Immuno-chromatographic reagent card for blood and method for preparing the same

Abstract

This invention relates to the field of medical testing and discloses a blood immunochromatographic reagent card and its preparation method. The method includes: using 80% purified water as a base solution, sequentially adding 0.404% tris(hydroxymethyl)aminomethane, 0.5-0.75% polyvinylpyrrolidone, and 0.5-0.1% serum albumin; sequentially adding 0.1-0.5% S9, 0-1% polysorbate-20, and 0.01% anti-RBC to the first base solution, stirring until completely dissolved after each addition; adjusting the pH of the solution with hydrochloric acid solution; bringing the volume to the required level with purified water and retesting the pH; uniformly treating the solution on glass fiber and soaking for 10-20 minutes; drying at 37°C to obtain the blood immunochromatographic reagent card. In this embodiment of the invention, the number of formulation components is reduced to decrease the error in the preparation operation, greatly simplifying the operation process and improving the convenience and applicability of the test.

Description

Technical Field

[0001] This invention relates to the field of medical testing, and more particularly to an immunochromatographic reagent card for blood and its preparation method. Background Technology

[0002] Immunochromatographic reagents are a rapid detection technology based on the principle of antigen-antibody specific reactions. Its core principle combines immunological reactions with chromatographic techniques, allowing analytes to specifically bind as they migrate on the chromatographic material. Color development using markers such as colloidal gold or fluorescent microspheres enables qualitative, semi-quantitative, and quantitative detection of the target analyte. Currently, immunochromatographic technology is developing towards higher sensitivity, multiplexing, easier operation, and integration with molecular diagnostics. Current products used for whole blood sample testing involve methods such as pre-treating whole blood samples, diluting whole blood samples, adding whole blood first and then diluent, and direct whole blood loading. Some of these methods have drawbacks, including the need for pre-treatment equipment, complex loading procedures, and poor whole blood sample throughput.

[0003] For immunochromatographic reagents, a key design feature is the direct addition of whole blood. Centrifugation and pretreatment of blood samples, such as separating serum or plasma, significantly increase preparation time and raise the operational threshold. Reliance on equipment makes them unsuitable for rapid on-site testing, preventing on-demand testing. Furthermore, operational errors, sample contamination, or biosafety risks may occur during sample separation and transfer. Therefore, a new technology is needed to address the technical problems of excessively long preparation times and cumbersome operations in existing immunochromatographic assays. Summary of the Invention

[0004] The main objective of this invention is to solve the technical problems of excessive preparation time and cumbersome operation in existing immunochromatographic detection methods.

[0005] The first aspect of the present invention provides an immunochromatographic reagent card for blood, comprising glass fiber and an immunochromatographic solution, wherein the immunochromatographic solution is dried and attached to the glass fiber, characterized in that the immunochromatographic solution comprises: tris(hydroxymethyl)aminomethane, polyvinylpyrrolidone, serum albumin, ethylenediamine polyoxyethylene polyoxypropylene block copolymer, polysorbate-20, anti-RBC, hydrochloric acid solution, and purified water.

[0006] A second aspect of the present invention provides a method for preparing an immunochromatographic reagent card for blood, the method comprising the steps of: Based on the total weight of the immunochromatographic reagents prepared for the target, take 80% of the mass of purified water as the base solution, and add 0.404% of the mass of tris(hydroxymethyl)aminomethane, 0.5-0.75% of the mass of polyvinylpyrrolidone, and 0.5-0.1% of the mass of serum albumin in sequence. Stir until completely dissolved after each addition to obtain the first basic solution. Add 0.1-0.5% by mass of ethylenediamine polyoxyethylene polyoxypropylene block copolymer, 0-1% by mass of polysorbate-20, and 0.01% by mass of anti-RBC to the first base solution in sequence. Stir until completely dissolved after each addition to obtain the second base solution. The pH of the second base solution was adjusted to 7.2-7.6 with hydrochloric acid solution to obtain an acid-base modified solution; The solution was prepared by adjusting the pH with purified water to the required volume and the pH was retested to be within the range of 7.2 to 7.6 to obtain the immunochromatographic reagent. The immunochromatographic reagent is uniformly applied to glass fibers and then immersed at 18-28°C for 10-20 minutes to obtain pretreated glass fibers. The pretreated glass fiber was dried at 37°C to obtain a blood immunochromatographic reagent card.

[0007] Optionally, in a first implementation of the second aspect of the present invention, after the step of uniformly coating the immunochromatographic reagent onto glass fiber and immersing it at 18-28°C for 10-20 minutes to obtain pretreated glass fiber, and before the step of drying the pretreated glass fiber at 37°C for 8-24 hours to obtain a blood immunochromatographic reagent card, the following step is further included: The pretreated glass fiber is immersed in a sodium casein solution for sealing treatment to obtain sealed pretreated glass fiber. The pretreated glass fibers that have undergone the sealing treatment are immersed in a trehalose solution for protective treatment to obtain protected pretreated glass fibers.

[0008] Optionally, in a second implementation of the second aspect of the present invention, the serum albumin includes bovine serum albumin.

[0009] Optionally, in a third implementation of the second aspect of the present invention, the anti-RBC includes: mouse-derived anti-RBC.

[0010] Optionally, in a fourth implementation of the second aspect of the present invention, after the step of drying the pretreated glass fiber at 37°C to obtain the blood immunochromatographic reagent card, the method further includes: Cut the blood immunochromatographic reagent card to the specified length and width.

[0011] Optionally, in a fifth implementation of the second aspect of the present invention, the specified length and width include: a rectangle with a width of 15 mm and a length of 300 mm.

[0012] Optionally, in a sixth implementation of the second aspect of the present invention, the step of uniformly treating the immunochromatographic reagent on the glass fiber and immersing it at 18-28°C for 10-20 minutes to obtain pretreated glass fiber includes: The immunochromatographic reagent is uniformly applied to glass fibers, and the fibers are soaked for 10-20 minutes at a temperature of 18-28°C and a relative humidity of 35-80%Rh to obtain pretreated glass fibers.

[0013] Optionally, in a seventh implementation of the second aspect of the present invention, the concentration of the hydrochloric acid solution includes 1 mol / L.

[0014] Optionally, in an eighth implementation of the second aspect of the present invention, the step of drying the pretreated glass fiber at a temperature of 37°C to obtain the blood immunochromatographic reagent card includes: The pretreated glass fiber was dried at 37°C for 8-24 hours to obtain a blood immunochromatographic reagent card.

[0015] In this embodiment of the invention, the detection preparation time is moved forward to the immunochromatographic reagent card for blood preparation. By designing the immunochromatographic reagent with the formulation of tris(hydroxymethyl)aminomethane, polyvinylpyrrolidone, serum albumin, ethylenediamine polyoxyethylene polyoxypropylene block copolymer, polysorbate-20, anti-RBC, hydrochloric acid solution, and purified water, the number of formulation components is reduced to reduce the error of the preparation operation. This greatly simplifies the operation process and improves the convenience and applicability of the detection, solving the technical problems of excessive preparation time and cumbersome operation in existing immunochromatographic detection schemes. Attached Figure Description

[0016] Figure 1 This is a schematic diagram of an embodiment of the method for preparing blood immunochromatographic reagent cards according to the present invention. Detailed Implementation

[0017] This invention provides a blood immunochromatographic reagent card and its preparation method.

[0018] The embodiments of the present invention will now be described in more detail with reference to the accompanying drawings. While some embodiments of the present invention are shown in the drawings, it should be understood that the present invention can be implemented in various forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided to provide a more thorough and complete understanding of the present disclosure. It should be understood that the accompanying drawings and embodiments are for illustrative purposes only and are not intended to limit the scope of protection of the present invention.

[0019] In the description of the embodiments disclosed in this invention, the term "comprising" and similar terms should be understood as open-ended inclusion, i.e., "including but not limited to". The term "based on" should be understood as "at least partially based on". The term "one embodiment" or "the embodiment" should be understood as "at least one embodiment". The terms "first", "second", etc., may refer to different or the same objects. Other explicit and implicit definitions may also be included below.

[0020] For ease of understanding, the specific process of the embodiments of the present invention is described below. Please refer to [link / reference]. Figure 1 A schematic diagram of an embodiment of the preparation method of the blood immunochromatographic reagent card in this invention is shown. The preparation method of the blood immunochromatographic reagent card includes: 101. Based on the total weight of the immunochromatographic reagents prepared for the target, take 80% of purified water as the base solution, and add 0.404% of tris(hydroxymethyl)aminomethane, 0.5-0.75% of polyvinylpyrrolidone, and 0.5-0.1% of serum albumin in sequence. Stir until completely dissolved after each addition to obtain the first base solution. Preferably, bovine serum albumin is used as the serum albumin.

[0021] 102. Add 0.1-0.5% by mass of ethylenediamine polyoxyethylene polyoxypropylene block copolymer, 0-1% by mass of polysorbate-20, and 0.01% by mass of anti-RBC to the first base solution in sequence. Stir until completely dissolved after each addition to obtain the second base solution. In this embodiment, ethylenediamine polyoxyethylene polyoxypropylene block copolymer and polysorbate-20 are used as surfactants to promote the flow of the sample on the glass fiber membrane. ethylenediamine polyoxyethylene polyoxypropylene block copolymer and polysorbate-20 are nonionic surfactants.

[0022] Preferably, murine anti-RBC antibodies are used. The RBC antibodies can act on red blood cells in whole blood samples, causing the red blood cells to adhere to the glass fiber membrane and not transfer to the NC membrane.

[0023] 103. Adjust the pH of the second base solution to 7.2-7.6 with hydrochloric acid solution to obtain an acid-base prepared solution; Preferably, the hydrochloric acid solution is 1 mol / L, and tris(hydroxymethyl)aminomethane and hydrochloric acid are used as a Tris-HCl buffer system to stabilize the solution environment.

[0024] 104. Adjust the solution to the required volume with purified water, and retest the pH to ensure it is within the range of 7.2 to 7.6 to obtain the immunochromatographic reagent; 105. The immunochromatographic reagent is uniformly applied to the glass fiber and soaked at 18-28°C for 10-20 minutes to obtain pretreated glass fiber; Preferably, the immunochromatographic reagent is uniformly applied to the glass fiber and then soaked for 10-20 minutes at a temperature of 18-28°C and a relative humidity of 35-80%Rh to obtain pretreated glass fiber.

[0025] Soaking the fiberglass membrane is to allow the liquid to adhere to the fiberglass membrane.

[0026] Preferably, the following specific implementation method is included between steps 105 and 106: 1051. The pretreated glass fiber is immersed in a sodium casein solution for sealing treatment to obtain sealed pretreated glass fiber; 1052. The pretreated glass fiber that has undergone the sealing treatment is immersed in a trehalose solution for protective treatment to obtain the pretreated glass fiber with protective treatment.

[0027] In steps 1051-1052, sodium casein salt is used as a sealing agent for glass fiber membranes. Through hydrophobic interactions and electrostatic adsorption, it covers non-specific protein binding sites on the surface of the glass fiber membrane, reducing the non-specific adsorption of impurities (such as albumin and immunoglobulins) in blood samples and lowering background interference.

[0028] Trehalose acts as a protective agent, maintaining the structure of the glass fiber membrane after immersion and drying. As a glass-forming agent, trehalose replaces water molecules during the drying process to maintain the membrane structure and protein conformation, preventing fiber collapse and membrane porosity decrease during drying, and protecting the stability of active ingredients such as anti-RBC.

[0029] 106. The pretreated glass fiber is dried at a temperature of 37°C to obtain a blood immunochromatographic reagent card.

[0030] In this embodiment, the pretreated glass fiber can be dried using an electric heating drying oven to obtain an immunochromatographic reagent card for blood.

[0031] Preferably, the pretreated glass fiber is dried at 37°C for 8-24 hours to obtain a blood immunochromatographic reagent card.

[0032] Furthermore, after step 106, the immunochromatographic reagent card containing the blood is cut to a specified length and width. Specifically, the specified length and width can be a rectangle with a width of 15mm and a length of 300mm.

[0033] Based on the preparation method of steps 101-106, an immunochromatographic reagent card for blood is prepared. The immunochromatographic reagent card for blood includes glass fiber and immunochromatographic solution. The immunochromatographic solution is dried and attached to the glass fiber. The immunochromatographic solution is characterized by comprising: tris(hydroxymethyl)aminomethane, polyvinylpyrrolidone, serum albumin, ethylenediamine polyoxyethylene polyoxypropylene block copolymer, polysorbate-20, anti-RBC, hydrochloric acid solution, and purified water.

[0034] Example 1 Take 80% of the mass of purified water required for the treatment solution. Add 0.404% of tris(hydroxymethyl)aminomethane, 0.5% of polyvinylpyrrolidone, and 1% of bovine serum albumin to the purified water in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0035] Add 0.5% by mass of ethylenediamine polyoxyethylene polyoxypropylene block copolymer and 0.01% of rodent-derived anti-RBC to the prepared solution in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0036] Adjust the pH of the solution to within the range of 7.4±0.2 using 1M hydrochloric acid solution, then dilute the prepared solution to the required volume using purified water, and retest the pH value to ensure it is within the range of 7.4±0.2.

[0037] The prepared treatment solution was evenly applied to the glass fiber and then soaked for 10 minutes under ambient temperature and humidity conditions (temperature 18~28℃, relative humidity 35~80%Rh).

[0038] The glass fibers treated with the treatment solution were dried at 37°C for 8-24 hours to obtain the blood immunochromatographic reagent card.

[0039] Cut the glass fiber membrane of the blood immunochromatographic reagent card to the specified length and width (15mm × 300mm).

[0040] Example 2 Take 80% of the mass of purified water required for the treatment solution. Add 0.404% of tris(hydroxymethyl)aminomethane, 0.75% of polyvinylpyrrolidone, and 1% of bovine serum albumin to the purified water in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0041] Add 0.2% S9, 0.5% polysorbate-20, and 0.01% murine anti-RBC by mass required for the treatment solution to the prepared solution in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0042] The pH of the solution was adjusted to 7.4±0.2 using 1M hydrochloric acid solution. The prepared solution was then brought to the required volume with purified water, and the pH was retested to be 7.4±0.2.

[0043] The prepared treatment solution is evenly applied to the glass fiber, and then immersed in it at room temperature and humidity (temperature 18~28℃, relative humidity 35~80%Rh) for 10 minutes. The glass fiber treated with the treatment solution is then dried at 37℃ for 8~24 hours.

[0044] Example 3 Take 80% of the mass of purified water required for the treatment solution, and add 0.404% of tris(hydroxymethyl)aminomethane, 0.75% of polyvinylpyrrolidone, and 0.5% of bovine serum albumin in the purified water in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0045] Add 0.1% S9, 1% Tween-20, and 0.01% murine anti-RBC by mass required for the treatment solution to the prepared solution in sequence. After each component is completely dissolved, add the next component and stir until completely dissolved.

[0046] The pH of the solution was adjusted to 7.4±0.2 using 1M hydrochloric acid solution. The prepared solution was then brought to the required volume with purified water, and the pH was retested to be 7.4±0.2.

[0047] The prepared treatment solution was evenly applied to the glass fiber and then soaked for 20 minutes under ambient temperature and humidity conditions (temperature 18~28℃, relative humidity 35~80%Rh).

[0048] The glass fibers treated with the treatment solution were dried at 37°C for 8-24 hours to obtain the blood immunochromatographic reagent card.

[0049] Cut the glass fiber membrane of the blood immunochromatographic reagent card to the specified length and width (15mm × 300mm).

[0050] Furthermore, although the operations are described in a specific order, this should be understood as requiring that such operations be performed in the specific order shown or in sequential order, or requiring that all illustrated operations be performed to achieve the desired result. In certain environments, multitasking and parallel processing may be advantageous. Similarly, although several specific implementation details are included in the above discussion, these should not be construed as limiting the scope of this disclosure. Certain features described in the context of individual embodiments may also be implemented in combination in a single implementation. Conversely, various features described in the context of a single implementation may also be implemented individually or in any suitable sub-combination in multiple implementations.

[0051] Although the subject matter has been described using language specific to structural features and / or methodological logic, it should be understood that the subject matter defined in the appended claims is not necessarily limited to the specific features or actions described above. Rather, the specific features and actions described above are merely illustrative examples of implementing the claims.

Claims

1. A blood immunochromatographic reagent card, comprising glass fibers and an immunochromatographic solution, wherein the immunochromatographic solution is dried and adhered to the glass fibers, characterized in that, The immunochromatographic solution comprises: tris(hydroxymethyl)aminomethane, polyvinylpyrrolidone, serum albumin, ethylenediamine polyoxyethylene polyoxypropylene block copolymer, polysorbate-20, anti-RBC, hydrochloric acid solution, and purified water.

2. A method for preparing a blood immunochromatographic reagent card, characterized in that, Including the following steps: Based on the total weight of the immunochromatographic reagents prepared for the target, take 80% of the mass of purified water as the base solution, and add 0.404% of the mass of tris(hydroxymethyl)aminomethane, 0.5-0.75% of the mass of polyvinylpyrrolidone, and 0.5-0.1% of the mass of serum albumin in sequence. Stir until completely dissolved after each addition to obtain the first basic solution. Add 0.1-0.5% by mass of ethylenediamine polyoxyethylene polyoxypropylene block copolymer, 0-1% by mass of polysorbate-20, and 0.01% by mass of anti-RBC to the first base solution in sequence. Stir until completely dissolved after each addition to obtain the second base solution. The pH of the second base solution was adjusted to 7.2-7.6 with hydrochloric acid solution to obtain an acid-base modified solution; The solution was prepared by adjusting the pH with purified water to the required volume and the pH was retested to be within the range of 7.2 to 7.6 to obtain the immunochromatographic reagent. The immunochromatographic reagent is uniformly applied to glass fibers and then immersed at 18-28°C for 10-20 minutes to obtain pretreated glass fibers. The pretreated glass fiber was dried at 37°C to obtain a blood immunochromatographic reagent card.

3. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, After the step of uniformly coating the immunochromatographic reagent onto glass fiber and immersing it at 18-28°C for 10-20 minutes to obtain pretreated glass fiber, and before the step of drying the pretreated glass fiber at 37°C for 8-24 hours to obtain the blood immunochromatographic reagent card, the following step is also included: The pretreated glass fiber is immersed in a sodium casein solution for sealing treatment to obtain sealed pretreated glass fiber. The pretreated glass fibers that have undergone the sealing treatment are immersed in a trehalose solution for protective treatment to obtain protected pretreated glass fibers.

4. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, The serum albumin includes bovine serum albumin.

5. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, The anti-RBC includes: murine anti-RBC.

6. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, After the step of drying the pretreated glass fiber at 37°C to obtain the blood immunochromatographic reagent card, the method further includes: Cut the blood immunochromatographic reagent card to the specified length and width.

7. The method for preparing blood immunochromatographic reagents according to claim 6, characterized in that, The specified length and width include: a rectangle with a width of 15mm and a length of 300mm.

8. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, The step of uniformly treating the immunochromatographic reagent onto the glass fiber and immersing it at 18-28°C for 10-20 minutes to obtain pretreated glass fiber includes: The immunochromatographic reagent is uniformly applied to glass fibers, and the fibers are soaked for 10-20 minutes at a temperature of 18-28°C and a relative humidity of 35-80%Rh to obtain pretreated glass fibers.

9. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, The concentration of the hydrochloric acid solution includes 1 mol / L.

10. The method for preparing blood immunochromatographic reagents according to claim 2, characterized in that, The step of drying the pretreated glass fiber at 37°C to obtain the blood immunochromatographic reagent card includes: The pretreated glass fiber was dried at 37°C for 8-24 hours to obtain a blood immunochromatographic reagent card.

Images