Test strips and their applications

Abstract

The present invention provides an albumin detection test strip and its application. The detection test strip includes a reaction layer adhered to and supported on a base plate, and an anti-interference layer is covered on the other side of the reaction layer relative to the base plate. The reaction layer is made of glass fiber material. The anti-interference layer is made of cotton fiber material. The present invention can effectively eliminate the false positive phenomenon when detecting albumin in high specific gravity urine or high pH urine by combining the reaction layer made of glass fiber material and the anti-interference layer made of cotton fiber material, thereby ensuring stable and accurate test results. The detection test strip of the present invention can be used for detecting urine component analysis and applied to urine combined detection test strips.

Description

Technical Field

[0001] The present invention relates to a test paper for detecting protein content in a liquid sample and an application thereof, in particular to a test paper for detecting protein content in urine and an application thereof. Background Art

[0002] Urinary microalbumin (MA, defined as a urinary albumin excretion rate of 20-200 μg / min or a 24-hour total urinary albumin content of 33-300 mg) can serve as a biochemical marker for pathological responses in diabetes and nephropathy. Furthermore, MA is also found in essential hypertension and can predict the development of cardiovascular disease. Therefore, clinical screening for urinary microalbumin is of great significance.

[0003] Currently, there are four main methods for screening for microalbuminemia: Method 1: random urine collection for A / C measurement; Method 2: 24-hour urine albumin excretion rate and simultaneous 24-hour urine creatinine measurement for calculation of creatinine clearance; Method 3: regular sample collection for albumin measurement; and Method 4: dry chemical test strips. While simple, Method 1 is expensive and unsuitable for clinical screening. Methods 2 and 3 are time-consuming. Dry chemical test strips are simple, rapid, and inexpensive to use for albumin determination.

[0004] Dry chemical test strips utilize the protein error principle of indicators. This principle states that when a pH indicator binds to a protein (particularly albumin), the pKa value of the indicator changes (generally increases). In a specific buffer system, the color of the test strip changes with the protein concentration in the test sample. Sulfonphthalein compounds are commonly used as indicators.

[0005] Conventional protein content testing (150-1000 mg / L) typically uses tetrabromophenolsulfonphthalein as an indicator and fibrous filter paper as a carrier for the assay, resulting in a sensitivity ranging from 150-300 mg / L. To improve sensitivity when measuring trace albumin, an improved sulfonphthalein indicator, such as bis(3',3"-iodo-5',5"-dinitrobenzene)-3,4,5,6-tetrabromophenolsulfonphthalein (DDPTP), is employed. This improved indicator, combined with fibrous filter paper as a carrier, increases sensitivity to approximately 80 mg / L.

[0006] However, a sensitivity of 80 mg / L still cannot meet clinical needs. When urine output is high, an albumin level of 30 mg / L is also considered an abnormal clinical result. Therefore, it is necessary to further improve the sensitivity of the test paper. Another way to improve sensitivity is to change the composition of the reagent carrier, using glass fiber instead of cotton fiber. Due to its unique structure, the carrier made of glass fiber can effectively filter protein. When the micro-albumin product made of glass fiber is immersed in the sample, the albumin in the sample is blocked by the surface of the glass fiber test paper and cannot enter the interior of the test paper and be evenly distributed like cotton fiber, thereby achieving the effect of enriching the measured substance on the test paper surface. In this way, the surface of the test paper can obtain a darker color reaction, thereby improving the sensitivity of the product. When glass fiber is used as the test paper carrier, its sensitivity can reach 30 mg / L. As can be seen from the comparative experiment in Example 1, when measuring micro-albumin in urine, the sensitivity of the test is higher when glass fiber is used as a reagent carrier than when cotton fiber is used as a reagent carrier.

[0007] However, while glass fiber can effectively improve the sensitivity of microalbumin detection, it cannot provide sufficient pH buffering for the microalbumin reaction. For samples with normal specific gravity (1.005-1.020) and pH (5.0-8.0), accurate and reliable test results can be obtained; however, for samples with high specific gravity (≥1.025), high pH (≥9.0), or high specific gravity (≥1.025) and high pH (≥9.0) coexisting, the test results will be much higher than the actual results. This requires a method to provide sufficient pH buffering to reduce the impact of high specific gravity or high pH samples on the test results. Summary of the Invention

[0008] The object of the present invention is to provide a test paper for detecting microalbumin, which can effectively reduce the influence of high specific gravity or high pH on the test results.

[0009] The test paper of the present invention comprises a base plate 11 on which a reaction layer 12 is adhered and supported, and an anti-interference layer 13 is covered on the other side of the reaction layer relative to the base plate. The base plate 11 of the present invention is made of a material that is impermeable to liquid samples, such as polypropylene, polystyrene, or polysulfone plastic sheets. The reaction layer 12 of the present invention is made of glass fiber material. The anti-interference layer 13 is made of cotton fiber material.

[0010] The beneficial effect of the present invention is that the combination of the reaction layer made of glass fiber material and the anti-interference layer made of cotton fiber material can effectively eliminate the influence of high specific gravity or high pH samples on albumin detection, thereby ensuring stable and accurate detection results. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1This is a schematic diagram of the structure of the first test paper of the present invention;

[0012] Figure 2 This is a schematic diagram of the decomposition of the second test paper of the present invention; DETAILED DESCRIPTION

[0013] The present invention will be described in detail below with reference to the accompanying drawings. These specific embodiments are merely limited examples without departing from the spirit of the present invention, and do not exclude other specific implementation plans that can be produced by combining the prior art with the present invention by those skilled in the art.

[0014] like Figure 1 As shown, the test paper of the present invention includes a base plate 11 and a reaction layer 12. The reaction layer 12 contains a detection reagent for receiving a liquid sample and reacting with the liquid sample to obtain a test result. An anti-interference layer 13 is covered on the other side of the reaction layer 12 relative to the base plate 11.

[0015] like Figure 2 As shown, the anti-interference layer is wider than the reactive layer, the middle portion of the anti-interference layer is above the reactive layer, and the end pieces 14 at both ends are overlapped on the bottom plate. The overlapped ends of the anti-interference layer are adhered to the bottom plate by hot pressing.

[0016] The bottom plate 11 of the present invention is made of a material that is impermeable to liquid samples, and the material is selected from polypropylene, polystyrene, polysulfone, etc.

[0017] The reaction layer 12 of the present invention includes a chemical reagent fixed on the reaction layer, and the chemical reagent reacts with the analyte in the sample to generate a result signal. The material of the reaction layer is selected from a suitable material such as glass fiber.

[0018] During the detection of microalbumin, the chemical reagents on the reaction layer include solution I in Table 1 and solution II in Table 2.

[0019] Table 1 Solution I

[0020] drug Dosage per 1000ml solution purified water 1000ml Tartaric acid buffered saline 0.3mol, pH 2.1

[0021] Table 2 Solution II

[0022] drug Dosage per 1000ml solution Toluene 1000ml DDPTP 1.5g PPG P2000 25g

[0023] DDPTP is bis(3,3'-iodo-5,5'-dinitrobenzene)-3,4,5,6-tetrabromophenolsulfonphthalein, and PPG P2000 is polypropylene glycerol P2000.

[0024] The material selected for the anti-interference layer 13 of the present invention can quickly diffuse and penetrate the added sample, such as cotton fiber and other suitable materials. The anti-interference layer can also be treated with a hydrophilic solution.

[0025] The thickness of the anti-interference layer is less than 0.07 mm, more preferably 0.03 mm.

[0026] The liquid samples mentioned in this specification and claims include liquid samples and samples dissolved in liquid.

[0027] Example 1 Comparison of Sensitivity of Glass Fiber and Cotton Fiber for Determination of Urine Microalbumin Concentration

[0028] Glass fiber Whatman GF / F filter paper (thickness between 0.3 mm and 0.5 mm) and cotton fiber Whatman 3MM filter paper (thickness between 0.3 mm and 0.5 mm) were immersed in solution I, dried in a 70°C oven for 25 minutes, and then immersed in solution II and dried in a 70°C oven for 15 minutes.

[0029] Solution I

[0030] drug Dosage per 1000ml solution purified water 1000ml Tartaric acid buffered saline 0.3mol, pH 2.1

[0031] Solution II

[0032] drug Dosage per 1000ml solution Toluene 1000ml DDPTP 1.5g PPG P2000 25g

[0033] DDPTP is bis(3,3"-iodo-5,5"-dinitrobenzene)-3,4,5,6-tetrabromophenolsulfonphthalein, and PPG P2000 is polypropylene glycerol P2000.

[0034] The above test strips were cut into 5mm×5mm squares and affixed to plastic cards. They were then immersed in urine with albumin contents of 10mg / L, 30mg / L, 80mg / L, and 150mg / L, a specific gravity of 1.015, and a pH of 7.0, calibrated using an Abbott C8000 fully automatic biochemical analyzer. The test strips were removed and excess solution was removed. After allowing the sample to sit for 1 minute, the reaction results were scanned at 520nm using a SpectroCam microfiber spectrometer from Avantes, the Netherlands, to measure the reflectivity. The results are shown in Table 3 below.

[0035] Table 3

[0036]

[0037] In Example 1, glass fiber or cotton fiber alone was used as a detection reagent carrier. The test results showed that when detecting trace albumin in urine with a normal specific gravity and pH value, the test strip using glass fiber alone as the carrier could clearly distinguish between samples with relatively low albumin concentrations, such as 10 mg / L and 30 mg / L, and thus could detect low-concentration albumin in urine. However, when cotton fiber was used as the detection reagent carrier, the test results could not clearly distinguish between samples with relatively low albumin concentrations, such as 10 mg / L and 30 mg / L, and therefore could not provide accurate results. Therefore, cotton fiber is not used as a detection reagent carrier in existing products.

[0038] Example 2 Effect of the Anti-interference Layer on Eliminating the Effect of High Specific Gravity or High pH of Urine

[0039] Glass fiber Whatman GF / DVA filter paper was immersed in Solution I, dried in a 70°C oven for 25 minutes, then immersed in Solution II. After removal, it was dried in a 70°C oven for 15 minutes to form a reactive layer. Cotton fiber Schoeller & Hoesch Grade 11 / LC 33 filter paper (0.03 mm thick) was immersed in Solution I and dried in a 70°C oven for 15 minutes to form an anti-interference layer.

[0040] Solution I

[0041] drug Dosage per 1000ml solution purified water 1000ml Tartaric acid buffered saline 0.3mol, pH 2.1

[0042] Solution II

[0043] drug Dosage per 1000ml solution Toluene 1000ml DDPTP 1.5g PPG P2000 25g

[0044] DDPTP is bis(3,3"-iodo-5,5"-dinitrobenzene)-3,4,5,6-tetrabromophenolsulfonphthalein, and PPG P2000 is polypropylene glycerol P2000.

[0045] The reaction layer was cut into 5mm wide strips and affixed to a plastic card. The anti-interference layer was then cut into 15-20mm wide strips and placed over the reaction layer, with both sides affixed to a larger card. The plastic card with only the reaction layer and the card with both the reaction layer and anti-interference layer were then cut into 5mm wide strips. The test strips were then immersed in urine solutions of varying specific gravities and pH values, calibrated to 10mg / L, 30mg / L, 80mg / L, and 150mg / L albumin levels on an Abbott C8000 fully automated biochemical analyzer. The test strips were removed and excess solution was removed. After allowing the solution to sit for 1 minute, the reaction results were scanned at 520nm using a SpectroCam microfiber spectrometer from Avantes (Netherlands), and the reflectance was measured. Using a normal urine sample with a pH of 7.0 and a specific gravity of 1.015 as a reference, the test results are shown in Table 4 below.

[0046] Table 4

[0047]

[0048] The results of Example 2 show that the test strip without the anti-interference layer produces significantly higher results than normal when detecting albumin in urine with a high pH or high specific gravity, resulting in a false positive. The test strip with the anti-interference layer also produces significantly higher results than normal when detecting albumin in urine with a high pH or high specific gravity, effectively eliminating the effects of high specific gravity or pH, eliminating false positives, and improving the accuracy of albumin detection.

[0049] Example 3 Effects of Anti-interference Layers of Different Thicknesses on Eliminating the Effects of High Specific Gravity or High pH of Urine

[0050] Whatman GF / DVA test paper was immersed in Solution I, oven-dried at 70°C for 25 minutes, then immersed in Solution II and oven-dried at 70°C for 15 minutes to form a reactive layer. Schoeller & Hoesch Grade 11 / LC 33 filter paper (0.03 mm) and Schweitzer Mauduit 260M4T filter paper (0.07 mm) of varying thickness were immersed in Solution I and oven-dried at 70°C for 15 minutes to form an anti-interference layer.

[0051] Solution I

[0052] drug Dosage per 1000ml solution purified water 1000ml Tartaric acid buffered saline 0.3mol, pH 2.1

[0053] Solution II

[0054] drug Dosage per 1000ml solution Toluene 1000ml DDPTP 1.5g PPG P2000 25g

[0055] DDPTP is bis(3,3"-iodo-5,5"-dinitrobenzene)-3,4,5,6-tetrabromophenolsulfonphthalein, and PPG P2000 is polypropylene glycerol P2000.

[0056] Cut the reaction layer into 5mm wide strips and stick them on a plastic card. Then cut the anti-interference layer into 15-20mm wide strips and cover the reaction layer, and stick both sides to the large card. Then cut the cards with the reaction layer and anti-interference layer into 5mm wide strips respectively. Immerse them in urine with an albumin content of 10mg / L, 30mg / L, 80mg / L, and 150mg / L calibrated by the Abbott C8000 fully automatic biochemical analyzer, a specific gravity of 1.015, and a pH of 7.0. Take out the test paper and absorb the excess solution. After leaving it naturally for 1 minute, use the miniature fiber optic spectrometer SpectroCam from Avantes of the Netherlands to scan the reaction results at 520nm and measure the reflectivity. The results are shown in Table 5 below

[0057] Table 5

[0058]

[0059] The measurement results of Example 3 show that the thickness of the anti-interference layer will affect the test results of microalbumin.

Claims

1. A protein detection test strip comprising a base plate (11) and a reaction layer (12) located on the base plate, characterized in that: An anti-interference layer (13) is covered on the other side of the reaction layer (12) relative to the bottom plate. The material of the reaction layer (12) is glass fiber, the material of the anti-interference layer (13) is cotton fiber, and the thickness of the anti-interference layer (13) is less than 0.07 mm.

2. The test paper according to claim 1, wherein: The reaction layer (12) was treated with solution I and solution II, respectively. The solution I consisted of 0.3 mol / 1000 ml of pH 2.1 tartaric acid buffer and 1000 ml / 1000 ml of purified water, and the solution II consisted of 1000 ml / 1000 ml of toluene, 1.5 g / 1000 ml of DDPTP, and 25 g / 1000 ml of PPG P2000.

3. The test paper according to claim 1, wherein: The anti-interference layer (13) is treated with a solution I consisting of 0.3 mol / 1000 ml of pH 2.1 tartaric acid buffer and 1000 ml / 1000 ml of purified water.

4. The test paper according to claim 1, wherein: The width of the anti-interference layer is greater than that of the reaction layer. The middle portion of the anti-interference layer is above the reaction layer, and both ends of the anti-interference layer are overlapped on the bottom plate.

5. The test paper according to claim 1, wherein: The protein is albumin.

6. The test paper according to claim 1, wherein: The cotton fiber is selected from Schoeller & Hoesch Grade 11 / LC 33 filter paper; the glass fiber is selected from Whatman GF / DVA filter paper.

7. The test paper according to claim 1, wherein: The thickness of the anti-interference layer (13) is 0.03 mm.

8. The test paper according to any one of claims 1 to 7 is used for detecting protein content.

9. The use according to claim 8, characterized in that The protein is albumin.

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