CN114544572A - BNP fluorescent quantitative detection test strip, kit and preparation method thereof - Google Patents

BNP fluorescent quantitative detection test strip, kit and preparation method thereof Download PDF

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Publication number
CN114544572A
CN114544572A CN202210161195.2A CN202210161195A CN114544572A CN 114544572 A CN114544572 A CN 114544572A CN 202210161195 A CN202210161195 A CN 202210161195A CN 114544572 A CN114544572 A CN 114544572A
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bnp
pad
detection
sample
membrane
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蒋鑫
王佩瑞
白婷
郑翠
徐思桐
杨博
伍健聪
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Shanghai I Reader Biological Technology Co ltd
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/531—Production of immunochemical test materials
    • G01N33/532—Production of labelled immunochemicals
    • G01N33/533—Production of labelled immunochemicals with fluorescent label
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/536—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids

Abstract

The invention discloses a BNP fluorescence quantitative detection test strip, a kit and a preparation method thereof, relating to the technical field of medical in-vitro assistance. The device comprises a combination pad, a sample pad, a detection membrane and an absorption pad which are sequentially arranged on a substrate along a chromatography direction; the binding pad is coated with fluorescent microspheres for marking BNP detection antibodies, and the detection membrane is respectively provided with a detection line for coating anti-BNP capture antibodies and a quality control line for coating X anti-YIgG antibodies. When the binding pad is coated with the fluorescent microspheres for marking the BNP detection antibody, the binding pad is arranged in front of the sample pad along the chromatography direction, so that the chromatography distance of the conjugate of the BNP antigen and the BNP detection antibody-fluorescent microspheres on the whole test strip can be increased, the full reaction of the BNP antigen and the BNP detection antibody is facilitated, the antibody on the subsequent detection membrane can be better combined with the BNP antigen which is chromatographed to the detection membrane with high affinity, and the detection sensitivity is improved.

Description

BNP fluorescent quantitative detection test strip, kit and preparation method thereof
Technical Field
The invention relates to the technical field of medical in-vitro assistance, in particular to a BNP fluorescence quantitative detection test strip, a kit and a preparation method thereof.
Background
Type B Natriuretic Peptide (BNP), a member of the family of natriuretic peptides that are secreted primarily by the heart, is a polypeptide consisting of 32 amino acid residues. BNP is the most valuable biomarker for diagnosing heart failure at present, and can be used for diagnosing various heart failure types, including asymptomatic heart failure, chronic decompensated heart failure and acute decompensated heart failure. At present, BNP (brain natriuretic peptide) less than 100pg/mL is recommended, can be used for the diagnosis of excluding acute compensatory heart failure and has higher negative predictive value; BNP is more than or equal to 35-100 pg/mL and is determined as a diagnosis threshold value of the heart failure, and the BNP is more than 400pg/mL, and the diagnosis of the heart failure is considered.
At present, methods for quantitatively detecting BNP include radioimmunoassay, enzyme-linked immunosorbent assay, chemiluminescence assay, and lateral flow immunochromatography. Compared with other methods, the lateral flow chromatography has the advantages of low cost, simple and quick operation, convenient carrying and the like, and is an important part in the field of medical in-vitro diagnosis. Meanwhile, in view of the significance of BNP for assisting in diagnosing heart failure, the development of the BNP immunofluorescence quantitative detection test strip with high sensitivity and high precision has a wide market value.
The necessary conditions that the BNP immunofluorescence quantitative detection test strip on the market can reach an extremely low detection limit at present are as follows: the antibody on the fiber membrane is used for marking a conjugate of biotin and fluorescein streptavidin, or marking fluorescein, quantum dots and the like, and the high-sensitivity detection is realized by a method of signal amplification and the like.
In view of this, the invention is particularly proposed.
Disclosure of Invention
The invention aims to provide a BNP immunofluorescence quantitative test strip independent of chemiluminescence and fluorescein.
The invention is realized by the following steps:
the invention provides a BNP fluorescent quantitative detection test strip which comprises a combination pad, a sample pad, a detection membrane and an absorption pad which are sequentially arranged on a substrate along a chromatography direction; the binding pad is coated with fluorescent microspheres for marking BNP detection antibodies, and the detection membrane is respectively provided with a detection line for coating anti-BNP capture antibodies and a quality control line for coating X anti-YIgG antibodies, wherein X is selected from sheep, chicken, horse, monkey, mouse, rabbit or pig, Y is selected from sheep, chicken, horse, monkey, mouse, rabbit or pig, and X is different from Y.
The inventor creatively discovers that when the binding pad is coated with the fluorescent microspheres for marking the BNP detection antibody, the chromatography distance of the conjugate of the BNP antigen and the BNP detection antibody-fluorescent microspheres on the whole test strip can be increased along the chromatography direction before the binding pad is arranged in front of the sample pad, so that the BNP antigen and the BNP detection antibody can fully react, the antibody on a subsequent detection membrane can be better combined with the BNP antigen which is chromatographed to the detection membrane with high affinity, and the detection sensitivity is improved.
The inventors surprisingly found that the combination pad and the sample pad sequentially arranged on the substrate along the chromatography direction can reduce the variation coefficient of the test value of the test strip, thereby being beneficial to improving the precision of the test strip and improving the test precision in batches or among batches.
In an alternative embodiment, the antibody coated on the quality control line is a goat anti-mouse IgG antibody.
The fluorescent microspheres include, but are not limited to, polystyrene microspheres, such as green, colored, orange, blue, etc. The particle size is 50-400 nm.
In a preferred embodiment of the present invention, the number of the sample pads is 1, and the lower edge of the bonding pad overlaps the upper edge of one end of the sample pad, and the lower edge of the other end of the sample pad overlaps the upper edge of the detection film.
The inventor finds that the test strip with the lap joint structure has the technical advantages of high detection signal intensity and small variation coefficient of the detection signal compared with other test strips with the lap joint structure, the variation coefficient is less than 10% at least, and the precision is obviously improved. In addition, the detection limit of the detection test strip provided by the invention is extremely low (reaching 10pg/mL), and reaches or exceeds the detection level after the cascade amplification of the conventional chemiluminescence and fluorescein. The linear detection range is 15-5000 pg/mL.
In a preferred embodiment of the present invention, the number of the sample pads is 1, the upper edge of the bonding pad overlaps the lower edge of one end of the sample pad, and the lower edge of the other end of the sample pad overlaps the upper edge of the detection membrane.
In a preferred embodiment of the present invention, the number of the sample pads is plural, and the lower edge of the combination pad overlaps the upper edge of one end of the first sample pad, the lower edge of the other end of the first sample pad overlaps the upper edge of one end of the second sample pad, and the lower edge of the last sample pad overlaps the upper edge of the detection membrane.
It should be noted that the overlapping may be equivalent to partial overlapping, and there is partial overlap in the structure.
In an alternative embodiment, the number of the sample pads includes, but is not limited to, 2, 3 or 4.
In a preferred embodiment of the present invention, the number of the sample pads is plural, and the lower edge of the combination pad overlaps the lower edge of one end of the first sample pad, the upper edge of the other end of the first sample pad overlaps the lower edge of one end of the second sample pad, and the lower edge of the last sample pad overlaps the upper edge of the detection membrane.
In an alternative embodiment, the number of the sample pads includes, but is not limited to, 2, 3 or 4.
In a preferred embodiment of the present invention, the bonding pad is made of glass fiber, polyester fiber or filter paper; the detection membrane is a nitrocellulose membrane or an acetate fiber membrane.
In a preferred embodiment of the present invention, the anti-BNP capture antibody coated on the detection line is an anti-BNP capture antibody solution, and the coating concentration is 2 mg/mL.
The capture antibody against BNP may be selected from commercially available antibodies or antibodies produced by hybridoma cells. The capture antibody of the anti-BNP and the BNP detection antibody have different BNP antigen binding epitopes.
In a preferred embodiment of the present invention, the X-anti-yig antibody coated on the quality control line is an X-anti-yig antibody solution, and the coating concentration is 1.2-2 mg/mL.
The invention also provides a preparation method of the BNP fluorescence quantitative detection test strip, which comprises the following steps: bonding a bonding pad, a sample pad, a detection membrane and an absorption pad on a substrate along the chromatography direction;
in an optional embodiment, the preparation method further comprises the steps of setting a detection line and a quality control line on the detection membrane before bonding; the preparation method also comprises the step of preparing a bonding pad before bonding, wherein the preparation of the bonding pad comprises the steps of soaking the glass fiber membrane, the polyester cellulose membrane or the filter paper in the bonding pad treatment solution of the fluorescent microspheres marked with the BNP detection antibody, and drying.
The adhesive may be an adhesive having tackiness.
The material of the sample pad includes, but is not limited to, glass fiber or polyester.
The above preparation method further comprises cutting the test strip into test strips of 3.2mm width by a cutter. And (5) putting the cut test strip into a plastic card, and pressing.
The invention also provides a BNP fluorescent quantitative detection kit which comprises the BNP fluorescent quantitative detection test strip. In other embodiments, the test kit further comprises a sample diluent.
The invention has the following beneficial effects:
the inventor finds that when the binding pad is coated with the fluorescent microspheres for marking the BNP detection antibody, the binding pad is arranged in front of the sample pad along the chromatography direction, so that the chromatography distance of the conjugate of the BNP antigen and the BNP detection antibody-fluorescent microspheres on the whole test strip can be increased, the full reaction of the BNP antigen and the BNP detection antibody is facilitated, the subsequent antibody on the detection membrane can be better combined with the BNP antigen which is chromatographed to the detection membrane with high affinity, and the detection sensitivity is improved. The inventors surprisingly found that the combination pad and the sample pad sequentially arranged on the substrate along the chromatography direction can reduce the variation coefficient of the test value of the test strip, thereby being beneficial to improving the precision of the test strip and improving the test precision in batches or among batches.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.
FIG. 1 is a schematic diagram of a test strip structure.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.
The features and properties of the present invention are described in further detail below with reference to examples.
In the following examples, the distance from the leftmost end of each set of test strips to the leftmost end of the NC membrane was the same, so the distance of the chromatography run by the sample was the same.
Example 1
The embodiment provides a BNP fluorescent quantitative test strip, which is shown in fig. 1, and in fig. 1: firstly, a sample pad is formed; ② a combination pad; ③ is a nitrocellulose membrane; and fourthly, the absorbent pad.
The preparation method comprises the following steps:
(1) preparation of nitrocellulose membrane containing capture antibody: respectively fixing the antibody membrane-scribing liquid (from salt wild meaning) containing the captured BNP antigen and the membrane-scribing liquid containing the goat anti-mouse IgG antibody (from Arista) on a nitrocellulose membrane to be respectively used as a test line (T) and a quality control line (C), and placing the nitrocellulose membrane and the test line (T) in a vacuum drying pump for drying for 2 hours for later use.
(2) Preparation of sample pad: and (3) selecting a commercial sample pad or soaking the glass fiber membrane in the sample pad treatment solution, placing the sample pad in a vacuum drying pump, and drying for 2 hours for later use.
(3) Preparation of the bonding pad: soaking the glass fiber membrane in a binding pad treatment solution containing fluorescent microspheres labeled with BNP detection antibody (from wild sense of salt), placing in a vacuum drying pump, and drying for 2h for later use.
(4) Sequentially sticking on a PVC bottom plate: conjugate pad, sample pad, nitrocellulose membrane and absorbent pad (i.e. conjugate pad-sample pad (lower edge of conjugate pad overlaps upper edge of one end of sample pad) -nitrocellulose membrane-absorbent pad) and cut into test strips of 3.2mm width using a cutter. The absorbent pad was purchased from GE.
(5) And (5) putting the cut test strip into a plastic card, and pressing. In other embodiments, a plastic card or housing may not be provided.
Example 2
Compared with example 1, the material is the same, the difference is only in the number of sample pads and the overlapping structure, and the structure is shown in fig. 1.
In this embodiment, along the chromatography direction, there are sequentially provided: sample pad-conjugate pad (both ends of conjugate pad are located under sample pad) -sample pad-nitrocellulose membrane-absorbent pad.
Example 3
Compared with example 1, the material is the same, the difference is only in the number of sample pads and the overlapping structure, and the structure is shown in fig. 1.
In this embodiment, along the chromatography direction, there are sequentially provided: conjugate pad-sample pad (part of the sample pad is below the conjugate pad) -sample pad (part of the sample pad is below the upper sample pad) -nitrocellulose membrane-absorbent pad.
In other embodiments, the number of sample pads may be adaptively adjusted according to the need, and is not limited to the two defined in this embodiment.
Example 4
Compared with the embodiment 1, the material is the same, the difference is only in the overlapping structure, and the structure is shown in the reference figure 1.
In this embodiment, along the chromatography direction, there are sequentially provided: conjugate pad-sample pad (top of conjugate pad lap joint with bottom of one end of sample pad, bottom of other end of sample pad lap joint with top of nitrocellulose membrane) -nitrocellulose membrane-absorbent pad.
Example 5
Compared with example 1, the material is the same, the difference is only in the number of sample pads and the overlapping structure, and the structure is shown in fig. 1.
In this embodiment, along the chromatography direction, there are sequentially provided: conjugate pad-sample pad (part of the sample pad is above the conjugate pad) -sample pad (part of the sample pad is above the upper sample pad) -nitrocellulose membrane-absorbent pad.
In other embodiments, the number of sample pads may be adaptively adjusted according to the need, and is not limited to the two defined in this embodiment.
Comparative example 1 (i.e., control group)
Compared with the embodiment 1, the material is the same, the difference is only in the overlapping structure, and the structure is shown in the reference figure 1.
In this embodiment, along the chromatography direction, there are sequentially provided: sample pad-conjugate pad (lower side of sample pad overlaps upper side of one end of conjugate pad, lower side of the other end of conjugate pad overlaps upper side of nitrocellulose membrane) -nitrocellulose membrane-absorbent pad.
The sample pad-conjugate pad length refers to the length of the sample pad-conjugate pad region from the left end to the right end of the test strip.
The sample pad-conjugate pad lengths of examples 1-5 and control 1 were all consistent and 20 mm. The test strips of examples 1-5 and comparative example 1 were 3.2mm in width; the length and width of the nitrocellulose membrane are both consistent and are 25mm and 3.2mm respectively.
It should be noted that: the lengths of the reagent strip conjugate pad-sample pad regions of the various embodiments are, in structural order (from left to right): control 1 (i.e., control) (19.5mm,5 mm); example 1(10mm,12 mm); example 2(10mm,7mm,8 mm); example 3(10mm,10mm,8 mm); example 4(10mm, 12 mm); example 5 (10mm,10mm,8 mm).
Experimental example 1
Using the BNP fluorescent quantitative test strips prepared in examples 1-5 and comparative example 1 to detect a sample solution (salt wild-sense BNP antigen) with a BNP concentration of 100pg/mL, each group of test strips was tested 10 times repeatedly, and the test line (T) signal, the quality control line (C) signal and the total signal were counted respectively. The results are given in table 1 below:
TABLE 1100 pg/mL BNP antigen sample solution test results table.
Figure BDA0003514774830000081
The results show that:
compared with the test strip with the traditional structure (i.e. the comparative example 1), the test strip structures prepared in the examples 1, 2, 3, 4 and 5 can improve the mean value of the T signal and the detection signal value.
From the results of 10 replicates, the coefficient of variation of the detected signal in example 1 was only 9.5%, which is the most excellent.
Experimental example 2
In this example, the detection limit and precision of comparative example 1 and example 1 were verified, respectively, by the following methods:
BNP antigen solutions of 0, 10, 50, 100, 200, 400, 800, 1600, 3200, 5500pg/mL were tested using the fluorescent quantitative BNP test strips prepared in comparative example 1 and example 1, respectively, and the test was repeated 3 times at each concentration to calculate an average value, as shown in table 2 below. A standard curve was made using four parameter fitting.
Table 2 mean values of the detection signals at each concentration.
Figure BDA0003514774830000091
Four parameter fitting (equation: y ═ (A-D)/[1+ (x/C) was used)B]+ D) respectively performing standard curve fitting on the signal mean values in the table 2, wherein the obtained standard curve equation parameter information is as follows:
Figure BDA0003514774830000092
verifying the detection limit: testing the BNP antigen solution with the concentration of 0pg/mL for 20 times to obtain a blank limit (mean value +2 SD); and testing the BNP antigen solution with the concentration of 10pg/mL for 25 times of repetition, and if no more than 3 test values in the 25 times of repeated test values are lower than the blank limit, considering the detection limit to be 10 pg/mL.
And (3) precision verification: the BNP antigen solution of 100 and 800pg/mL is tested, 10 times of each test is carried out for each concentration, and the variation coefficient of each concentration test value is calculated.
The detection results are shown in table 3 below, and compared with comparative example 1, the detection sensitivity of example 1 is significantly improved, and the detection limit is 10 pg/mL.
Table 3 table of verification limit results.
Group of Detection limit (number less than or equal to blank limit)
Comparative example 6
Example 1 0
As shown in Table 4, the precision of example 1 was significantly improved compared to that of comparative example 1, and the coefficients of variation of BNP antigen solutions of 100 and 800pg/mL were all less than 10%.
Table 4 table of precision verification results.
Figure BDA0003514774830000101
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims (10)

1. A BNP fluorescence quantitative detection test strip is characterized by comprising a combination pad, a sample pad, a detection membrane and an absorption pad which are sequentially arranged on a substrate along a chromatography direction; the binding pad is coated with fluorescent microspheres for marking BNP detection antibodies, and the detection membrane is respectively provided with a detection line for coating anti-BNP capture antibodies and a quality control line for coating X anti-YIgG antibodies, wherein X is selected from sheep, chickens, horses, monkeys, mice, rabbits or pigs, Y is selected from sheep, chickens, horses, monkeys, mice, rabbits or pigs, and X is different from Y.
2. The BNP fluorescence quantitative detection test strip of claim 1, wherein the number of the sample pads is 1, and the lower edge of the combination pad overlaps the upper edge of one end of the sample pad, and the lower edge of the other end of the sample pad overlaps the upper edge of the detection membrane.
3. The BNP fluorescence quantitative detection test strip of claim 1, wherein the number of the sample pads is 1, and the upper edge of the combination pad overlaps the lower edge of one end of the sample pad, and the lower edge of the other end of the sample pad overlaps the upper edge of the detection membrane.
4. The BNP fluorescence quantitative detection test strip of claim 1, wherein the number of the sample pads is plural, and the lower edge of the combination pad overlaps the upper edge of one end of the first sample pad, the lower edge of the other end of the first sample pad overlaps the upper edge of one end of the second sample pad, and the lower edge of the last sample pad overlaps the upper edge of the detection membrane;
preferably, the number of sample pads is 2, 3 or 4.
5. The BNP fluorescent quantitative detection test strip of claim 1, wherein the number of the sample pads is plural, and the lower edge of the combination pad is overlapped with the lower edge of one end of the first sample pad along the chromatographic direction, the upper edge of the other end of the first sample pad is overlapped with the lower edge of one end of the second sample pad, and the lower edge of the last sample pad is overlapped with the upper edge of the detection membrane;
preferably, the number of sample pads is 2, 3 or 4.
6. The BNP fluorescence quantitative detection test strip of any one of claims 1 to 5, wherein the material of the bonding pad is glass fiber, polyester cellulose membrane or filter paper; the detection membrane is a nitrocellulose membrane or an acetate fiber membrane.
7. The BNP fluorescent quantitative detection test strip according to any one of claims 1 to 5, wherein the anti-BNP capture antibody coated on the detection line is an anti-BNP capture antibody solution, and the coating concentration is 2 mg/mL.
8. The BNP fluorescent quantitative detection test strip of any one of claims 1 to 5, wherein the X anti-YIgG antibody coated on the quality control line is X anti-YIgG antibody liquid, and the coating concentration is 1.2mg/mL to 2 mg/mL.
9. The preparation method of the BNP fluorescence quantitative detection test strip of any one of claims 1 to 8, which comprises the following steps: bonding a bonding pad, a sample pad, a detection membrane and an absorption pad on a substrate along the chromatography direction;
preferably, the preparation method further comprises the steps of setting a detection line and a quality control line on the detection film in a point mode before bonding; the preparation method further comprises the step of preparing a binding pad before bonding, wherein the preparation of the binding pad comprises the steps of soaking the glass fiber membrane, the polyester cellulose membrane or the filter paper in the binding pad treatment solution of the fluorescent microspheres marked with the BNP detection antibody, and drying.
10. A BNP fluorescence quantitative determination kit, characterized in that it comprises the BNP fluorescence quantitative determination test strip of any one of claims 1 to 8.
CN202210161195.2A 2022-02-22 2022-02-22 BNP fluorescent quantitative detection test strip, kit and preparation method thereof Pending CN114544572A (en)

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