Sampling extracting solution for total IgE antibody detection product, total IgE antibody detection kit and application of total IgE antibody detection kit

Abstract

The invention discloses a sampling extracting solution for a total IgE antibody detection product, a total IgE antibody detection kit and application of the total IgE antibody detection kit, and the sampling extracting solution comprises the following components in percentage by mass: 0.5-2% of a component A, 0.5-2% of a component B and the balance of a component C, 0.3% to 1.2% of Tris; 0.3% to 1.2% of Na2HPO4; 0.3% to 1.2% of NaCl; 0.01%-0.1% of a component B; the balance of purified water; wherein the component A is Triton X100 or Tween 20, and the component B is Tween 20; and the component A is prolin 300 or NaN3, and the component B is prolin 300 or NaN3. According to the present invention, with the extraction liquid, the detected substance can be completely and efficiently extracted from the sampling test paper, and the components of the extraction liquid are matched with the reaction system of the reagent so as to improve the antigen-antibody binding efficiency to a certain extent; soft materials such as sampling filter paper are adopted to suck ocular surface liquid, so that hard materials can be prevented from directly making contact with the surfaces of eyeballs, substance exchange between other liquid and the human body is avoided, a subject can conduct sampling by himself / herself, the risk can be remarkably reduced in the sampling process, operation is easy, and professionals do not need to cooperate for operation.

Description

Technical Field

[0001] This invention relates to the field of biological detection technology, specifically to a sampling extraction solution, a total IgE antibody detection kit, and their applications for total IgE antibody detection products. Background Technology

[0002] In vitro diagnostic reagents typically involve collecting human samples (various body fluids / cells / tissues, etc.) from the subject's body using specific sampling methods, and then conducting in vitro tests.

[0003] Some commercially available total IgE antibody test kits primarily utilize the double-antibody sandwich method and colloidal gold immunochromatography to qualitatively detect total IgE antibodies in ocular surface fluid. The sample is dropped into the sample well of the test card, and due to chromatography, the sample moves forward along the nitrocellulose membrane. When the sample contains total IgE antibodies, these antibodies bind to colloidal gold-labeled anti-human IgE monoclonal antibodies during chromatography. When the concentration of total IgE antibodies in the sample is equal to or higher than the detection limit, a complex forms in the detection zone (T) and aggregates, resulting in color development. The remaining portion continues to migrate to the region immobilized with goat anti-mouse IgG, where it binds and aggregates again in the control zone (C), also resulting in color development. When the sample contains no total IgE antibodies or the antibody concentration is below the detection limit, there is little or no complex aggregation in the detection zone (T), and therefore no visible color development occurs. In this case, the test result is negative. The remaining portion continues to migrate to the region immobilized with goat anti-mouse IgG, where it binds and aggregates again in the control zone (C), resulting in color development.

[0004] Collecting ocular surface fluid samples is less convenient compared to collecting urine or feces. Currently, tear sample collection for total IgE antibody detection products mainly uses the following two methods:

[0005] 1. Directly drawing the reagent from the surface of the eyeball using a capillary tube is a high-risk method that must be performed by trained professionals, and extreme care must be taken during the procedure to avoid harming the subject's eyes. Furthermore, it is highly likely to cause eye discomfort in the subject, and for some subjects with dry eyes, the amount drawn is extremely small, severely affecting the sensitivity of the reagent.

[0006] 2. Use physiological saline to rinse the ocular surface. This method requires the subject to lie flat on a suitable device, and then a professional will instill standard physiological saline into the subject's eyes. After the subject blinks to allow the ocular surface to be fully rinsed, the subject will close their eyes tightly and squeeze out the rinsing solution, which will then be received by a professional at the corner of the eye.

[0007] Neither of the above two sampling methods can be completed independently by the subject, and both carry certain risks. Summary of the Invention

[0008] The purpose of this invention is to overcome the shortcomings of the prior art and provide a sampling extraction solution for total IgE antibody detection products, used to extract ocular surface fluid samples for testing.

[0009] To solve the above-mentioned technical problems, the objective of this invention is achieved through the following technical solution:

[0010] A sampling extract for use in a total IgE antibody detection product, comprising, by weight percentage:

[0011]

[0012] Purified water balance;

[0013] Component A is either Triton X100 or Tween 20.

[0014] Component B is either proclin 300 or NaN3.

[0015] In this invention, the sampling extraction solution is used to extract the analyte from a soft material containing ocular surface fluid to obtain an ocular surface fluid test sample. This ocular surface fluid test sample is used for the detection of total IgE antibodies. In specific applications, it can be used in conjunction with the sampling process of a total IgE antibody detection kit to extract the sampled filter paper after sampling, thereby obtaining a test sample for the detection of total IgE antibodies.

[0016] To improve elution efficiency, component A acts as a surfactant. Surfactants enhance the hydrophilicity of the sampling filter paper, thereby improving extraction efficiency. Additionally, to ensure optimal antigen-antibody binding conditions are not disrupted, and to maintain the stability of the colloidal gold, and to prevent dilution of key ions in the original reaction system by directly adding purified water, key chemicals from the original reaction system are added to the extract to maintain the original reaction system. Therefore, Tris, Na₂HPO₄, and NaCl are also added. Ocular surface fluid samples are primarily neutral or slightly alkaline. The optimal buffering range of Tris-hydrochloric acid buffer is 7.10-9.00, and the normal pH of human ocular surface fluid samples is generally 7.4-7.6. Tris-hydrochloric acid buffer effectively covers the pH range of human ocular surface fluid samples, so no further chemicals are needed. To ensure the stability of the extract, component B is also added to inhibit microbial growth and improve stability.

[0017] Therefore, the sampling extraction solution in this invention can extract the analyte from the sampling filter paper. The surfactant improves the hydrophilicity of the sampling filter paper, thereby increasing the extraction efficiency. Furthermore, the addition of chemicals such as Tris, Na2HPO4, and NaCl to the extraction solution ensures that the original reaction system of the reagents remains largely unchanged, guaranteeing optimal conditions for the binding of total IgE antigen and antibody, as well as the stability of the colloidal gold. Additionally, component B added to the sampling extraction solution of this invention inhibits microbial growth, ensuring the stability of the extraction solution.

[0018] Preferably, by weight percentage, it includes:

[0019]

[0020] The remaining amount of purified water.

[0021] Preferably, component A is Tween20.

[0022] Preferably, component B is proclin 300.

[0023] Preferably, the pH of the sampling extract is 7.0-10.0.

[0024] Preferably, the pH of the sampling extract is 8.0-8.5.

[0025] The present invention also provides a method for preparing a sampling extract for a total IgE antibody detection product as described above, characterized in that it comprises:

[0026] According to the mass percentage, add component A, Tris, Na2HPO4, NaCl and component B to purified water, adjust the pH to 7.0-10.0, and obtain the sampling extract.

[0027] This invention also provides a method for obtaining a sample for detecting total IgE antibodies in ocular surface fluid, comprising:

[0028] A soft sampling material is used to collect ocular surface fluid;

[0029] The soft material from which ocular surface fluid was collected was placed in the sampling extraction solution as described above to extract the ocular surface fluid sample to be tested.

[0030] In this invention, after using a soft material to absorb ocular surface fluid, if it is necessary to extract the ocular surface fluid sample from the soft material for subsequent sample addition and testing, a good extraction effect is required. The sampling extract solution used in this invention has the following advantages:

[0031] 1. It has good extraction effect and can elute the analyte (total IgE antibody) from soft materials (such as sampling filter paper);

[0032] 2. It conforms to the reaction system of the reagent itself, and the components in the extract will not disrupt the optimal conditions for antigen-antibody binding, nor will it disrupt the stability of colloidal gold (colloids are a metastable state);

[0033] 3. It can eliminate or reduce differences between different subject samples to a certain extent (non-test item differences), such as pH;

[0034] 4. High stability, preventing deterioration and failure within the product's shelf life.

[0035] Preferably, the method of using a sampling soft material to aspirate ocular surface fluid includes:

[0036] One end of the sampling soft material is placed inside the conjunctival sac on the outer side of the lower eyelid, while the other end hangs outside the lower eyelid, so that the end of the soft material inside the conjunctival sac can absorb ocular surface fluid.

[0037] Existing methods for sampling ocular surface fluid typically involve direct contact between a rigid object (capillary pipette) and the eyeball. This requires a high level of expertise from the operator and may result in other fluids entering the ocular surface during extraction, potentially leading to substance exchange with the body. Therefore, existing methods are risky and complex. In contrast, this invention allows for collection using a soft material (such as sampling filter paper) in contact with the ocular surface. The specific collection method is as follows:

[0038] In this invention, a soft sampling material (such as sampling filter paper) can be sandwiched inside the conjunctival sac on the outer side of the lower eyelid, such as in the outer 1 / 3 of the conjunctival sac, with the other end hanging down outside the lower eyelid. With the eyes closed and looking slightly upwards, blinking is permitted. After sufficient ocular surface fluid has been absorbed (e.g., tear film chromatography to the 20mm mark), the sampling filter paper is placed into an extraction tube containing the sampling extract. The sampling filter paper is mixed in the extract for at least 30 seconds, while simultaneously squeezing the sampling filter paper at least 5 times through the outer wall of the extraction tube to ensure the sample is fully eluted into the extraction tube.

[0039] As can be seen from the above, the sampling method using soft materials in this invention is convenient to operate and has low risk, thus effectively reducing sampling risk and operational difficulty.

[0040] The present invention also provides a total IgE antibody detection kit, comprising a total IgE antibody detection plate, a sampling soft material for collecting the sample to be tested, and the above-mentioned sample extract solution for extracting the analyte from the sampling soft material.

[0041] Preferably, the sampling soft material is sampling filter paper.

[0042] The present invention also provides the application of the total IgE antibody detection kit as described above in the detection of total IgE antibodies.

[0043] The advantages of this invention compared to the prior art are:

[0044] (1) The extraction solution of this invention has a good extraction effect, which can extract the total IgE antibody analyte from the sampling material (such as eluting from the sampling filter paper), and it conforms to the reaction system of the reagent itself. The components in the extraction solution do not disrupt the optimal conditions for antigen-antibody binding, and do not damage the stability of colloidal gold (colloids are a metastable state). Moreover, it can eliminate or reduce the differences between different subject samples (non-test item differences), such as pH, to a certain extent. In addition, the extraction solution has good stability, effectively preventing deterioration and failure within the product's shelf life. Therefore, the extraction solution of this invention is used for total IgE antibody detection. When used in conjunction with the sampling process of the total IgE antibody detection kit to extract from the sampling filter paper after sampling, the extraction solution can fully and efficiently extract the analyte from the sampling test paper, and its components match the reaction system of the reagent, which can improve the antigen-antibody binding efficiency to a certain extent.

[0045] (2) The present invention uses soft materials such as sampling filter paper to absorb ocular surface fluid, which avoids direct contact between hard materials and the ocular surface, and there is no other liquid exchange with the human body. Subjects can sample themselves, the sampling process can significantly reduce the risk, and it is easy to operate without the need for professional personnel to cooperate in the operation.

[0046] The present invention will be further described below with reference to the accompanying drawings and specific embodiments. Attached Figure Description

[0047] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the following description of the embodiments will be briefly introduced. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0048] Figure 1 This is a schematic diagram of the total IgE detection process in this embodiment. Detailed Implementation

[0049] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0050] Example 1

[0051] The preparation of the sampling extract includes:

[0052] Add 1% Tween 20, 0.6% Tris, 0.6% Na2HPO4, 0.7% NaCl, and 0.03% Proclin 300 to purified water by mass percentage, and adjust the pH of the solution to 8.0.

[0053] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0054] Example 2

[0055] The preparation of the sampling extract includes:

[0056] Add 0.6% Tween 20, 0.6% Tris, 0.6% Na2HPO4, 0.7% NaCl, and 0.03% Proclin 300 to purified water by mass percentage, and adjust the pH of the solution to 8.5.

[0057] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0058] Example 3

[0059] The preparation of the sampling extract includes:

[0060] Add 1% Triton X100, 0.7% Tris, 0.7% Na2HPO4, 0.6% NaCl, and 0.02% Proclin 300 to purified water by mass percentage, and adjust the pH of the solution to 8.5.

[0061] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0062] Example 4

[0063] The preparation of the sampling extract includes:

[0064] Add 1% Tween 20, 0.4% Tris, 0.5% Na2HPO4, 0.4% NaCl, and 0.02% Proclin 300 to purified water by mass percentage, and adjust the pH of the solution to 7.5.

[0065] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0066] Example 5

[0067] The preparation of the sampling extract includes:

[0068] Add 1% Tween20, 0.8% Tris, 0.8% Na2HPO4, 0.7% NaCl, and 0.03% NaN3 to purified water by mass percentage, and adjust the pH of the solution to 9.0.

[0069] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0070] Comparative Example 1

[0071] The preparation of the sampling extract includes:

[0072] Add 0.9% NaCl and 0.05% proclin 300 to the purified water by weight percentage, and adjust the pH to 7.0.

[0073] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0074] Comparative Example 2

[0075] The preparation of the sampling extract includes:

[0076] Add 10% Tween20 and 0.03% Proclin 300 to the purified water by weight percentage, and adjust the pH to 8.0.

[0077] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0078] Comparative Example 3

[0079] The preparation of the sampling extract includes:

[0080] Add 0.9% NaCl, 5% Triton X100, and 0.02% Proclin 300 to the purified water by mass percentage, and adjust the pH to 8.5.

[0081] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0082] Comparative Example 4

[0083] The preparation of the sampling extract includes:

[0084] Add 0.2% Tris, 5% Tween20, and 0.03% Proclin300 to the purified water by weight percentage, and adjust the pH to 9.0.

[0085] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0086] Comparative Example 5

[0087] Prepare a 0.01M PBS solution using purified water by mass percentage, then add 5% Tween 20 and 0.03% Proclin 300 to the solution and adjust the pH to 8.5.

[0088] The prepared extract was used to extract samples for testing, and a sensitivity comparison test of total IgE antibody detection reagents was conducted.

[0089] The test results of Example 1 and Comparative Examples 1 to 5 are shown in Table 1.

[0090] Table 1

[0091]

[0092] Note: "+" indicates a positive result, meaning that the concentration can be detected. The lower the detection concentration, the higher the sensitivity of the detection system. "-" indicates a negative result, meaning that the concentration cannot be detected.

[0093] The above results show that Example 1 can detect a minimum total IgE antibody standard of 2.0 IU / mL; Examples 2, 3, 4, and 5 can detect a minimum total IgE antibody standard of 3.5 IU / mL; Comparative Examples 2 and 3 can detect a minimum total IgE antibody standard of 7.5 IU / mL; and Comparative Examples 1, 4, and 5 can only detect a minimum total IgE antibody standard of 20 IU / mL. The sensitivity of all detection systems using Comparative Examples 1-5 is significantly lower than that of Examples 1-5, with Example 1 exhibiting the highest sensitivity. Therefore, the extracts prepared in Examples 1-5 of this invention can be used in total IgE antibody detection kits. When using this total IgE antibody detection kit for total IgE detection, as... Figure 1 As shown, the steps are as follows:

[0094] The ocular fluid was collected by contacting a soft sampling material (sampling filter paper) with the ocular surface.

[0095] The sampling filter paper used to absorb ocular surface fluid is placed in the extraction solution to extract the sample to be tested.

[0096] Finally, the extracted test samples are subjected to total IgE detection (added to a total IgE antibody detection plate) to obtain the total IgE detection results.

[0097] Specifically, a tear sampling filter paper is placed in the conjunctival sac at the outer 1 / 3 of the lower eyelid, with the other end hanging outside the lower eyelid. The eyes are closed, and the gaze is slightly upward, allowing for free blinking. After sufficient ocular surface fluid has been absorbed (tear chromatography to the 20mm mark), the sampling filter paper is placed in the extraction tube. The sampling filter paper should be mixed in the extraction solution for at least 30 seconds, while simultaneously squeezing the sampling filter paper at least 5 times through the outer wall of the extraction tube to ensure the sample is fully eluted into the extraction tube.

[0098] Therefore, whether the extract can fully extract / elute the analyte from the sampling filter paper will have a crucial impact on the performance (sensitivity) of the reagent.

[0099] In this embodiment, a soft material (such as sampling filter paper) is used to collect samples from the ocular surface, which effectively reduces sampling risks and operational difficulties. After the ocular surface fluid is absorbed by the soft material (sampling filter paper), the analyte is extracted using the extraction solution of this invention, facilitating subsequent sample addition and testing.

[0100] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any person skilled in the art can easily conceive of various equivalent modifications or substitutions within the technical scope disclosed in the present invention, and these modifications or substitutions should all be covered within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A sampling extraction solution for total IgE antibody detection products, characterized in that, By weight percentage, including: Purified water balance; Component A is either Triton X100 or Tween 20. Component B is either proclin 300 or NaN3.

2. The sampling and extraction solution for total IgE antibody detection products according to claim 1, characterized in that, By weight percentage, including: The remaining amount of purified water.

3. The sampling and extraction solution for total IgE antibody detection products according to claim 2, characterized in that, Component A is Tween20; component B is proclin 300.

4. The sampling and extraction solution for total IgE antibody detection products according to any one of claims 1 to 3, characterized in that, The pH of the sample extraction solution is 7.0-10.

0.

5. A method for preparing a sampling extract solution for a total IgE antibody detection product as described in any one of claims 1 to 4, characterized in that, include: According to the mass percentage, add component A, Tris, Na2HPO4, NaCl and component B to purified water, adjust the pH to 7.0-10.0, and obtain the sampling extract.

6. A method for obtaining test samples for detecting total IgE antibodies in ocular surface fluid, characterized in that, include: A soft sampling material is used to collect ocular surface fluid; The soft sampling material after absorbing ocular surface fluid is placed in the sampling extraction solution as described in any one of claims 1 to 4 to extract the ocular surface fluid sample to be tested.

7. The method for obtaining test samples for total IgE antibody detection in ocular surface fluid according to claim 6, characterized in that, The method of using a sampling soft material to aspirate ocular surface fluid includes: One end of the sampling soft material is placed inside the conjunctival sac on the outer side of the lower eyelid, while the other end hangs outside the lower eyelid, so that the end of the soft material inside the conjunctival sac can absorb ocular surface fluid.

8. A total IgE antibody detection kit, characterized in that, It includes a total IgE antibody detection plate, a soft sampling material for collecting the sample to be tested, and a sampling extractant for extracting the analyte from the soft sampling material, which is any one of claims 1 to 4.

9. The total IgE antibody detection kit as described in claim 8, characterized in that, The sampling soft material is sampling filter paper.

10. The application of a total IgE antibody detection kit as described in any one of claims 8 to 9 in the detection of total IgE.

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