阅读说明：本技术 一种甲胎蛋白免疫层析试纸条及其制备方法和应用 (Alpha fetoprotein immunochromatographic test strip and preparation method and application thereof ) 是由 冯涛 陈国动 周慧 张楠 朱红甜 于 2019-11-28 设计创作，主要内容包括：本发明提供一种甲胎蛋白免疫层析试纸条及其制备方法和应用,所述甲胎蛋白免疫层析试纸条包括样品垫、结合垫、硝酸纤维素膜和吸水垫和背衬板,所述结合垫上结合有复合荧光微球探针,所述复合荧光微球探针为甲胎蛋白单克隆抗体与碳纳米管以及水溶性量子点形成的微球探针,所述硝酸纤维素膜上检测线和质控线,其中检测线上含有甲胎蛋白抗体,所述质控线上含有羊抗鼠IgG。本发明的甲胎蛋白免疫层析试纸条制备方法简单、易操作,所述甲胎蛋白免疫层析试纸条用于甲胎蛋白检测荧光强度显著增强,检测准确,灵敏度明显增强,最低检测限为0.02ng/mL。(The invention provides an alpha fetoprotein immunochromatographic test strip and a preparation method and application thereof, the alpha fetoprotein immunochromatographic test strip comprises a sample pad, a combination pad, a nitrocellulose membrane, a water absorption pad and a backing plate, wherein a composite fluorescent microsphere probe is combined on the combination pad, the composite fluorescent microsphere probe is a microsphere probe formed by an alpha fetoprotein monoclonal antibody, a carbon nano tube and water-soluble quantum dots, a detection line and a quality control line are arranged on the nitrocellulose membrane, the detection line contains an alpha fetoprotein antibody, and the quality control line contains goat anti-mouse IgG. The alpha fetoprotein immunochromatographic test strip is simple in preparation method and easy to operate, the fluorescence intensity of the alpha fetoprotein detection is obviously enhanced, the detection is accurate, the sensitivity is obviously enhanced, and the minimum detection limit is 0.02 ng/mL.)

1. The utility model provides an alpha fetoprotein immunochromatographic test strip, which is characterized in that, alpha fetoprotein immunochromatographic test strip includes sample pad, combination pad, cellulose nitrate membrane and water absorption pad and backing plate, it has compound fluorescence microsphere probe to combine on the combination pad, compound fluorescence microsphere probe is the microsphere probe that alpha fetoprotein monoclonal antibody and carbon nanotube and water-soluble quantum dot formed, last detection line and the quality control line of cellulose nitrate membrane, wherein contain the alpha fetoprotein antibody on the detection line, contain sheep anti mouse IgG on the quality control line.

2. The alpha fetoprotein immunochromatographic test strip of claim 1, wherein the carbon nanotubes are multi-walled carbon nanotubes or single-walled carbon nanotubes.

3. The alpha fetoprotein immunochromatographic test strip according to claim 1 or 2, wherein the water-soluble quantum dots are carboxyl-coated CdTe quantum dots or carboxyl-coated CdTe/ZnSe core-shell quantum dots.

4. The alpha fetoprotein immunochromatographic test strip according to any one of claims 1 to 3, wherein the preparation method of the composite fluorescent microsphere probe is as follows: incubating aminated carbon nanotubes and alpha fetoprotein monoclonal antibodies at 37 ℃ under the action of a condensing agent, centrifuging, redissolving precipitates after centrifugation, adding the condensing agent and carboxylated water-soluble quantum dots, incubating at 37 ℃, and centrifuging to obtain the composite fluorescent microsphere probe.

5. The alpha fetoprotein immunochromatographic test strip according to claim 4, wherein when the aminated carbon nanotube and the alpha fetoprotein monoclonal antibody are incubated at 37 ℃ under the action of a condensing agent, the condensing agent used is any one of 1-ethyl- (3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and N-hydroxythiosuccinimide or a combination of at least two of the 1-ethyl- (3-dimethylaminopropyl) carbodiimide and the N-hydroxythiosuccinimide.

6. The alpha fetoprotein immunochromatographic test strip according to claim 4 or 5, wherein the incubation time of the aminated carbon nanotube and the alpha fetoprotein monoclonal antibody at 37 ℃ under the action of the condensing agent is 30min-2 h.

7. The immunochromatographic strip for alpha fetoprotein according to any one of claims 4 to 6, wherein when a condensing agent and a carboxylated water-soluble quantum dot are added, the condensing agent is any one of 1-ethyl- (3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and N-hydroxythiosuccinimide or a combination of at least two of the above.

8. The alpha fetoprotein immunochromatographic test strip according to any one of claims 4 to 7, wherein the condensing agent and the carboxylated water-soluble quantum dot are added, and the incubation time at 37 ℃ is 30min-2 h.

9. The method for preparing the alpha fetoprotein immunochromatographic test strip according to any one of claims 1 to 8, comprising the steps of:

spraying the composite fluorescent microsphere probe on a bonding pad to obtain the bonding pad sprayed with the composite fluorescent microsphere probe, then respectively scribing the alpha fetoprotein antibody and goat anti-mouse IgG on a nitrocellulose membrane to obtain a detection line and a quality control line, and assembling a sample pad, the bonding pad sprayed with the composite fluorescent microsphere probe, the nitrocellulose membrane with the detection line and the quality control line and a water absorption pad on a back plate to obtain the alpha fetoprotein immunochromatography test strip;

preferably, the scribing is performed with a scribe.

10. Use of the alpha-fetoprotein immunochromatographic test strip according to any one of claims 1 to 8 in the preparation of an alpha-fetoprotein detection device.

Technical Field

The invention belongs to the technical field of immunodetection, and relates to an alpha fetoprotein immunochromatography test strip, and a preparation method and application thereof.

Background

The primary liver cancer is one of the most common clinical malignant tumors, the incidence rate of the primary liver cancer is on the rise in the world, and the method has important significance for the accurate diagnosis of the primary liver cancer and the treatment of the liver cancer.

Alpha fetoprotein is a specific clinical index for diagnosing primary liver cancer, currently, immunoassay is mainly adopted for detecting the alpha fetoprotein, and CN103558396A discloses a quantitative detection method of the alpha fetoprotein, which comprises the following steps: (1) mixing the alpha fetoprotein antibody with a hydroformylation glucoamylase solution to prepare an enzyme-labeled antibody; loading the alpha fetoprotein antibody on the nano gold magnetic particles; (2) adding the alpha-fetoprotein antigen sample and a series of alpha-fetoprotein antigen standard samples with different concentrations into an immunoreaction interface of the antibody-loaded nano gold magnetic particles for incubation, measuring the glucose concentration by a glucometer to obtain a linear equation of the corresponding relation between the alpha-fetoprotein concentration and the glucose concentration, and substituting the glucose concentration value of the alpha-fetoprotein antigen sample into the linear equation to calculate the alpha-fetoprotein concentration of the alpha-fetoprotein antigen sample.

CN108709914A discloses a method for rapidly detecting alpha fetoprotein, which comprises the steps of coating a single-arm carbon nanotube alpha fetoprotein antibody dispersion liquid on a paper base material, determining the resistance of a paper-based carbon nanotube sensor according to the resistance change of the paper-based sensor before and after the antibody is combined with an antigen by utilizing the characteristic of specific combination of the alpha fetoprotein and the antibody thereof, and detecting the content of a target antigen by recording the resistance change of the sensor within a certain time to realize the detection of the alpha fetoprotein.

Although these detection methods can detect alpha-fetoprotein, they are complicated to operate and time-consuming, and therefore, it is desired in the art to develop a method for detecting alpha-fetoprotein simply and rapidly.

Disclosure of Invention

Aiming at the defects of the prior art, the invention aims to provide an alpha fetoprotein immunochromatographic test strip, and a preparation method and application thereof.

In order to achieve the purpose, the invention adopts the following technical scheme:

on one hand, the invention provides an alpha fetoprotein immunochromatographic test strip which comprises a sample pad, a combination pad, a nitrocellulose membrane, a water absorption pad and a backing plate, wherein a composite fluorescent microsphere probe is combined on the combination pad, the composite fluorescent microsphere probe is a microsphere probe formed by an alpha fetoprotein monoclonal antibody, a carbon nano tube and water-soluble quantum dots, a detection line and a quality control line are arranged on the nitrocellulose membrane, the detection line contains an alpha fetoprotein antibody, and the quality control line contains goat anti-mouse IgG.

In the invention, the alpha fetoprotein immunochromatographic test strip developed by a microspherical probe formed by the alpha fetoprotein monoclonal antibody, the carbon nano tube and the water-soluble quantum dot takes the carbon nano tube as a core, the water-soluble quantum dot and the alpha fetoprotein monoclonal antibody are coupled, and a large number of water-soluble quantum dots are combined on the surface of the carbon nano tube, so that the fluorescence intensity is obviously enhanced, and the sensitivity of the test strip for detecting the alpha fetoprotein is obviously enhanced.

Preferably, the carbon nanotubes are multi-walled carbon nanotubes or single-walled carbon nanotubes.

Preferably, the water-soluble quantum dots are carboxyl-coated CdTe quantum dots or carboxyl-coated CdTe/ZnSe core-shell quantum dots.

In the invention, the carboxyl-coated CdTe quantum dot or the carboxyl-coated CdTe/ZnSe core-shell quantum dot can be prepared by the prior art.

Preferably, the preparation method of the composite fluorescent microsphere probe comprises the following steps: incubating aminated carbon nanotubes and alpha fetoprotein monoclonal antibodies at 37 ℃ under the action of a condensing agent, centrifuging, redissolving precipitates after centrifugation, adding the condensing agent and carboxylated water-soluble quantum dots, incubating at 37 ℃, and centrifuging to obtain the composite fluorescent microsphere probe.

Preferably, when the aminated carbon nanotube and the alpha fetoprotein monoclonal antibody are incubated at 37 ℃ under the action of a condensing agent, the condensing agent used is any one of 1-ethyl- (3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and N-hydroxythiosuccinimide or the combination of at least two of the 1-ethyl- (3-dimethylaminopropyl) carbodiimide and the N-hydroxythiosuccinimide.

Preferably, the time for incubating the aminated carbon nanotube and the alpha fetoprotein monoclonal antibody at 37 ℃ under the action of the condensing agent is 30min-2h, such as 30min, 35min, 45min, 1h, 1.2h, 1.5h, 1.8h or 2 h.

Preferably, when the condensing agent and the carboxylated water-soluble quantum dot are added, the condensing agent is any one or a combination of at least two of 1-ethyl- (3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and N-hydroxythiosuccinimide.

Preferably, the condensing agent and the carboxylated water-soluble quantum dot are added, and the incubation time at 37 ℃ is 30min-2h, such as 30min, 35min, 45min, 1h, 1.2h, 1.5h, 1.8h or 2 h.

In another aspect, the invention provides a preparation method of the above-mentioned alpha fetoprotein immunochromatographic test strip, which comprises the following steps:

and (2) spraying the composite fluorescent microsphere probe on the bonding pad to obtain the bonding pad sprayed with the composite fluorescent microsphere probe, then respectively scribing the alpha fetoprotein antibody and goat anti-mouse IgG on a nitrocellulose membrane to obtain a detection line and a quality control line, and assembling the sample pad, the bonding pad sprayed with the composite fluorescent microsphere probe, the nitrocellulose membrane with the detection line and the quality control line and a water absorption pad on a back plate to obtain the alpha fetoprotein immunochromatographic test strip.

Preferably, the scribing is performed with a scribe.

In the invention, the preparation method of the alpha fetoprotein immunochromatographic test strip is simple, efficient and easy to operate.

In another aspect, the invention provides an application of the above-mentioned alpha-fetoprotein immunochromatographic test strip in the preparation of an alpha-fetoprotein detection device.

Compared with the prior art, the invention has the following beneficial effects:

the alpha fetoprotein immunochromatographic test strip developed by a microsphere probe formed by an alpha fetoprotein monoclonal antibody, a carbon nano tube and water-soluble quantum dots takes the carbon nano tube as a core, is coupled with the water-soluble quantum dots and the alpha fetoprotein monoclonal antibody, and a large number of water-soluble quantum dots are combined on the surface of the carbon nano tube, so that the fluorescence intensity is obviously enhanced, the detection is accurate, the sensitivity of the detection of the alpha fetoprotein is obviously enhanced, and the minimum detection limit is 0.02 ng/mL.

Drawings

FIG. 1 is a schematic structural diagram of the alpha fetoprotein immunochromatographic test strip of the present invention.

Detailed Description

The technical solution of the present invention is further explained by the following embodiments. It should be understood by those skilled in the art that the examples are only for the understanding of the present invention and should not be construed as the specific limitations of the present invention.