample introduction card, an enzyme region card and a substrate card, wherein a sample introduction region and a super-inert edge surrounding and completely surrounding the sample introduction region are arranged on the sample introduction card, and an enzyme region and a hydrophobic edge surrounding and completely surrounding the enzyme region are arranged on the enzyme region card; a substrate area and a hydrophobic edge surrounding the substrate area completely are disposed on the substrate card.
2. The enzyme inhibition rapid test card according to claim 1, wherein the sample introduction region, the enzyme region, and the substrate region are formed by using a test paper or other porous sheet or plate-like material capable of withstanding organic solvents as a substrate material, wherein the enzyme region fixes the enzyme reagent on the test paper substrate, and the substrate region fixes the color developing substrate on the test paper substrate.
3. The enzyme inhibition rapid test card according to claim 1, wherein the sample card, the enzyme region card and the substrate card are connected with each other or are not connected with each other or are connected with any two of them.
4. The enzyme inhibition rapid test card according to claim 3, wherein the sample card, the enzyme region card and the substrate card are connected together into a whole; when the card is integrated, the sample feeding area and the enzyme area, and the enzyme area and the substrate area on the card can be completely overlapped through folding and aligning.
5. The enzyme inhibition rapid test card according to claim 4, wherein the sample card, the enzyme region card and the substrate card are linearly arranged in sequence and connected into a long strip shape.
6. The enzyme inhibition rapid test card according to claim 4, wherein the connection is L-shaped, the enzyme region card is located at the corner of L, and the sample introduction card and the substrate card are respectively located at two adjacent edges of L.
7. The enzyme inhibition rapid test card according to claim 1, wherein said sample region, enzyme region and substrate region are circular in shape; the area of the enzyme region is smaller than or equal to that of the sample introduction region, and the area of the enzyme region is smaller than or equal to that of the substrate region.
8. The enzyme inhibition rapid test card according to claim 3, wherein the enzyme inhibition rapid test card is composed of a sample introduction card and a conventional enzyme inhibition rapid test card, the conventional enzyme inhibition rapid test card is composed of an enzyme region card and a substrate card which are connected, a sample introduction region and a super-inert edge surrounding and completely surrounding the sample introduction region are arranged on the sample introduction card; the enzyme area card is provided with an enzyme area and a hydrophobic edge which completely surrounds the enzyme area; a substrate area and a hydrophobic edge surrounding the substrate area completely are disposed on the substrate card.
9. Use of the enzyme inhibition rapid test card according to any one of claims 1 to 8 for detecting organophosphorus and carbamate pesticide residues in an organic solvent extraction sample.
10. The application according to claim 9, characterized in that the method of application is:
(1) dripping an organic phase extracting solution of a sample to be detected in a sample introduction area, and placing at room temperature until an organic solvent is completely volatilized;
(2) dripping phosphate buffer solution into the enzyme area to keep the enzyme area wet; overlapping and attaching the sample introduction card and the enzyme region card in a folding or aligning mode, completely attaching and contacting the sample introduction region and the enzyme region to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(3) after the incubation is completed, separating the sample introduction card from the enzyme region card, removing the contact between the sample introduction region and the enzyme region, overlapping and attaching the enzyme region card and the substrate card in a folding or aligning way and the like, completely attaching and contacting the enzyme region and the substrate region to promote enzyme color development reaction, reacting for 3min at 37 ℃, then separating the enzyme region card from the substrate card, and observing the enzyme region to obtain a sample color development image;
(4) taking phosphate buffer solution as a control solution, carrying out simultaneous detection with the sample according to the same steps of the steps (1), (2) and (3), and observing an enzyme area to obtain a control detection graph;
(5) comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues; the color development result can also be acquired by equipment such as a camera, a scanner and the like, and data is processed by software such as images and the like, so that the pesticide residue level in the sample is quantitatively analyzed.
11. An independent sample introduction card used in combination with a conventional enzyme inhibition rapid test card for detecting pesticide residues in organic solvent extraction samples, wherein the conventional enzyme inhibition rapid test card is formed by connecting an enzyme region card and a substrate card, and an enzyme region and a hydrophobic edge which completely surrounds the enzyme region are arranged on the enzyme region card; providing a substrate area on said substrate card and a hydrophobic edge surrounding said substrate area completely; the card is characterized in that a sample inlet area and a super-inert edge which completely surrounds the sample inlet area are arranged on the independent sample inlet card.
12. The self-contained sample card of claim 11, wherein the sample inlet region comprises a substrate material selected from the group consisting of a test strip and other porous sheet or plate materials that are resistant to organic solvents.
13. The individual sample card of claim 12 wherein the sample entry region is circular in shape and has a diameter greater than or equal to the diameter of the enzyme region.
14. Use of the card for independent sample injection according to any one of claims 11 to 13 for detecting organophosphorus and carbamate pesticide residues in an organic solvent extraction sample.
15. The application according to claim 14, characterized in that the method of application comprises the steps of:
(1) dripping the organic phase extracting solution of the sample to be detected in a sample introduction area of the independent sample introduction card, and placing the card at room temperature until the organic solvent is completely volatilized;
(2) adding phosphate buffer solution with pH 8.0 and 100mM to the enzyme area of a conventional enzyme inhibition rapid test card purchased in advance, and keeping the enzyme area wet;
(3) completely covering the sample introduction area of the independent sample introduction card on the enzyme area of the rapid-determination card in an alignment and lamination manner, enabling the sample introduction area to be completely laminated and contacted with the enzyme area so as to promote enzyme inhibition reaction, and incubating for 10min at 37 ℃;
(4) after the incubation is completed, the independent sample introduction card and the enzyme area card are separated, the contact between the sample introduction area and the enzyme area is removed, the enzyme area card and the substrate card are overlapped and attached according to the instruction conditions of the enzyme inhibition rapid test card, and the enzymatic reaction and result judgment are carried out.
16. The use according to claim 15, wherein in step (4), the enzymatic reaction and the determination of the result are carried out by the following steps:
completely attaching and contacting the enzyme area and the substrate area, promoting enzyme color development reaction, reacting for 3min at 37 ℃, separating the enzyme area card from the substrate card, and observing the enzyme area to obtain a sample color development image; according to the experiment requirement, a proper amount of phosphate buffer solution is dripped into the substrate area to keep the color reaction in a wet state;
using a phosphate buffer solution of 100mM and pH 8.0 as a control solution, performing detection simultaneously with the sample in the same manner as in steps (1), (2), (3), and (4), and observing the enzyme region to obtain a control detection map;
comparing the sample chromogenic image with a control detection image, and qualitatively analyzing the pesticide residue level in the sample: the color development of the sample is obviously lighter than that of a contrast color development, and the detection result is judged to be positive, which indicates that the sample contains organophosphorus and carbamate pesticide residues; the color development of the sample is unchanged relative to the contrast color, the detection result is judged to be negative, and the sample does not contain organophosphorus and carbamate pesticide residues. The color development result can also be acquired by a camera, a scanner and other equipment, and data is processed by utilizing software such as images, so that the pesticide residue level in the sample can be quantitatively analyzed.
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| CN2019113491659 | 2019-12-24 | ||
| CN201911349165.9A CN111239115A (en) | 2019-12-24 | 2019-12-24 | Enzyme inhibition rapid test card suitable for detecting organophosphorus and carbamate pesticide residues in organic phase and using method thereof |
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| CN202010505846.6A Pending CN111721761A (en) | 2019-12-24 | 2020-06-05 | Enzyme inhibition rapid test card and independent sample introduction card suitable for detecting pesticide residue in organic solvent extraction sample and using method thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115453036A (en) * | 2022-09-29 | 2022-12-09 | 中国农业科学院农业质量标准与检测技术研究所 | Method for rapidly detecting pesticide by combining thin-layer chromatography and enzyme inhibition principle |
| CN116550402A (en) * | 2023-07-03 | 2023-08-08 | 中国农业大学 | 3D paper-based microfluidic device and method for rapidly detecting malathion |
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2020
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- 2020-06-05 CN CN202010505846.6A patent/CN111721761A/en active Pending
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115453036A (en) * | 2022-09-29 | 2022-12-09 | 中国农业科学院农业质量标准与检测技术研究所 | Method for rapidly detecting pesticide by combining thin-layer chromatography and enzyme inhibition principle |
| CN116550402A (en) * | 2023-07-03 | 2023-08-08 | 中国农业大学 | 3D paper-based microfluidic device and method for rapidly detecting malathion |
| CN116550402B (en) * | 2023-07-03 | 2023-09-22 | 中国农业大学 | 3D paper-based microfluidic device and method for rapidly detecting malathion |
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