Test strip for rapid quantitative detection of tissue cell protein

Abstract

The present invention relates to a test piece for rapid and quantitative detection of tissue cell proteins. Specifically, the present invention provides a test piece for detecting tissue cell proteins, the test piece comprising a substrate, and a sample addition area, a colloidal gold area, a protein detection area and a water absorption area are sequentially provided on the substrate from one end to the other end of the substrate; the protein detection area comprises a plurality of detection points, the detection points are arranged in a plurality of columns, and the centers of the detection points in each column are interconnected to form a straight line, and the straight line is perpendicular to the flow direction of the sample to be tested; the detection points on the protein detection area present an alternating array. The test piece of the present invention can rapidly and accurately qualitatively and quantitatively detect tissue cell proteins.

Description

Technical Field

[0001] The present invention relates to the field of detection, and in particular to a test piece for rapid quantitative detection of tissue cell protein. Background Art

[0002] Previous tumor treatments, especially chemotherapy, had a wide range of drug effects. While killing tumor cells, they also killed a large number of normal cells, leading to unbearable side effects. With the rapid progress of medical research, scientists have gradually developed tumor targeted therapeutic drugs with strong targeting and low toxicity and side effects. At present, this targeted therapy has become an increasingly popular trend in tumor treatment due to its good therapeutic effect and low toxicity and side effects.

[0003] Surgery or biopsy will accelerate the infiltration and spread of tumor cells. The medical community unanimously recognizes that timely and effective treatment is a necessary means to prevent its spread. However, how to quickly and accurately determine the molecular type of a patient's tumor in clinical practice and then determine whether the patient is suitable for a certain targeted treatment plan is still a difficult problem that the medical community urgently needs to solve.

[0004] Therefore, there is an urgent need in the art to develop a method that can quickly and quantitatively detect tissue cell protein levels, especially cell protein levels associated with tumor cells, to provide guidance for the treatment of diseases. Summary of the invention

[0005] The object of the present invention is to provide a test piece capable of quickly and quantitatively detecting tissue cell proteins, especially cell proteins associated with tumor cells.

[0006] In a first aspect of the present invention, a test piece for detecting tissue cell protein is provided, wherein the test piece comprises a substrate, and a sample addition area, a colloidal gold area, a protein detection area and a water absorption area are sequentially provided on the substrate from one end to the other end of the substrate;

[0007] The protein detection area includes a plurality of detection points, the detection points are arranged in a plurality of columns, the centers of the detection points in each column are connected to form a straight line, and the straight line is perpendicular to the flow direction of the sample to be tested;

[0008] The detection spots on the protein detection zone are arranged in an alternating array.

[0009] In another preferred embodiment, the detection points are arranged in n+1 columns, wherein n=3-30, preferably 3-20, more preferably 3-15, more preferably 5-12, and n is a positive integer (such as 1, 2, 3, 4, 9, 15 or 30, etc.).

[0010] In another preferred embodiment, the shapes of the detection points are the same or different.

[0011] In another preferred embodiment, the shape of the detection point is circular.

[0012] In another preferred embodiment, the inner diameters of the detection points are the same or different.

[0013] In another preferred embodiment, the distance between the centers of two adjacent detection points in each column is the same.

[0014] In another preferred embodiment, the number of detection points contained in each column is 2-30, preferably 2-20, more preferably 2-10, and most preferably 2-6.

[0015] In another preferred embodiment, the number of detection points in each column differs by 1-2.

[0016] In another preferred embodiment, the straight lines formed by connecting the centers of the circles of each column of detection points are parallel to each other.

[0017] In another preferred embodiment, the vertical distance between two adjacent columns is 1.0-1.8 mm, preferably 1.2-1.6 mm, and more preferably 1.3-1.5 mm. In another preferred embodiment, the distance between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween is equal or unequal.

[0018] In another preferred embodiment, the distance between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap between the two adjacent detection points in the column is 1.3-2.1 mm, preferably 1.4-1.8 mm, and more preferably 1.6-1.8 mm.

[0019] In another preferred embodiment, the distance between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap between the two adjacent detection points is equal or unequal.

[0020] In another preferred example, two adjacent detection points D1 and D2 in one column, the gap between D1 and D2 corresponds to a detection point D3 in another adjacent column, wherein the distance between the centers of the two detection points D1 and D3 is 1.3-2.1mm, preferably 1.4-1.8mm, more preferably 1.6-1.8mm; the distance between the centers of the two detection points D2 and D3 is 1.3-2.1mm, preferably 1.4-1.8mm, more preferably 1.6-1.8mm.

[0021] In another preferred embodiment, an equilateral triangle is formed between two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap between the two adjacent detection points in the column.

[0022] In another preferred embodiment, the inner diameter of each detection point is 0.5-1.2 mm, preferably 0.6-1.0 mm, and more preferably 0.7-0.9 mm.

[0023] In another preferred embodiment, the distance between the centers of two adjacent detection points in each column is 1.3-2.1 mm, preferably 1.4-1.8 mm, and more preferably 1.6-1.8 mm.

[0024] In another preferred embodiment, the distance between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween is 1.3-2.1 mm, preferably 1.4-1.8 mm, and more preferably 1.6-1.8 mm.

[0025] In another preferred embodiment, the test piece further includes a quality control area located on the substrate.

[0026] In another preferred embodiment, the quality control area is located between the protein detection area and the water absorption area.

[0027] In another preferred embodiment, the colloidal gold region is loaded with one or more complexes selected from the following group: colloidal gold antibody complex, colloidal silver antibody complex;

[0028] The colloidal gold-antibody complex is formed by coupling colloidal gold with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof;

[0029] The colloidal silver antibody complex is formed by coupling colloidal silver with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof.

[0030] In another preferred embodiment, the colloidal gold region is provided with glass fiber, and the composite is coated on the glass fiber.

[0031] In another preferred embodiment, the tissue cell protein to be detected is HER2 protein.

[0032] In another preferred embodiment, the colloidal gold region is also loaded with IgG colloidal gold complex, IgG colloidal silver complex, or a combination thereof.

[0033] In another preferred embodiment, the detection points of the protein detection zone are loaded with one or more antibodies selected from the following group: monoclonal antibodies against the tissue protein to be detected, and polyclonal antibodies against the tissue protein to be detected.

[0034] In another preferred embodiment, the colloidal gold area includes a colloidal gold complex loaded with HER2 antibodies, and the detection point includes an anti-HER2 antibody.

[0035] In another preferred embodiment, the protein detection area is provided with a nitrocellulose membrane, and the monoclonal antibody or polyclonal antibody against the tissue protein to be detected is coated on the glass fiber.

[0036] In another preferred embodiment, the HER2 antibody in the HER2 antibody colloidal gold complex is a rabbit-derived anti-human HER2 antibody.

[0037] In another preferred embodiment, the sample adding area includes a sample adding pad.

[0038] In another preferred embodiment, the water absorbing area includes absorbent paper.

[0039] In a second aspect, the present invention provides a kit for detecting tissue cell protein, the kit comprising:

[0040] (i) a test piece for detecting tissue cell protein as described in the first aspect of the present invention;

[0041] (ii) Instructions for use.

[0042] In another preferred embodiment, the instruction manual records the use of the test piece for detecting tissue cell protein as described in the first aspect of the present invention to detect tissue cell protein.

[0043] The third aspect of the present invention provides a use of the test piece as described in the first aspect of the present invention in detecting tissue cell proteins.

[0044] In another preferred embodiment, the detection is an in vitro detection.

[0045] In another preferred embodiment, the detection includes qualitative and / or quantitative detection.

[0046] In another preferred embodiment, the detection is an auxiliary detection.

[0047] In another preferred embodiment, the detection is a non-therapeutic and non-diagnostic detection.

[0048] A fourth aspect of the present invention provides a method for detecting tissue cell protein, the method comprising using the test piece for detecting tissue cell protein according to the first aspect of the present invention to detect tissue cell protein in a sample to be tested.

[0049] In another preferred embodiment, the detection is an in vitro detection.

[0050] In another preferred embodiment, the detection includes qualitative and / or quantitative detection.

[0051] In another preferred embodiment, the detection is an auxiliary detection.

[0052] In another preferred embodiment, the detection is a non-therapeutic and non-diagnostic detection.

[0053] In another preferred embodiment, the sample includes cell lysate, tissue lysate, plasma, serum, whole blood, urine or sputum.

[0054] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described below (such as embodiments) can be combined with each other to form a new or preferred technical solution. Due to space limitations, they will not be described one by one here. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] Figure 1 Schematic diagram of the structure of the test piece of Example 1.

[0056] Figure 2 It is the positive color development of HER2 protein standard solution of different concentrations at the protein detection point of the test piece. According to the sequence of numbers 1-5, the concentrations of HER2 standard protein are 4ng / ml, 20ng / ml, 100ng / ml, 500ng / ml and 2500ng / ml.

[0057] Figure 3 The standard curve drawn for different concentrations of HER2 protein standard solution and its total absorbance value is as follows: Figure 2 As shown, the correlation coefficient R 2 =0.99.

[0058] Figure 4 The results are from immunohistochemistry.

[0059] Figure 5 Determine the results for Western blot detection.

[0060] Figure 6 The results of HER2 protein expression detection of different subjects by the test piece of Example 1 are shown.

[0061] Figure 7 The test results of the test piece of Example 1 and the test piece of Comparative Example 1 for the HER2 protein standard solution of the same concentration, wherein the left side is the test result of the parallel array test point of Comparative Example 1, and the right side is the test result of the alternating array test point of Example 1 DETAILED DESCRIPTION

[0062] After extensive and in-depth research, the inventor unexpectedly discovered a test piece capable of rapid and quantitative detection of tissue cell proteins through a large amount of screening and exploration. In the test piece, the detection points on the protein detection area present a specific arrangement. The detection piece can perform rapid and accurate quantitative detection of tissue cell proteins. On this basis, the inventor completed the present invention.

[0063] the term

[0064] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0065] As used herein, the terms "comprise", "include", and "contain" are used interchangeably and include not only closed definitions, but also semi-closed and open definitions. In other words, the terms include "consisting of", "consisting essentially of".

[0066] As used in this article, the term "HER2 protein" and "proto-oncogene human epidermal growth factor receptor-2 (HER2)" are used interchangeably. HER2 refers to the target protein for targeted therapy of breast cancer. HER2-positive (overexpressed or amplified) breast cancer has a good therapeutic effect on targeted drugs such as Herceptin that target HER2, and the treatment model is also very different from that of other types of breast cancer.

[0067] Test piece

[0068] The present invention provides a test piece for detecting tissue cell protein, wherein the test piece can perform rapid qualitative and quantitative detection on the level of tissue cell protein, especially rapid qualitative and quantitative detection on the level of cell protein associated with tumor cells.

[0069] The test piece of the present invention may also be referred to as a reagent strip or a test card.

[0070] Typically, the test sheet comprises a substrate, and from one end to the other end of the substrate, a sample addition area, a colloidal gold area, a protein detection area and a water absorption area are sequentially provided on the substrate;

[0071] The protein detection area includes a plurality of detection points, the detection points are arranged in a plurality of columns, the centers of the detection points in each column are connected to form a straight line, and the straight line is perpendicular to the flow direction of the sample to be tested;

[0072] The detection spots on the protein detection zone are arranged in an alternating array.

[0073] In the present invention, it should be understood that the alternating array means that the detection points in two adjacent columns are arranged alternately. In other words, the alternating array means that the gap between two adjacent detection points in one column corresponds to a detection point in another adjacent column.

[0074] In the present invention, the substrate may also be referred to as a backing, a substrate, etc., which plays a supporting role.

[0075] In the present invention, the flow direction of the sample to be tested is the direction from the sample addition area to the water absorption area, and is perpendicular to the straight line formed by connecting the centers of each column of detection points.

[0076] Sample loading area

[0077] In the test piece of the invention, the sample adding area is used for adding the sample to be tested. The sample adding area may include a sample adding pad. Preferably, the sample adding pad is a fiber membrane.

[0078] In another preferred embodiment, the sample to be tested may include cell lysate, tissue lysate, plasma, serum, whole blood, urine or sputum. Preferably, the sample to be tested contains tissue cell protein, such as HER2 protein.

[0079] Colloid area

[0080] In the test piece of the invention, the colloidal gold area may contain a gold reaction pad.

[0081] The colloidal gold region may be loaded with one or more complexes selected from the following group: colloidal gold antibody complex, colloidal silver antibody complex.

[0082] In another preferred embodiment, the colloidal gold-antibody complex is formed by coupling colloidal gold with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof.

[0083] In another preferred embodiment, the colloidal silver-antibody complex is formed by coupling colloidal silver with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof.

[0084] In another preferred embodiment, the tissue cell protein to be detected is HER2 protein.

[0085] In another preferred embodiment, the colloidal gold region is also loaded with IgG colloidal gold complex.

[0086] In another preferred embodiment, the colloidal gold region is provided with glass fiber, and the composite is coated on the glass fiber.

[0087] Protein detection area

[0088] In the test piece of the invention, the specific structure of the protein detection area enables the test piece to perform rapid qualitative and quantitative detection of tissue cell protein levels.

[0089] In the present invention, the shape of the detection point is not particularly limited as long as the purpose of the present invention is met. A preferred shape of the detection point includes (but is not limited to): circle, square, or a combination thereof. More preferably, the shape of the detection point is circle.

[0090] In this article, it should be understood that if the shape of the detection point is not a circle, when defining features such as the inner diameter of the detection point, the detection point needs to be simulated into a circle. The specific simulation process is: take the two farthest points on the shape of the detection point as the diameter and simulate it into a circle.

[0091] In the present invention, the inner diameter of the detection point refers to the diameter of the detection point. For example, when the detection point is circular, the inner diameter refers to the diameter of the circular detection point.

[0092] In a preferred embodiment of the present invention, the inner diameter of each detection point is 0.5-1.2 mm, preferably 0.6-1.0 mm, and more preferably 0.7-0.9 mm.

[0093] In another preferred embodiment, the distance between the centers of two adjacent detection points in each column is 1.3-2.1 mm, preferably 1.4-1.8 mm, and more preferably 1.6-1.8 mm.

[0094] In another preferred embodiment, the distance between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween is 1.3-2.1 mm, preferably 1.4-1.8 mm, and more preferably 1.6-1.8 mm.

[0095] In another preferred embodiment, the detection points are arranged in n+1 columns, wherein n=3-30, preferably 3-20, more preferably 3-15, more preferably 5-12, and n is a positive integer (such as 1, 2, 3, 4, 9, 15 or 30, etc.).

[0096] In another preferred embodiment, the vertical distance between two adjacent columns is 1.0-1.8 mm, preferably 1.2-1.6 mm, and more preferably 1.3-1.5 mm. In the present invention, the vertical distance between two adjacent columns refers to the vertical distance between the straight lines formed by connecting the centers of the detection points of the two adjacent columns. In other words, the "vertical distance between two adjacent columns" refers to the vertical distance between the straight line (L1) formed by connecting the centers of the detection points of one column and the straight line (L2) formed by connecting the centers of the detection points of another adjacent column, that is, the vertical distance between the straight line (L1) and the straight line (L2).

[0097] Quality Control Area

[0098] The test piece of the present invention may further include a quality control area on the substrate. The quality control area may be provided with a quality control line, preferably, the quality control line is perpendicular to the flow direction. Preferably, the quality control area is located between the protein detection area and the water absorption area.

[0099] The quality control line can be used to indicate whether the detection process (such as the chromatography process) is normal. For example, in the test piece of the present invention, the sample addition area is coated with a colloidal gold complex of IgG, and the quality control line is coated with an antibody of the IgG. The IgG and the antibody of the IgG play a role of quality control, indicating whether a certain detection process (chromatography process) is normal; that is, if the IgG can reach the quality control line normally through chromatography, the antibody of IgG on the quality control line binds to the IgG, thereby developing color; and if the test card has expired or been damaged, the IgG cannot reach the quality control line normally, and the color cannot be developed.

[0100] Therefore, those skilled in the art can select IgG for use in the present invention at their own discretion. In a specific embodiment, the IgG includes, but is not limited to, rabbit IgG, horse IgG, mouse IgG, and the like. Accordingly, those skilled in the art can select IgG antibodies for use in the present invention at their own discretion, as long as the IgG antibodies are antibodies to the IgG. In a specific embodiment, the IgG antibodies include, but are not limited to, goat anti-rabbit IgG, goat anti-horse IgG, and the like.

[0101] In the present invention, the sample adding area, the colloidal gold area, the protein detection area and the water absorption area are located on the substrate.

[0102] Water absorption area

[0103] In the present invention, the water absorption area may include absorbent paper. The water absorption area drives the sample to be tested to flow from the sample addition area through the colloidal gold area and the protein detection area in sequence to the water absorption area, thereby ensuring the smooth progress of the detection.

[0104] The main advantages of the present invention include:

[0105] 1. The test piece of the present invention can quickly and accurately perform qualitative and quantitative determination of tissue cell proteins (such as cancer cell tissue protein HER2) with high sensitivity, greatly improving the effect of disease diagnosis and treatment.

[0106] 2. The existing technology mainly detects tissue cell proteins (such as cancer cell tissue protein HER2) through high performance liquid chromatography-tandem mass spectrometry, high performance liquid chromatography, biochip technology and other detection methods, but these technologies have obvious defects, such as complex operation, expensive equipment, time-consuming, unsuitable for on-site detection, etc. In addition, the methods for detecting tissue cell proteins (such as cancer cell tissue protein HER2) in the prior art have high requirements for instruments and operators, and require a long time, resulting in high detection costs. The test piece described in the present invention is easy to operate, low in cost and fast in determination, which overcomes the complex, time-consuming and labor-intensive instrument detection in the prior art.

[0107] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples without specifying specific conditions are usually based on conventional conditions or the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are calculated by weight.

[0108] Example 1

[0109] Test piece

[0110] The structural diagram of the test piece is as follows Figure 1 As shown, the test sheet includes a substrate, and from one end to the other end of the substrate, a sample pad, a colloidal gold area, a protein detection area and a blotting paper are sequentially arranged on the substrate. After the sample to be tested is added to the sample pad, it flows through the colloidal gold area, the protein detection area and the blotting paper in sequence;

[0111] Colloidal gold area: The colloidal gold area is loaded with HER2 antibody colloidal gold complex, which is coated on glass fiber;

[0112] Protein detection area: The protein detection area contains multiple detection points, and the detection points are loaded with anti-HER2 mouse antibodies fixed on the nitrocellulose membrane; all detection points have the same shape and are circular, and the detection points are arranged in multiple columns, and the centers of the detection points in each column are connected to form a straight line, and the straight line is perpendicular to the flow direction of the sample to be tested. The detection points on the protein detection area are presented in an alternating array, and the distance between the centers of two adjacent detection points in each column is the same, and the straight lines formed by the centers of the detection points in each column are parallel to each other. The inner diameter d1 of each detection point is 0.8mm, and the distance d2 between the centers of adjacent detection points in each column is 1.66mm. The distances between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap between them are d3 and d4, respectively, d3=1.66mm and d4=1.66mm.

[0113] Example 2

[0114] Quantitative detection of HER2 protein

[0115] The test piece of Example 1 was used to detect HER2 protein standard solutions of different concentrations. The specific detection method is as follows: the HER2 protein standard was diluted in phosphate buffer in proportion to 4ng / ml, 20ng / ml, 100ng / ml, 500ng / ml and 2500ng / ml. Then 100μl of HER2 protein standard dilutions of different concentrations were added to the sample addition area of ​​the test piece. After 5 minutes, the positive color development of the protein detection point of the test piece of HER2 protein standard solutions of different concentrations was as follows: Figure 2 As shown, in Figure 2In the sequence 1-5, the concentrations of HER2 standard protein are 4ng / ml, 20ng / ml, 100ng / ml, 500ng / ml, and 2500ng / ml. The number of positive color detection points and their absorbance values ​​of each test piece are counted by an absorbance scanning detector, and the total absorbance value (Ab525nm) is calculated. The total absorbance value is the sum of the absorbance values ​​of all detection points for 525nm wavelength light, as shown in Table 1. Then, a standard curve is drawn based on different concentrations of HER2 protein standard solutions and the total absorbance value (Ab525nm). The standard curve is as shown in Table 1. Figure 3 shown.

[0116] Table 1 Correspondence between HER2 concentration and total absorbance value

[0117]

[0118]

[0119] A=0.68C+22.10, correlation coefficient R 2 =0.9966

[0120] From Table 1 and Figure 3 It can be seen from the standard curve that the test piece of Example 1 can accurately and quantitatively measure the HER2 protein standard solution. Therefore, the test piece of Example 1 can be used to quantitatively detect the concentration of HER2 protein, and the accuracy of the quantitative result is high.

[0121] Example 3

[0122] Example 3: The expression of HER2 protein in breast cancer of 5 subjects was determined by immunohistochemistry, protein blotting and the test piece of Example 1, and the accuracy of the test piece of Example 1 in detecting HER2 protein expression in breast cancer was compared. Among them, immunohistochemistry is a routine clinical detection method, and protein blotting is used as a third-party gold standard. In the immunohistochemistry, protein blotting and detection methods of Example 1, HER2 protein (labeled as 1, 2, 3, 4 and 5 respectively) in breast cancer tissues of 5 subjects was determined, and specimens with the same serial number were all derived from breast cancer tissues of the same subject.

[0123] Immunohistochemistry

[0124] The postoperative tumor specimens were fixed with formaldehyde and embedded in paraffin; after sectioning, they were dewaxed, hydrated, permeabilized, and blocked, and then mouse HER2 antibodies were added, and finally incubated with goat anti-mouse IgG secondary antibodies labeled with horseradish peroxidase, and finally color developed and filmed under a microscope. The darker the color, the higher the expression of the detected protein. Specimens of the same tissue from different subjects were tested by immunohistochemistry, and the test results were as follows: Figure 4As shown:

[0125] Gold standard western blot detection

[0126] Tumor specimens were homogenized and broken with SDS lysis buffer to extract total protein. After determining the protein concentration, the same mass of total protein was taken, heated and denatured, separated by 8% SDS-polyacrylamide gel, and transferred to a 0.45μm nitrocellulose membrane. Finally, it was blocked, incubated with primary antibody and fluorescein-labeled secondary antibody, and scanned for imaging. Specimens from the same tissue from different subjects were tested by Western blotting. The results of Western blotting are as follows Figure 5 As shown:

[0127] Test piece detection of Example 1

[0128] The test strip of Example 1 was used to detect the HER2 protein expression level in breast cancer tissues of 5 subjects. The specific detection method is as follows: After cutting about 0.1g of fresh breast cancer tissue into pieces, add 250 microliters of protein lysis solution and grind it to completely lyse it, then heat it at 95 degrees for 5 minutes and quickly cool it on ice. Then take 50 microliters of tissue lysis solution and add it to the sample area of ​​the test strip. After 5 minutes, the HER2 protein expression detection results of different subjects are as follows: Figure 6 shown.

[0129] from Figure 4-6 It can be seen that there are certain differences between the test results of the test piece of Example 1 and the traditional immunohistochemistry test results, but are consistent with the third-party gold standard (protein blotting test) results. The test results of specimens 4 and 5 show that the trend of immunohistochemistry is obviously inconsistent with the results of the gold standard protein blotting test, but the test results of the test piece of Example 1 are highly consistent with the protein blotting test results, indicating that the detection accuracy of the test piece of Example 1 is higher than that of traditional immunohistochemistry detection. It can be seen that the values ​​of the test piece of Example 1 between different specimens are more consistent with the results of protein blotting detection, indicating that the detection accuracy of the test piece of Example 1 is higher than that of traditional immunohistochemistry detection.

[0130] Comparative Example 1

[0131] Comparative Example 1 provides a test piece, which is different from Example 1 in that the detection points on the protein detection area are in a parallel array.

[0132] According to the method for quantitatively detecting HER2 protein in Example 2, the test results of the test piece in Example 1 and the test piece in Comparative Example 1 for the HER2 protein standard solution of the same concentration are as follows: Figure 7 shown.

[0133] from Figure 7It can be seen that the number of positive color development detection points of the test piece of comparative example 1 is significantly smaller than the number of positive color development protein detection points of the test piece of embodiment 1. The more color development detection points there are, the better the accuracy and sensitivity of the determination structure. The test piece of embodiment 1 is significantly better than the test piece of comparative example 1. The total absorbance values ​​obtained by the absorbance scanning detector are 2170 and 2930, respectively (the larger the absorbance value, the higher the accuracy and sensitivity). The total absorbance value is the sum of the absorbance values ​​of all detection points to light of 525nm wavelength.

[0134] All documents mentioned in the present invention are cited as references in this application, just as each document is cited as reference individually. In addition, it should be understood that after reading the above teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the claims attached to this application.

Claims

1. A test piece for detecting tissue cell protein, characterized in that: The test sheet comprises a substrate, and a sample adding area, a colloidal gold area or a colloidal silver area, a protein detection area and a water absorption area are sequentially arranged on the substrate from one end to the other end of the substrate; The protein detection area includes a plurality of detection points, the detection points are arranged in a plurality of columns, the centers of the detection points in each column are connected to form a straight line, and the straight line is perpendicular to the flow direction of the sample to be tested; The detection points on the protein detection area are in an alternating array, wherein the alternating array means that the gap between two adjacent detection points in one column corresponds to a detection point in another adjacent column; The inner diameter of each detection point is 0.9-1.2 mm, the distance between the centers of two adjacent detection points in each column is 1.4-1.8 mm, and, The vertical distance between two adjacent columns is 1.0-1.8 mm, wherein the vertical distance between two adjacent columns refers to the vertical distance between straight lines formed by connecting the centers of the detection points of the two adjacent columns.

2. The test piece according to claim 1, wherein: The detection points are arranged in n+1 columns, where n=3-30, and n is a positive integer.

3. The test piece according to claim 2, characterized in that: n=3-20, n is a positive integer.

4. The test piece according to claim 1, wherein: The number of detection points contained in each column ranges from 2 to 30.

5. The test piece according to claim 4, characterized in that: The number of detection points contained in each column is 2-20.

6. The test piece according to claim 4, characterized in that: Each column contains 2 to 10 detection points.

7. The test piece according to claim 1, wherein: The inner diameter of each detection point is 1.0-1.2mm.

8. The test piece according to claim 1, wherein: The distance between the centers of two adjacent detection points in each column is 1.6-1.8 mm.

9. The test piece according to claim 1, wherein: The distance between the centers of two adjacent detection points in each column is the same.

10. The test piece according to claim 1, wherein: The distances between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween are 1.3-2.1 mm respectively.

11. The test piece according to claim 10, wherein: The distances between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween are 1.4-1.8 mm, respectively.

12. The test piece according to claim 10, wherein: The distances between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween are 1.6-1.8 mm respectively.

13. The test piece according to claim 1, wherein: The vertical distance between two adjacent columns is 1.2-1.6 mm.

14. The test piece according to claim 13, wherein: The vertical distance between two adjacent columns is 1.3-1.5 mm.

15. The test piece according to claim 1, wherein: The distances between the centers of two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap therebetween are equal or unequal.

16. The test piece according to claim 1, wherein: An equilateral triangle is formed between two adjacent detection points in one column and a detection point in another adjacent column corresponding to the gap between the two adjacent detection points in the column.

17. The test piece according to claim 1, wherein: The test piece also includes a quality control area located on the substrate.

18. The test piece according to claim 1, wherein: The colloidal gold region or colloidal silver region is loaded with one or more complexes selected from the following group: colloidal gold antibody complex, colloidal silver antibody complex; The colloidal gold-antibody complex is formed by coupling colloidal gold with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof; The colloidal silver antibody complex is formed by coupling colloidal silver with an antibody selected from the following group: a monoclonal antibody to the tissue cell protein to be detected, a polyclonal antibody to the tissue cell protein to be detected, or a combination thereof.

19. The test piece according to claim 1, wherein: The detection points in the protein detection zone are loaded with one or more antibodies selected from the following group: monoclonal antibodies against the tissue protein to be detected, and polyclonal antibodies against the tissue protein to be detected.

20. The test piece according to claim 1, wherein: The tissue cell protein to be detected is HER2 protein.

21. A kit for detecting tissue cell protein, characterized in that: The kit comprises: (i) a test piece for detecting tissue cell protein according to any one of claims 1 to 20; (ii) Instructions for use.

22. Use of the test piece according to any one of claims 1 to 20 in preparing a kit for detecting tissue cell protein.

23. An in vitro, non-therapeutic and non-diagnostic method for detecting tissue cell proteins, characterized in that: The method comprises using the test piece for detecting tissue cell protein according to any one of claims 1 to 20 to detect tissue cell protein in the sample to be tested.

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