Hydrogen peroxide rapid detection kit and use method

Abstract

Disclosed in the present invention are a hydrogen peroxide rapid detection kit and a use method thereof. The hydrogen peroxide rapid detection kit comprises a sample diluent and a test strip dried with a detection color developing agent and a quality control color developing agent. The sample diluent comprises 100-500 mM of ammonium oxalate, 0.3-2.0 mM of 8-hydroxyquinoline, 50-500 mM of sodium chloride and 10-100 mM of a first pH regulator with the pH of 4.5-7.0.

Description

Hydrogen peroxide rapid detection kit and usage method Technical Field

[0001] The present invention belongs to the field of biological detection technology, and in particular relates to a rapid hydrogen peroxide detection reagent and a use method. Background Art

[0002] Numerous studies have found that CAT expression is downregulated and activity is significantly reduced in various solid tumor tissues, including lung, esophageal, intestinal, liver, and cervical cancers. This leads to increased accumulation of hydrogen peroxide in cells and tissues, creating oxidative stress. This leads to oxidative damage to biomacromolecules such as DNA, proteins, and lipids, and increased genomic instability. This in turn transmits pro-tumor growth signals, a key mechanism driving tumor initiation and progression. If solid tumors rely solely on diffusion from surrounding tissues for their nutrients, their growth will be extremely slow or dormant, with tumors remaining around 1-2 mm in size. To overcome this growth constraint, tumor angiogenesis is necessary, requiring the formation of tumor blood vessels to continuously absorb nutrients from the blood supply and transport metabolic waste. The high diffusivity of hydrogen peroxide allows tumors to release hydrogen peroxide to transmit growth signals before angiogenesis occurs. Studies have demonstrated that hydrogen peroxide activates hypoxia-inducible factor-1, which in turn upregulates the expression of a series of angiogenic proteins, including vascular endothelial growth factor, guiding angiogenesis into the tumor. By oxidizing the cysteine ​​sulfhydryl groups in proteins, hydrogen peroxide can also directly or indirectly activate signal transduction proteins that promote the occurrence and development of tumors, such as EGFR, PKC, MAPK, CDK, JAK, Ras, NF-κB, c-Myc, and AP-1, and inhibit the activity of phosphatases with tumor suppressor functions such as PTEN and PTPs, making the cancer-promoting signal pathways more continuously promote the occurrence and development of tumors, and can lead to tumor cell resistance to chemotherapy drugs and targeted drugs such as EGFR.

[0003] Therefore, the detection of hydrogen peroxide in human samples can be used not only for early screening of tumors, but also for follow-up monitoring of tumor treatment and prognosis. In addition, hydrogen peroxide detection is also a common method for gynecological inflammation examinations. The development of hydrogen peroxide detection methods that can be conveniently self-tested will help women who are concerned about their own health to conduct self-examination and monitoring. The existing common methods of hydrogen peroxide detection generally require professional operation and processing, which is not conducive to universal application. Since the early symptoms of tumors are very subtle or asymptomatic, early screening of tumors requires strong convenience, ease of use and cost control to adapt to popular application scenarios such as home / community monitoring. It is necessary to develop hydrogen peroxide detection technologies and products that are convenient, easy to use, easy to assemble and produce, and have long-term storage and standby costs that can be controlled. Summary of the Invention

[0004] In some embodiments, a hydrogen peroxide detection kit is introduced, comprising: a sample diluent and a test strip dried with a detection color developer and a quality control color developer, wherein the sample diluent comprises 100-500 mM ammonium oxalate, 0.3-2.0 mM 8-hydroxyquinoline, 50-500 mM sodium chloride, and 10-100 mM first pH adjuster with a pH of 4.5-7.0.

[0005] In the hydrogen peroxide detection kit of some embodiments, the first pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, or 2-(N-morpholino)ethanesulfonic acid.

[0006] In the hydrogen peroxide detection kit of some embodiments, the detection color developer includes 2.0%-8.0% (w / v) starch, 0.5-5.0 mM acetic acid, 0.5%-2.0% (w / v) Brij-35, 1.0-7.0 M potassium iodide, 2.0-10.0 mM 8-hydroxyquinoline, 50-300 mM calcium chloride, 5-100 mM second pH adjuster and 3-15 mM L-ascorbic acid or sodium thiosulfate.

[0007] In the hydrogen peroxide detection kit of some embodiments, the second pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, sodium bicarbonate, or tris(hydroxymethylaminomethane).

[0008] In the hydrogen peroxide detection kit of some embodiments, the quality control color developing reagent includes 5-30 mM neutral red and 10-60 mM sodium bicarbonate.

[0009] In other embodiments, the test strip in the hydrogen peroxide detection kit further includes an intermediate line reagent.

[0010] In some embodiments of the hydrogen peroxide detection kit, the intermediate line reagent includes 5%-15% (w / v) Brij-35 and 5%-15% (w / v) sodium lauryl sulfate.

[0011] In some embodiments of the hydrogen peroxide detection kit, the test strip has a size of 68 mm × 6 mm × 0.7 mm, 3 μL of detection color developer is dried on one end of the test strip, 3 μL of quality control color developer is dried on the other end of the test strip, and 3 μL of intermediate line reagent is dried between the detection color developer position and the quality control color developer position of the test strip.

[0012] In some embodiments of the hydrogen peroxide detection kit, the sample diluent includes 500 mM ammonium oxalate, 2 mM 8-hydroxyquinoline, 450 mM sodium chloride, and 20 mM sodium dihydrogen phosphate-disodium hydrogen phosphate at pH 6.0.

[0013] In the hydrogen peroxide detection kit of some embodiments, the detection color reagent includes 2% (w / v) starch, 1 mM acetic acid, 250 mM calcium chloride, 2% (w / v) Brij-35, 2M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate and 10 mM sodium bicarbonate.

[0014] In the hydrogen peroxide detection kit of some embodiments, the intermediate line reagent comprises 10% (w / v) Brij-35 and 10% (w / v) SDS.

[0015] In the hydrogen peroxide detection kit of some embodiments, the quality control color developing reagent includes 15 mM neutral red and 45 mM sodium bicarbonate.

[0016] In some embodiments, a method for detecting hydrogen peroxide is introduced, comprising the following steps: a. taking a sample to be tested and adding it to a sample diluent in a hydrogen peroxide detection kit to obtain a mixed solution 1; b. pipetting the mixed solution 1 to a sample addition area of ​​a test strip in the hydrogen peroxide detection kit and allowing it to stand for the test strip to develop color.

[0017] The detection method of some embodiments includes: after sampling with a throat swab sampler, immersing the sampling end into a sampling tube with a built-in sample diluent, breaking it along the score, leaving the sampling end in the tube, sealing the sampling tube, and shaking the sampling tube so that the sampling end is completely immersed in the sample diluent to obtain a mixed solution.

[0018] In some embodiments, the detection method, step b, includes: adding a drop of the mixed solution to the sample addition area of ​​the test strip, allowing it to stand until the quality control color developer area fully changes color to purple-red, and observing the color development in the detection color developer area.

[0019] Use of the kit or detection method of some embodiments in detecting lung cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1: Schematic diagram of a test strip and test results.

[0021] Figure 2: A photo of a finished test strip and the test results.

[0022] Figure 3: Schematic diagram of the composition of a kit.

[0023] Figure 4: Front view of the sampling tube containing the mixture of sample and sample diluent.

[0024] Figure 5: A finished product of a test kit and a real photo of the test results.

[0025] Figure markings: S: sample addition area; 321: T area; 322: R area; 323: C area; KA1: sampling end of the sampler; KB1: sampling tube body; KB2: tube cover; KB3: sample diluent; KC1: colorimetric plate housing; KC2: colorimetric area; KC3: quality control area; KC4: sample addition hole; KC5: card slot. DETAILED DESCRIPTION

[0026] In some embodiments, the present invention introduces a hydrogen peroxide detection kit, comprising: a sample diluent and a test strip dried with a detection color developer and a quality control color developer, wherein the sample diluent comprises 100-500 mM ammonium oxalate, 0.3-2.0 mM 8-hydroxyquinoline, 50-500 mM sodium chloride, and 10-100 mM first pH adjuster with a pH of 4.5-7.0.

[0027] In the hydrogen peroxide detection kit of some embodiments, the first pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, or 2-(N-morpholino)ethanesulfonic acid.

[0028] In the sample diluent formula of the present invention, 100-500mM ammonium oxalate can not only complex and precipitate various oxidative metal ions, but also inhibit Fe 3+ 、Fe 2+ The decomposition of hydrogen peroxide mediated by metal ions helps stabilize the hydrogen peroxide in the sample; 8-hydroxyquinoline further enhances the stability of hydrogen peroxide; the pH adjuster maintains the system pH between 4.5 and 7.0, while nitrite's oxidizing properties require a pH of ≤4.0, eliminating interference from nitrite. Sodium chloride provides an appropriate ionic strength for the reaction. The sample diluent of the present invention not only stabilizes hydrogen peroxide dissolved and released from the sample, but also helps prevent plastic materials such as polypropylene and polyethylene from promoting the decomposition of hydrogen peroxide. Furthermore, it does not interfere with the rapid oxidation of iodide ions by hydrogen peroxide.

[0029] In some embodiments of the hydrogen peroxide detection kit, a quality control area is set at the lower end of the test strip, a quality control color developer is added to the quality control area, and the amount of the quality control color developer is 1-2 μL / mm. 9-11 mm below the quality control area is a colorimetric area, a detection color developer is added to the colorimetric area, and the amount of the detection color developer is 1-2 μL / mm. The test strip is dried at 50°C for 5 minutes.

[0030] In the hydrogen peroxide detection kit of some embodiments, the detection color reagent includes 2.0%-8.0% (w / v) starch, 0.5-5.0 mM acetic acid, 0.5%-2.0% (w / v) polyoxyethylene lauryl ether (Brij-35), 1.0-7.0 M potassium iodide, 2.0-10.0 mM 8-hydroxyquinoline, 50-300 mM calcium chloride, 5-100 mM second pH adjuster and 3-15 mM L-ascorbic acid or sodium thiosulfate.

[0031] In the hydrogen peroxide detection kit of some embodiments, the second pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, sodium bicarbonate, or tris(hydroxymethylaminomethane).

[0032] In the hydrogen peroxide detection kit of some embodiments, the quality control color developing reagent includes 5-30 mM neutral red and 10-60 mM sodium bicarbonate.

[0033] In the detection color developer formula of the present application, 2.0%-8.0% (w / v) starch and 1.0-7.0M potassium iodide provide the color development basis, 0.5-5.0mM acetic acid and 0.5%-2.0% (w / v) Brij-35 can promote the dissolution of starch mother liquor and the drying and fixation of the detection indicator on the test strip during the production and preparation process. The local high concentration of Brij-35 can retard the diffusion of the starch solution and enhance the clear cohesion of the color development; 2.0-10.0mM 8-hydroxyquinoline, 3-15mM vitamin C or sodium thiosulfate can eliminate the iodine molecular background, and 5-100mM pH adjuster is used to adjust the system to an alkaline pH, so that the test strip can be preserved for a long time at room temperature or even under hot and dry conditions of 40-45°C.

[0034] In other embodiments, the test strip in the hydrogen peroxide detection kit further includes an intermediate line reagent.

[0035] In some embodiments of the hydrogen peroxide detection kit, the intermediate line reagent includes 5%-15% (w / v) Brij-35 and 5%-15% (w / v) sodium lauryl sulfate.

[0036] In some embodiments of the hydrogen peroxide detection kit, the test strip has a size of 68 mm × 6 mm × 0.7 mm, 3 μL of detection color developer is dried on one end of the test strip, 3 μL of quality control color developer is dried on the other end of the test strip, and 3 μL of intermediate line reagent is dried between the detection color developer position and the quality control color developer position of the test strip.

[0037] In some embodiments, the test strip in the hydrogen peroxide detection kit is shown in FIG1 . The sample addition area S 320 is defined 13 mm from the bottom of the test strip. Sample addition area S 320 corresponds to the sample addition well, and the sample solution to be tested can be dripped or immersed in this area. 12 mm above area S is defined as colorimetric area T 321, to which a detection color developer is dripped. 5 mm above area S is the center line area R 322. 5 mm above area R is the quality control area C 323, to which a quality control color developer is dripped. After drying the test strip at 50°C for 5 minutes, the test strip can be used directly for sample testing or can be sealed and dried at room temperature for storage.

[0038] In some embodiments of the hydrogen peroxide detection kit, the sample diluent includes 500 mM ammonium oxalate, 2 mM 8-hydroxyquinoline, 450 mM sodium chloride, and 20 mM sodium dihydrogen phosphate-disodium hydrogen phosphate at pH 6.0.

[0039] In the hydrogen peroxide detection kit of some embodiments, the detection color reagent includes 2% (w / v) starch, 1 mM acetic acid, 250 mM calcium chloride, 2% (w / v) Brij-35, 2M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate and 10 mM sodium bicarbonate.

[0040] In the hydrogen peroxide detection kit of some embodiments, the intermediate line reagent comprises 10% (w / v) Brij-35 and 10% (w / v) SDS.

[0041] In the hydrogen peroxide detection kit of some embodiments, the quality control color developing reagent includes 15 mM neutral red and 45 mM sodium bicarbonate.

[0042] In some embodiments, a method for detecting hydrogen peroxide is introduced, comprising the following steps: a. taking a sample to be tested and adding it to a sample diluent in a hydrogen peroxide detection kit to obtain a mixed solution 1; b. pipetting the mixed solution 1 to a sample addition area of ​​a test strip in the hydrogen peroxide detection kit and allowing it to stand for the test strip to develop color.

[0043] The detection method of some embodiments includes: after sampling with a throat swab sampler, immersing the sampling end into a sampling tube with a built-in sample diluent, breaking it along the score, leaving the sampling end in the tube, sealing the sampling tube, and shaking the sampling tube so that the sampling end is completely immersed in the sample diluent to obtain a mixed solution.

[0044] In some embodiments, the detection method, step b, includes: adding a drop of the mixed solution to the sample addition area of ​​the test strip, allowing it to stand until the quality control color developer area fully changes color to purple-red, and observing the color development in the detection color developer area.

[0045] Detection methods in some embodiments are shown in FIG2 . When using the present hydrogen peroxide test strip to test a sample, the sample is mixed with a sample diluent consisting of 500 mM ammonium oxalate, 2.0 mM 8-hydroxyquinoline, 450 mM sodium chloride, and 20 mM sodium dihydrogen phosphate / disodium hydrogen phosphate aqueous solution at a volume ratio of 1:4. The lower end of the test strip is then immersed in the sample diluent, but not submerging the colorimetric area. The strip is then left to stand for approximately 10 seconds, and the C zone 323 and T zone 321 are observed. If the C zone 323 appears purple-red, indicating a successful test, the T zone 321 appears distinctly blue-black; a positive result indicates that the hydrogen peroxide content in the sample is ≥1.0 mM. If the colorimetric area appears only a faint background color or light gray, a negative result indicates that the hydrogen peroxide content in the sample is <1.0 mM.

[0046] After testing the purified water control sample and 1.5mM hydrogen peroxide as the positive control sample using this hydrogen peroxide detection test strip as described above, it can be observed that the quality control area of ​​the sample is purple-red, indicating that the operation is effective; the colorimetric area of ​​the negative control sample is a light background color, indicating a negative result; the colorimetric area of ​​the positive control sample is a distinct blue-black color, indicating a positive result.

[0047] The hydrogen peroxide detection kit and its use method in some embodiments are shown in Figures 3-5, including a colorimetric card case KC1, a throat swab sampler KA, a sampling tube KB with a built-in sample diluent, and a colorimetric card KC with a built-in test strip. The colorimetric card KC is provided with a sample addition hole KC4. (The colorimetric card KC is a colloidal gold card case with an external specification of 70×20mm, a sample addition hole specification of 6×3mm, and a reading window specification of 16×3.5mm. There is a card slot KC5 for fixing the test strip inside, with a specification of 60×4mm. The test strip is fixed in the card slot KC5 and consists of a 60mm×4mm (length×width) filter paper. The reagents in the quality control area KC3 and the colorimetric area KC2 are dry and fixed on the filter paper of the test strip at 38mm and 24mm from the lower end.

[0048] The sample diluent KB3 built into the sampling tube body KB1 includes 500mM ammonium oxalate, 2.0mM 8-hydroxyquinoline, 450mM sodium chloride and 20mM pH 6.0 sodium dihydrogen phosphate-disodium hydrogen phosphate mixed aqueous solution, and the volume of the sample diluent is 300μL.

[0049] It should be noted that there is a card slot KC5 for fixing the test strip inside the colorimetric plate KC. The middle line area R area can be set without setting the card slot KC5. After the sampling and testing operation is completed according to the aforementioned usage steps, a clear purple-red color can be observed in the quality control area, indicating that the sample and operation are qualified and the results are reliable. If the hydrogen peroxide content in the sample is ≥1.0mM, a clear blue-black color will be observed in the colorimetric area after the test is completed.

[0050] In some embodiments, a hydrogen peroxide detection kit and a method of using the same are used in the preliminary detection of lung cancer.

[0051] The present invention can preliminarily distinguish between abnormal increase of airway hydrogen peroxide related to lung cancer and airway inflammation caused by external stimuli such as infection and allergy. Airway reactive oxygen species include hydrogen peroxide and O 2- OH - , lipid peroxides, etc. Among them, only hydrogen peroxide can directly diffuse into airway mucus. 2- and OH - The half-life is very short, and it only exists briefly in the cell and is quickly converted into hydrogen peroxide and lipid or protein peroxides. Hydrogen peroxide is usually cleared efficiently and quickly by CAT. Therefore, the increase in airway reactive oxygen species related to inflammation mainly exists in the cell in the form of lipid or protein peroxides. Lung cancer usually does not have typical manifestations of airway inflammation. The abnormal increase in its related airway reactive oxygen species is mainly due to the accumulation of hydrogen peroxide caused by CAT downregulation and airway acidification, which can directly diffuse and manifest in airway mucus. Airway nitric oxide (NO) can be increased in some airway inflammations or lung cancers, mainly in the form of nitrite (NO 2- ) in the airway mucus and exhibit oxidizing properties under highly acidic conditions. Dissolved oxygen and oxidizing metal ions are also common oxidizing substances in the airways.

[0052] The present invention uses mild sample dilution and dissolution conditions, avoiding the use of surfactants to reduce the release of intracellular lipid or protein peroxides by lysis; uses weak acidity of pH 6.0 and metal ion complexation conditions, avoiding the use of stronger acidic conditions to prevent nitrite or dissolved oxygen from acquiring strong oxidizing properties, and complexes and passivates metal ions that may be present in the sample and can actively participate in redox reactions, so that the detected increase in oxidizing properties can specifically indicate an abnormal increase in airway hydrogen peroxide, which is closely related to lung cancer, and avoid confusion with common airway inflammation.

[0053] In the present invention, unless otherwise specified, all raw material components are commercially available products well known to those skilled in the art. The technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all.

[0054] Example 1

[0055] A hydrogen peroxide detection kit comprises a sample diluent consisting of a mixed aqueous solution of 500 mM ammonium oxalate, 2 mM 8-hydroxyquinoline, 450 mM sodium chloride and 20 mM pH 6.0 pH regulator.

[0056] The test strip is 68 mm long and 6 mm wide. The T zone is set 27 mm from the bottom of the test strip, the R zone is set 5 mm above the T zone, and the C zone is set 5 mm above the R zone.

[0057] Zone C quality control colorimetric reagent: 5 mM neutral red, 10 mM sodium bicarbonate.

[0058] T-zone detection color reagent: 2% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0059] R zone middle line reagents: 5% SDS, 5% Brij-35.

[0060] Example 2

[0061] Zone C quality control colorimetric reagent: 15 mM neutral red, 35 mM sodium bicarbonate.

[0062] T-zone detection color reagent: 2% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0063] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0064] Example 3

[0065] Zone C quality control colorimetric reagent: 30 mM neutral red, 60 mM sodium bicarbonate.

[0066] T-zone detection color reagent: 2% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0067] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0068] Example 4

[0069] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0070] T-zone detection color reagent: 4% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0071] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0072] Example 5

[0073] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0074] T-zone detection color reagent: 6% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0075] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0076] Example 6

[0077] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0078] T-zone detection color reagent: 8% starch, 0.5% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0079] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0080] Example 7

[0081] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0082] T-zone detection color reagent: 2% starch, 1% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0083] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0084] Example 8

[0085] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0086] T-zone detection color reagent: 2% starch, 2% Brij-35, 0.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0087] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0088] Example 9

[0089] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0090] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0091] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0092] Example 10

[0093] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0094] T-zone detection color reagent: 2% starch, 2% Brij-35, 2.5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0095] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0096] Example 11

[0097] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0098] T-zone detection color reagent: 2% starch, 2% Brij-35, 5 mM acetic acid, 1 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0099] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0100] Example 12

[0101] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0102] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0103] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0104] Example 13

[0105] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0106] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 4 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0107] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0108] Example 14

[0109] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0110] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 7 M potassium iodide, 2 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0111] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0112] Example 15

[0113] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0114] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0115] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0116] Example 16

[0117] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0118] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 8 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0119] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0120] Example 17

[0121] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0122] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 10 mM 8-hydroxyquinoline, 3 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0123] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0124] Example 18

[0125] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0126] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 7 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0127] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0128] Example 19

[0129] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0130] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 11 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0131] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0132] Example 20

[0133] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0134] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 50 mM calcium chloride.

[0135] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0136] Example 21

[0137] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0138] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 35 mM sodium bicarbonate, 50 mM calcium chloride.

[0139] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0140] Example 22

[0141] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0142] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 65 mM sodium bicarbonate, 50 mM calcium chloride.

[0143] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0144] Example 23

[0145] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0146] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 100 mM sodium bicarbonate, 50 mM calcium chloride.

[0147] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0148] Example 24

[0149] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0150] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 100 mM calcium chloride.

[0151] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0152] Example 25

[0153] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0154] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 200 mM calcium chloride.

[0155] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0156] Example 26

[0157] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0158] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 300 mM calcium chloride.

[0159] Reagents for the middle line of the R region: 5% SDS, 5% Brij-35. Others are the same as in Example 1.

[0160] Example 27

[0161] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0162] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 300 mM calcium chloride.

[0163] The reagents for the middle line of the R region are: 10% SDS, 10% Brij-35. The rest are the same as in Example 1.

[0164] Example 28

[0165] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0166] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 5 mM sodium bicarbonate, 300 mM calcium chloride.

[0167] The reagents for the middle line of the R region are: 15% SDS, 15% Brij-35. The rest are the same as in Example 1.

[0168] Example 29

[0169] Zone C quality control colorimetric reagent: 15 mM neutral red, 45 mM sodium bicarbonate.

[0170] T-zone detection color reagent: 2% starch, 2% Brij-35, 1 mM acetic acid, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, 10 mM sodium bicarbonate, 250 mM calcium chloride.

[0171] The reagents for the middle line of the R region are: 10% SDS, 10% Brij-35. The rest are the same as in Example 1.

[0172] Example 30

[0173] A hydrogen peroxide detection kit comprises a sample diluent consisting of a mixed aqueous solution of 100 mM ammonium oxalate, 50 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator.

[0174] The test strip is 68 mm long and 6 mm wide. The T zone is set 27 mm from the bottom of the test strip, the R zone is set 5 mm above the T zone, and the C zone is set 5 mm above the R zone.

[0175] Zone C quality control colorimetric reagent: 3 μL of 15 mM neutral red and 45 mM sodium bicarbonate solution.

[0176] T zone detection color developer: 3 μL of a mixed aqueous solution of 2% (w / v) starch, 2% (w / v) Brij-35, 1 mM acetic acid, 250 mM calcium chloride, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, and 10 mM sodium bicarbonate.

[0177] R zone middle line reagent: 3 μL of a mixed solution of 10% (w / v) SDS and 10% (w / v) Brij-35.

[0178] Example 31

[0179] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 300 mM ammonium oxalate, 50 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator, with the remaining components being the same as those in Example 30.

[0180] Example 32

[0181] A hydrogen peroxide detection kit, wherein the sample diluent comprises a mixed aqueous solution of 500 mM ammonium oxalate, 50 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator, with the remaining components being the same as those in Example 30.

[0182] Example 33

[0183] A hydrogen peroxide detection kit, wherein the sample diluent comprises a mixed aqueous solution of 500 mM ammonium oxalate, 200 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator, with the remaining components being the same as those in Example 30.

[0184] Example 34

[0185] A hydrogen peroxide detection kit, wherein the sample diluent comprises a mixed aqueous solution of 500 mM ammonium oxalate, 350 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator, with the remaining components being the same as those in Example 30.

[0186] Example 35

[0187] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 0.3 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0188] Example 36

[0189] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 1 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0190] Example 37

[0191] A hydrogen peroxide detection kit, wherein the sample diluent comprises a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 1.5 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator, with the remaining components being the same as those in Example 30.

[0192] Example 38

[0193] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 10 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0194] Example 39

[0195] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 25 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0196] Example 40

[0197] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 50 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0198] Example 41

[0199] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 100 mM pH 4.5 pH regulator. The remaining steps are the same as those in Example 30.

[0200] Example 42

[0201] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 25 mM pH 6.0 pH regulator, with the remaining components being the same as those in Example 30.

[0202] Example 43

[0203] A hydrogen peroxide detection kit, wherein the sample diluent comprises: a mixed aqueous solution of 500 mM ammonium oxalate, 500 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 25 mM pH 7.0 pH regulator, with the remaining components being the same as those in Example 30.

[0204] Example 44

[0205] A hydrogen peroxide detection kit, wherein the sample diluent comprises: 500 mM ammonium oxalate, 450 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 25 mM pH 6.0 sodium dihydrogen phosphate-disodium hydrogen phosphate mixed aqueous solution. The remaining components are the same as those in Example 30.

[0206] Example 45

[0207] A hydrogen peroxide detection kit, wherein the sample diluent comprises: 500 mM ammonium oxalate, 450 mM sodium chloride, 2 mM 8-hydroxyquinoline, and 20 mM pH 6.0 sodium dihydrogen phosphate-disodium hydrogen phosphate mixed aqueous solution. The remaining components are the same as those in Example 30.

[0208] Experimental Example 1

[0209] The present invention compares throat swab sampling with the sputum sampling disclosed in the patent application document with application number CN 2022115399358. In previous experimental studies, it was found that after rinsing the mouth and cleaning the mouth, the user drank a sip of water, patted the chest or back 10 times, took a deep breath, and coughed hard 6 times, prompting a small amount of airway mucus to be discharged from the airway and accumulated in the mouth. The oral fluid was then coughed up and collected for detection, and the airway-specifically expressed mucin MUC5AC or its mRNA was detected 100%, indicating that airway mucus samples could be collected. However, during the expansion and promotion, it was found that some people, especially young children, had difficulty coughing up oral fluid, resulting in sampling failure. In order to ensure the success rate of sampling, the throat swab sampling method and the autonomous sputum sampling method of the patent application document with application number CN 2022115399358 were adopted respectively.

[0210] Throat swab sampling method: After following the aforementioned procedures to encourage a small amount of airway mucus to drain from the airway and collect in the mouth, the user does not need to cough it up to collect it. Instead, they wipe the mouth with a throat swab sampler, then immerse the sampling tip of the sampler in the sample diluent and break it along the notch, leaving both the sample and the sampling tip in the sample diluent to complete the sampling. CN 2022115399358 Self-experienced sputum sampling method: After following the aforementioned procedures to encourage a small amount of airway mucus to drain from the airway and collect in the mouth, the user coughs it up to collect it.

[0211] 30 healthy volunteers were recruited, including 20 adults and 10 children aged 1-5 years old. Two sampling methods were compared once a day at different times for three consecutive days. Among them, adults took samples independently throughout the process, while children were guided by their parents to cough 6 times, and then their parents took samples from the children using throat swabs. After sampling, the sampling tube weight was compared before and after sampling to calculate the sampling weight; the sampling volume was measured using a sampling pipette to determine that the throat swab sampling method provided by the present invention was successful for both adult and child volunteers, with minimal sampling fluctuations: the sampling volume was approximately 100±15μL (volume) or 100±15mg (weight). In contrast, the sputum sampling method of patent application document CN 2022115399358 successfully sampled adults in the morning and evening, but the sampling volume was approximately 850±185μL (volume) or 850±150mg (weight). Five adult samples failed in the afternoon, and three samples failed in each period for children. Specific statistics are shown in the table below.

[0212] Table 1 Test statistics of throat swab and sputum sampling of volunteers

[0213] The kits described in Examples 1 to 29 were used to detect whether the airway mucus samples provided by the donor contained trace amounts of airway hydrogen peroxide, thereby detecting whether the donor had a higher risk of lung cancer. The negative control was purified water, and the positive control was 1.5 mM hydrogen peroxide.

[0214] The specific steps are:

[0215] 1. Sampling: When taking samples with the pharyngeal swab sampler KA, you can directly insert the pharyngeal swab sampler KA into your mouth and hold it in your mouth for about 3 seconds to take a sample. For airway mucus testing, after rinsing your mouth, the user should drink a sip of water, pat their chest or back 10 times, take a deep breath, and cough and gargle forcefully 6 times to encourage a small amount of airway mucus to be discharged from the airway and gathered in the mouth. Then, insert the pharyngeal swab sampler KA into your mouth and hold it in your mouth for about 3 seconds to take a sample.

[0216] 2. After sampling, unscrew the cap KB2 of the sampling tube KB, immerse the sampling tip KA1 of the throat swab sampler KA into the sample diluent in the tube, and break it along the notch on the throat swab. Tighten the cap and shake vigorously for about 5 seconds. Then, tap the bottom of the tube on the table 5 times. Unscrew the cap and drip all the liquid in the tube into the sample well of the colorimetric plate KC.

[0217] 3. Then let it stand until the quality control area C shows a distinct purple-red color, indicating that the sample and operation are qualified and the result is reliable. If the T window shows a distinct blue-black color, it is positive, indicating that the hydrogen peroxide content in the sample is ≥1.0mM, indicating a high risk of lung cancer. It is recommended to go to a hospital or other professional institution for in-depth examination related to lung cancer.

[0218] The results showed that the negative compliance rate of the test strips prepared in Examples 1 to 29 was 100%, and the positive compliance rate was 100%. Obvious purple-red color was observed in the C area, indicating that the operation was correct; no obvious color was observed in the negative control colorimetric area, indicating that the test result was negative; obvious blue-black color was observed in the positive control colorimetric area, indicating that the test result was positive. Therefore, the test strips prepared in Examples 1 to 29 can all achieve expected test results.

[0219] Experimental Example 2

[0220] Referring to the operating steps in Experimental Example 1, the hydrogen peroxide test was performed using the kits of Examples 30 to 45. The negative control was purified water, and the positive control was 1.5 mM hydrogen peroxide. It was found that in the test strips of the kits of Examples 30 to 45, an obvious purple-red color was observed in the C area, indicating that the operation was correct; no obvious color was observed in the colorimetric area of ​​the negative control, indicating that the test result was negative; and an obvious blue-black color was observed in the colorimetric area of ​​the positive control, indicating that the test result was positive. Therefore, the kits of Examples 30 to 45 can all achieve the expected test results.

[0221] Experimental Example 3

[0222] For test strip specifications and preparation, see Example 45. Following the procedures in Experimental Example 1, 100 mg of a bovine serum albumin (BSA) / skim milk powder solution and 100 μL of a 3.0 mM FeCl₃ / CuSO₄ / NaNO₂ solution were added to 100 μL of a 2.0 mM H₂O₂ solution, respectively. Each experiment was repeated three times. The results are shown in Table 2. The hydrogen peroxide detection method showed no color development for any substance other than hydrogen peroxide, demonstrating the method's good specificity.

[0223] Table 2 Specificity test results

[0224] Experimental Example 4

[0225] For test strip specifications and preparation, refer to Example 45, and refer to the operating procedures in Experimental Example 1. Hydrogen peroxide concentrations of 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, and 1.6 mM were prepared for gradient testing. Each sample was tested 20 times in duplicate, for a total of three batches. The results are shown in Table 3. Samples with hydrogen peroxide concentrations as low as 0.8-1.5 mM clearly developed color, while samples with concentrations above 1.5 mM developed 100% clear color. At 0.7 mM, the color development rate was 0, meaning 100% no color was developed. This sample dilution not only achieved good hydrogen peroxide detection sensitivity but also ensured excellent specificity.

[0226] Table 3 Detection results of hydrogen peroxide at different concentrations

[0227] At the same time, further expanded testing was performed on the method of patent application document with application number CN 2022115399358, and 0.6mM / 0.7mM / 1.5mM / 1.6mM hydrogen peroxide tests were also performed as shown in the following table. The results are shown in Table 4. The present invention can better control the background of 0.7mM hydrogen peroxide and more clearly indicate the difference between 1.5mM and 0.7mM hydrogen peroxide samples.

[0228] Table 4 Test results of deep cough sampling method CN 202211539935.8

[0229] Experimental Example 5

[0230] For test strip specifications and preparation, refer to Example 45, and follow the procedures in Experimental Example 1. Add 3 μL of a 15 mM neutral red and 45 mM sodium bicarbonate solution to the control area; 3 μL of a 10% (w / v) SDS and 10% (w / v) Brij-35 solution to the centerline area; and 3 μL of a 2% (w / v) starch, 2% (w / v) Brij-35, 1 mM acetic acid, 250 mM calcium chloride, 2 M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate, and 10 mM sodium bicarbonate solution to the detection area. The sample diluent consists of a 500 mM ammonium oxalate, 2 mM 8-hydroxyquinoline, 450 mM sodium chloride, and 20 mM pH 6.0 pH adjuster solution. Purified water was used as the negative control, and 1.5 mM H₂O₂ was used as the positive control. The interfering substances tested are listed in Table 5. The interfering substance content in the sample was standardized at 2% of the sample volume based on the typical usage of the substance, which was higher than the volume ratio of the substance remaining in the sample after routine use and rinsing. Each sample was tested three times, for a total of three batches of tests. Observational statistics showed that after the operation was completed, a distinct purple-red color was observed in Area C, indicating correct operation. No significant color was observed in the negative control test area, indicating a negative test result. A distinct blue-black color was observed in the positive control test area, indicating a positive test result. Therefore, within the tested concentration range, the interfering substances did not affect the test results, indicating no interference.

[0231] Table 5 Interfering substances

[0232] Experimental Example 6

[0233] In this example, 1200 test kits were produced. First, the production batch number and production plan (Table 6) were developed, and materials were prepared (Table 7).

[0234] Table 6. 1200 sets of planned quantity groups

[0235] Table 7. Bill of Materials

[0236] Key process - test strip preparation: On a 270g ordinary qualitative filter paper cut into a size of 300mm×68mm, mark the test area with a pencil line approximately 27mm from the bottom, mark the middle line area with a pencil line 5mm above it as the quality control area, and mark the quality control area with a pencil line 5mm above it as the quality control area. According to the method in the main text of the instruction manual, use a spotting pen to mark the quality control area with 15mM neutral red and 45mM sodium bicarbonate solution. The middle line area is a mixed solution of 10% (w / v) SDS and 10% (w / v) Brij-35. Use a spotting pen to mark the test area with a mixed aqueous solution of 2% (w / v) starch, 2% (w / v) Brij-35, 1mM acetic acid, 250mM calcium chloride, 2M potassium iodide, 5mM 8-hydroxyquinoline, 15mM sodium thiosulfate, and 10mM sodium bicarbonate, with a natural pH value; dry at 50°C for 5 minutes. After the test strips are prepared, random inspections of key processes should be carried out to ensure that the test strips are of qualified quality.

[0237] Preparation and packaging of sample diluent: Prepare 360 ​​ml of the sample diluent according to the formula of the sample diluent of the test kit in Example 45, and dispense it into sampling tubes at 300 μL / portion. Seal the sampling tubes tightly, place the sampling tubes in a ziplock bag containing a desiccant, seal it, and temporarily store it at room temperature to obtain the semi-finished product. After passing the random inspection, carry out outer packaging.

[0238] Assembling the test board: Place the test strip into the lower card slot, ensuring the bottom end of the test strip is flush with the slot. Secure the upper card slot to the lower card slot, ensuring that the C and T positions correspond to the test strip's quality control and test areas, respectively. After on-site inspection by the workshop inspector confirming satisfactory assembly, a pass mark is affixed to the top of the test board. The test board is then placed in an aluminum foil bag containing desiccant, sealed, and temporarily stored at room temperature to produce a semi-finished product. Once it passes random inspection, it is packaged.

[0239] Outer packaging: Place the semi-finished products and instructions that have passed the random inspection into the outer packaging box, affix the certificate of conformity, and put it into storage for later use.

[0240] Quality inspection: random inspection of key processes. After the test strips are prepared, random inspection of key processes is carried out to ensure that the test strips are of qualified quality.

[0241] Appearance: The appearance and size of the test strip should conform to the settings.

[0242] Negative reference product compliance rate: Take 10 samples of the company's negative reference products for testing, and the test results should all be negative. Positive reference product compliance rate: Take 10 samples of the company's positive reference products for testing, and the test results should all be positive.

[0243] Detection limit: Repeat the test 20 times with the company's detection limit reference product, and at least 19 of the test results should be positive.

[0244] Semi-finished product inspection

[0245] Appearance: All components are complete, the aluminum foil bag and desiccant packaging are intact, without leakage, damage, or moisture regain; the label is clear, a qualified mark is affixed to the top of the indicator slot, and the window position matches the corresponding area on the test strip.

[0246] Negative Reference Material Compliance Rate: Directly use the results of random inspections of key processes. Positive Reference Material Compliance Rate: Directly use the results of random inspections of key processes. Detection Limit: Directly use the results of random inspections of key processes. Repeatability: Take the company's repeatability reference material and repeat the test 10 times. The test results should all be positive and uniform in color.

[0247] Sample processing efficiency: Samples were collected according to the airway mucus sampling method described in the example. Five negative samples obtained by mixing and aliquoting airway mucus samples from 10 healthy volunteers and five positive samples obtained by mixing and aliquoting airway mucus samples from 10 lung cancer patients were tested separately. The test results of the negative samples should all be negative, and the test results of the positive samples should all be positive.

[0248] Finished product inspection

[0249] Appearance: All components must be complete, with the aluminum foil bag and desiccant packaging intact and free of leaks, damage, or moisture. The label must be legible, with a qualified mark affixed to the top of the indicator slot, and the window position must match the corresponding area on the test strip. The finished product should weigh ≥ 9.0g, indicating sufficient reagents. Performance: After visual verification, performance indicators from key process inspections and semi-finished product inspections will be used.

[0250] Healthy volunteer testing of this hydrogen peroxide detection kit: 50 healthy volunteers, aged 2-40, working in high-tech industries, with a gender ratio of approximately 50:50. They had undergone a comprehensive physical examination within the past year and showed no abnormalities, history of heavy smoking, work in heavily polluting industries, lung cancer, asthma, or other lung diseases, nor a family history of related diseases. Some of the women were pregnant or breastfeeding and felt well. Staff members presided over registration, distributed the finished kit, and provided a hands-on demonstration. Volunteers then performed the test according to the requested method and reported the results to the staff. After verification, the staff conducted statistical analysis of the results. The results showed that all 50 C-windows showed a distinct purple-red color, indicating 100% accuracy. All 50 T-windows showed no significant coloration, indicating a low risk of lung cancer in the volunteers.

[0251] Lung cancer patient testing of this hydrogen peroxide detection kit: 24 lung cancer patient volunteers, aged 40-70 years, of all occupations, with a gender ratio of approximately 50%. All volunteers had been diagnosed with lung cancer and had not undergone surgical treatment. None of the women were pregnant or breastfeeding. Staff members presided over registration, distributed the finished kit, and provided a user experience demonstration. If a foreign object was felt in the throat, volunteers should cough gently to clear their throat and expel phlegm. Then, they should perform the test according to the instructions and report the results to the staff. After verification by the staff, the results were statistically analyzed. The results showed that all 24 C-windows showed a distinct purple-red color, indicating 100% correct operation. All 24 T-windows showed a distinct blue-black color, indicating abnormally elevated airway H2O2 levels and a higher risk of lung cancer.

[0252] Testing of this hydrogen peroxide detection kit in patients with lung nodules: 80 volunteers with lung nodules, aged 20-70 years, of all occupations, with a gender ratio of approximately 50:50, were recruited. They had lung nodules diagnosed with non-lung cancer by low-dose spiral CT scans this year. Women were not pregnant or breastfeeding. Staff members presided over registration, distributed the finished kit, and provided a user guide and operation demonstration. If a foreign object was felt in the throat, the volunteers should cough gently to clear their throat and expel phlegm. Then, they should perform the test according to the instructions and report the results to the staff. After verification, the staff performed statistical analysis of the results. The results showed that all 80 C-windows showed a distinct purple-red color, indicating 100% accuracy. All 80 T-windows showed no significant coloration, indicating a low risk of lung cancer in these volunteers.

[0253] This hydrogen peroxide detection kit was tested on patients with upper respiratory tract inflammation. Thirty patients each with colds, coughs, rhinitis, and pharyngitis were recruited. They were aged 20-40 years, with no occupational restrictions and a gender ratio of approximately 50-50. They had undergone a comprehensive physical examination within the past year and had no other abnormalities, a history of heavy smoking or work in heavily polluting industries, a history of lung cancer, asthma, or other lung diseases, and no family history of related diseases. Women were not pregnant or breastfeeding. Staff members presided over registration, distributed the finished kit, and provided a hands-on demonstration. After clearing their noses and coughing gently to clear their throats and eliminate phlegm, volunteers performed the test according to the instructions and reported the results to the staff. After verification, the staff performed statistical analysis. The results showed that all 90 C-windows showed a distinct purple-red color, indicating 100% accuracy. All 90 T-windows showed no significant coloration, indicating a low risk of lung cancer in these volunteers.

[0254] The application of this hydrogen peroxide detection kit in lung cancer physical examination screening: a staff member is arranged on-site at the physical examination center to be responsible for the registration form and the distribution and recovery of the kit, and to provide necessary answers and guidance when the subjects have questions, and to play a promotional explanation video in a rolling manner. The method of using the kit is the same as described in the main text of the instruction manual. Physical examination subjects can be tested whenever they arrive, and the average time is about 5 minutes. When there are multiple people present, they can test themselves in parallel, and it only takes about 10 minutes in total to test 30 people. Based on this estimate, about 1,440 people can be tested in 1 day (8 hours). Low-dose spiral CT generally takes 3-5 minutes, but subjects usually need to wait in line. It takes about 150 minutes to test 30 people, and 96 people can be completed in 8 hours. The present invention can improve the efficiency of physical examination screening by about 15 times. In addition, it usually takes 1-3 days for low-dose spiral CT to produce results, while the present invention can produce results within 5 minutes.

[0255] Community application of this hydrogen peroxide detection kit: 100 sets of the finished kit of the present invention were distributed in a pilot community supermarket, and promotional posters with QR codes were posted. Users could scan the codes with their mobile phones to watch a video explanation. After registering online, they could pick up the kit on-site and perform the test themselves. They followed the method described in the instructions for the kit and then updated the test results online. Test results were summarized and counted. Positive results were counted according to the user's reported results, while negative results were counted according to the user's reported results. Those who did not report a result were also counted as negative. The logic behind this is that positive results are more likely to stimulate users' desire to communicate and have a low underreporting rate, while negative results may reduce users' desire to communicate and have a high underreporting rate. Statistics show that there were 2 positive cases, with a positive rate of 2%, which is higher than the lung cancer incidence rate in the general population (0.08%). Considering that 70% of the trial users had lung nodules, the lung nodule positivity rate was 3.6%, and the total positive rate of 2% is reasonable. Community hospitals are a key component of national health construction. Currently, lung cancer screening cannot be performed in community hospitals, requiring examinations at central hospitals, which is not conducive to lung cancer screening. The kit of the present invention facilitates residents to participate in lung cancer screening in community hospitals and is expected to develop into a powerful supplement to national health construction.

[0256] Home self-testing use of this hydrogen peroxide detection kit: 30 volunteers with lung nodules were recruited to use the kit of the present invention to take samples for testing at home, once a month, and go to the hospital for regular follow-up examinations as prescribed by the doctor. The volunteers were aged 20-70 years old, with a male-female ratio of about 50%. The user will send the test results back through the network or the staff will follow up by phone or WeChat to collect usage information and register the test results. The statistical results showed that there were 2 positive results, one of whom was a female, 28 years old. A physical examination that year found a 4mm lung nodule. The first 3 test kits were negative, and the fourth test kit was positive. She went to the hospital for a follow-up examination. The right upper lung nodule increased to 6.5mm. The doctor made a comprehensive judgment and recommended surgical treatment. The right lung nodule was wedge resection under general anesthesia and thoracoscopic surgery was performed. Microinvasive adenocarcinoma was confirmed by pathology. Another patient was a 35-year-old male with a 12-year smoking history. During a physical examination the previous year, a 3mm lung nodule was found. The test kit tested negative on the first 10 occasions, but positive on the 11th. He went to the hospital for a follow-up examination. The nodule in the left upper lobe had grown to 5mm. The doctor made a comprehensive assessment and recommended surgery. Pathology confirmed the diagnosis as microinvasive adenocarcinoma. CT follow-up is complex and varied, and the cycles are relatively long, which is not conducive to the timely diagnosis of lung nodules and the timely detection of lung cancer. The present invention facilitates patients to self-test and track at home, which is conducive to promoting patients to go to the hospital for a follow-up examination.

[0257] Storage and stability testing of this hydrogen peroxide detection kit: The test plate (including test strips) was sealed and dry assembled within the finished kit packaging. The kit should be stored at room temperature (25°C ± 5°C) or refrigerated, with a humidity of ≤60%. The negative reference sample was prepared by mixing airway mucus samples from healthy volunteers and aliquoting them in 1ml portions. Standard absorbance determination indicated a H2O2 content of <0.5mM. The positive reference sample was prepared by mixing airway mucus samples from lung cancer patients and aliquoting them in 1ml portions. Standard absorbance determination indicated a H2O2 content of ≥1.0mM. The repeatability reference sample was 1.5mM H2O2, and the limit of detection reference sample was 1.0mM H2O2. The test method was performed according to the manufacturer's instructions. Each batch was tested three times for each of the negative / positive / repeatability criteria, with a limit of detection of five times. One batch was tested monthly. Preliminary statistics from six consecutive tests over six consecutive months indicate that the compliance rate for the negative / positive / repeatability criteria reached 100%, meeting quality requirements.

[0258] Accelerated stability testing of the hydrogen peroxide detection kit was performed: Three batches of the present invention's kit and the kit using the CN 202211539935.8 method were stored in the dark at 45°C and 50°C, respectively, for one week. The test was then performed according to the instructions for the reference sample. Each batch was tested three times for negative / positive / repeatability, with a detection limit of 5. The statistical results are shown in Table 8. The method and kit of the present invention all achieved 100% compliance with the reference sample for the negative / positive / repeatability detection limit, meeting quality requirements. However, the CN 202211539935.8 method showed a decrease in the compliance rate with the reference sample. This indicates that the accelerated stability of the present invention's kit is superior to that of the CN 202211539935.8 method.

[0259] Table 8 Accelerated stability test results

[0260] Transport stability and portability of this hydrogen peroxide detection kit: Transport stability and portability are important considerations for point-of-care (POCT) product applications. The better these characteristics, the more suitable the product is for rapid on-site testing. Three batches of the present invention's kit and the patented kit from CN 2022115399358, 20 units per batch, were stored at 40°C for 7 days. The kits were then dropped from a height of 1.5 meters (3 times) for a shock drop test. The outer and inner packaging of the kits were inspected for damage. The kits were then tested using negative, positive, limit of detection, and repeatability references. The results are shown in Table 9: The inner and outer packaging of the present invention's kits were intact, and the compliance rates for negative / positive / repeatability / limit of detection all reached 100%, meeting quality requirements. However, one batch of the CN 2022115399358's kit had partial outer packaging damage, but the inner packaging was intact. Testing statistics showed that the compliance rates for negative / positive / repeatability / limit of detection did not reach 100%. Therefore, the kit of the present invention has better transportation stability and portability than the kit of the patent CN 2022115399358.

[0261] Table 9 Transport stability test results

[0262] The principle of the present invention is mainly based on the fact that hydrogen peroxide can quickly oxidize iodine ions under various conditions to generate iodine molecules (I2), which combine with starch to develop color. However, in order to perform qualitative detection on the test strip, three technical difficulties need to be overcome: 1) The combination of iodine and starch needs to be condensed to enhance the clarity and readability of the results. The color development reaction of the combination of iodine and starch needs to be carried out in a solution state, and the dissolution of iodine and starch on the test strip can cause diffusion dilution and weaken the readability of the test results, resulting in a serious decrease in detection sensitivity. 2) It is necessary to eliminate the background interference of iodine molecules caused by dissolved oxygen and oxygen in the air. Potassium iodide solution and test paper can be slowly oxidized and deteriorated by dissolved oxygen and oxygen in the air, resulting in a small amount of iodine molecular background. It is necessary to select a reducing agent of suitable type and concentration to remove and control it. 3) It is necessary to eliminate the interference of oxidizing metal ions and nitrites and other oxidizing substances that may exist in the sample, as well as proteins and enzymes. The present invention solves these three difficulties through experimental testing and optimization. In general, the main invention points and beneficial effects of the present invention are:

[0263] 1. The design of the present invention focuses on improving the versatility of sample collection and processing and the anti-interference and stability of the color development reaction system. The present invention establishes an internationally pioneering system for sample lysis processing, preservation and interference elimination. While releasing hydrogen peroxide from the sample, it stabilizes the hydrogen peroxide in the solution and simultaneously eliminates the interference of metal ions, nitrites and proteins, thereby enhancing the sensitivity and specificity of hydrogen peroxide detection from biological samples. Colorimetric methods such as iodine titration are commonly used methods for hydrogen peroxide determination, but they require precise measurement and the reagents used need to be prepared and used immediately, which are not convenient for POCT instant rapid testing. The hydrogen peroxide test is based on a rapid redox reaction and needs to eliminate the interference of various substances that can actively participate in the redox reaction. Samples of biological origin may usually contain metal ions, nitrites and other interfering substances with strong oxidizing properties under acidic conditions. The present invention eliminates these interferences by controlling the pH of the system to a weakly acidic condition and supplementing it with a chelating agent such as potassium ferrocyanide that works under acidic conditions. Samples of biological origin are usually rich in proteins, and the sulfhydryl groups of cysteine ​​or the disulfide bonds of cystine they constitute are in a folded and protected state in the initial state of the sample. During the sample lysis or detection process, they may be exposed to the solution system and react with hydrogen peroxide or the reducing agent used for detection, thereby interfering with the detection. Samples of biological origin may also be rich in catalase, peroxidase and amylase, which may also degrade the hydrogen peroxide released in the sample or the starch used for color development, thereby interfering with the detection. How to conveniently and quickly eliminate the interference of these proteins without causing the loss of hydrogen peroxide in the sample is also a difficult problem that plagues the detection of hydrogen peroxide in biological samples. The present invention uses 8-hydroxyquinoline to simultaneously precipitate macromolecules such as nucleic acids and proteins and stabilize the hydrogen peroxide in the sample, thereby significantly improving the repeatability of the detection. It also has a bactericidal and disinfecting effect, which helps to prevent the spread of pathogens that may be present in biological samples. The formulation of the reagent for hydrogen peroxide detection provided by the present invention allows the reagent used for detection to be not only stored for a long time at room temperature for future use, but also to be stably preserved under hot conditions of 40-45°C. This significantly improves the convenience and ease of use of hydrogen peroxide detection compared to traditional methods that require fresh preparation or low-temperature storage.

[0264] 2. When used for airway sample testing, the present invention shares the advantages of Chinese invention patent application CN 2022115399358, which facilitates point-of-care (POCT) testing. Furthermore, it offers the following significant enhancements and improvements over the CN 2022115399358 method: A) Enhanced sampling success rate and versatility. The CN 2022115399358 method is limited to airway sampling and requires the user to cough up a sample, a process that some users struggle to master. However, the present invention utilizes throat swab sampling, achieving a 100% sampling success rate. This effectively overcomes the limitations of some users who struggle to cough up a sample, improving the versatility and specificity of airway sampling for early screening. Furthermore, the throat swab sample volume can be determined within a 15% fluctuation range, making it easier to control sample volume deviations compared to coughed-out sampling. The present invention is applicable not only to airway sampling but also to various sampling sites accessible to throat swab samplers, such as those in gynecology, facilitating daily self-testing. B) Enhanced readability of test results. When the method and kit of the present invention are used for detection, hydrogen peroxide samples as low as 0.8-1.5mM can be clearly colored, and hydrogen peroxide samples with a concentration higher than 1.5mM can be 100% clearly colored, while the color development rate of 0.7mM hydrogen peroxide or 3mM sodium nitrite or 3mM ferric chloride or 3mM copper sulfate is 0, that is, 100% no color. Therefore, the present invention can not only achieve good hydrogen peroxide detection sensitivity, but also ensure good specificity. Compared with the CN 2022115399358 method, the present invention can better control the background of 0.7mM hydrogen peroxide and more clearly indicate the difference between 1.5mM and 0.7mM hydrogen peroxide samples. C) Improved economic practicality. The test kit provided by CN 2022115399358 requires customized sampling tubes and production processes. The test kit provided by the present invention uses standard colloidal gold cartridges and polyethylene screw-cap tubes, which simplifies the production process, reduces packaging, is compatible with the colloidal gold POCT automated assembly line, greatly improves production throughput, reduces the production cost and packaging and transportation cost of the test kit, extends the shelf life of the product, and enables the product to maintain stable quality for a long time under hot conditions, which is conducive to the popularization and promotion of the product.

[0265] 3. Compared with traditional exhaled breath testing. Due to the solubility and diffusivity of hydrogen peroxide, the vast majority of hydrogen peroxide in the airway dissolves in the airway mucus, with only a trace amount being distributed in the exhaled breath. In addition, the airway mucus sampling of the present invention can eliminate airflow interference compared to conventional exhaled breath sampling. Therefore, the present invention has better anti-interference ability, is more sensitive, more specific, and easier to use.

[0266] 4. Compared with conventional colorimetric or fluorescence methods. These conventional methods require professional pipetting operations, the drawing of standard curves, and the dilution of samples into the linear range before measurement. The use process of the present invention is an integrated operation that does not require quantification and does not require additional equipment and facilities. The color threshold of the colorimetric area has eliminated background interference, and the brown color of the quality control area is usually weaker than the yellow-brown color of the positive color of the colorimetric area, which is convenient for comparison to determine the positive result. Therefore, the present invention is conducive to qualitative and intuitive judgment, and is simpler, faster, and more suitable for popularization.

[0267] 5. Compared with various blood testing methods. Numerous blood testing methods for tumors exist, with varying blood requirements and performance. However, widespread application is limited by the need for blood draws. This present invention does not require blood draws, offers superior safety and non-invasiveness, and can more sensitively and directly indicate the hydrogen peroxide metabolism status at the sampling site than blood testing.

[0268] 6. Compared with various DNA and protein detection methods. Since DNA has good stability, not only chemical stability but also genetic stability, the detection of tumor DNA is usually a relatively long-term correlation test and cannot reflect the occurrence and development of tumors in a timely manner. Different proteins have different half-lives, but they can usually be up to several months or even longer. Therefore, they cannot reflect the dynamic changes related to tumors in a very timely manner. The present invention is based on the limited half-life of hydrogen peroxide, has good dynamics, is suitable for indicating dynamic changes related to tumors, and is expected to develop into a powerful means of screening and monitoring various lung cancer-related screenings. The present invention can help potential patients detect the risk of lung cancer as early as possible and receive relevant examinations such as CT / PET-CT in a timely manner, so as to facilitate early diagnosis and treatment and restore health.

[0269] In summary, the present invention specifically solves the problem of insufficient convenience and ease of use in hydrogen peroxide detection. It has the advantages of convenience and ease of use, reliable performance, and moderate price. It is conducive to popularization and application, and is suitable for various hydrogen peroxide detections that can be sampled using a throat swab sampler.

[0270] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A hydrogen peroxide rapid detection kit, characterized in that: include: A sample diluent, the sample diluent comprising 100-500 mM ammonium oxalate, 0.3-2.0 mM 8-hydroxyquinoline, 50-500 mM sodium chloride and 10-100 mM first pH adjuster with a pH of 4.5-7.0; Dry the test strips with the detection color developer and the quality control color developer.

2. The kit according to claim 1, characterized in that The first pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate or 2-(N-morpholino)ethanesulfonic acid.

3. The kit according to claim 1, characterized in that The detection color developing agent includes 2.0%-8.0% (w / v) starch, 0.5-5.0 mM acetic acid, 0.5%-2.0% (w / v) Brij-35, 1.0-7.0 M potassium iodide, 2.0-10.0 mM 8-hydroxyquinoline, 50-300 mM calcium chloride, 5-100 mM second pH adjusting agent and 3-15 mM L-ascorbic acid or sodium thiosulfate.

4. The kit according to claim 3, characterized in that: The second pH adjuster includes at least one of sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium acetate, sodium bicarbonate or tris(hydroxymethyl)aminomethane.

5. The kit according to claim 1, characterized in that: The quality control color developing reagent includes 5-30 mM neutral red and 10-60 mM sodium bicarbonate.

6. The kit according to claim 1, characterized in that The test strip also includes an intermediate line reagent.

7. The kit according to claim 6, characterized in that The intermediate line reagent includes 5%-15% (w / v) Brij-35 and 5%-15% (w / v) sodium dodecyl sulfate.

8. The kit according to any one of claims 1 to 7, characterized in that: The test strip has a size of 68mm×6mm×0.7mm, 3μL of detection colorimetric agent is dried on one end of the test strip, 3μL of quality control colorimetric agent is dried on the other end of the test strip, and 3μL of intermediate line reagent is dried between the detection colorimetric agent position and the quality control colorimetric agent position of the test strip.

9. The kit according to claim 8, characterized in that The sample diluent includes 500 mM ammonium oxalate, 2 mM 8-hydroxyquinoline, 450 mM sodium chloride, and 20 mM sodium dihydrogen phosphate-disodium hydrogen phosphate at pH 6.

0.

10. The kit according to claim 8, characterized in that The detection color developing agent includes 2% (w / v) starch, 1 mM acetic acid, 250 mM calcium chloride, 2% (w / v) Brij-35, 2M potassium iodide, 5 mM 8-hydroxyquinoline, 15 mM sodium thiosulfate and 10 mM sodium bicarbonate.

11. The kit according to claim 8, characterized in that The intermediate line reagent includes 10% (w / v) Brij-35 and 10% (w / v) SDS.

12. The kit according to claim 8, characterized in that The quality control color developing reagent includes 15 mM neutral red and 45 mM sodium bicarbonate.

13. A method for detecting hydrogen peroxide, characterized in that: The following steps are involved: a. Take the sample to be tested and add it to the sample diluent in the kit as claimed in any one of claims 1 to 12 to obtain a mixed solution; b. Pipette the mixed solution into the sample application area of ​​the test strip in the kit according to any one of claims 1 to 12 and let it stand until the test strip develops color.

14. The detection method according to claim 13, characterized in that: a includes: after sampling with a throat swab sampler, immersing the sampling end into a sampling tube with a built-in sample diluent, breaking it along the notch, leaving the sampling end in the tube, sealing the sampling tube, and shaking the sampling tube so that the sampling end is completely immersed in the sample diluent to obtain a mixed solution one.

15. The detection method according to claim 13, characterized in that: b includes: adding one drop of the mixed solution to the sample addition area of ​​the test strip, leaving it to stand until the quality control color developer area fully changes color to purple-red, and observing the color development in the detection color developer area.

16. Use of the kit according to any one of claims 1 to 7 or the detection method according to claims 8 to 15 in detecting lung cancer.

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