Immunofluorescence test reagent strip for measuring cytotoxin-related protein A antibodies as well as preparation method and application of immunofluorescence test reagent strip

A related protein, immunofluorescence technology, applied in biological testing, measuring devices, material inspection products, etc., can solve the problem of complex pathogenesis, and achieve the effect of simple and convenient operation, stable and accurate test results

Inactive Publication Date: 2018-05-22
ANIMAL AND PLANT & FOOD DETECTION CENTER JIANGSU ENTRY EXIT INSPECTION AND QUARANTINE BUREAU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Atherosclerosis (AS) is caused by the joint action of multiple factors, and the pathogenesis is complex, which has not yet been fully elucidated.

Method used

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  • Immunofluorescence test reagent strip for measuring cytotoxin-related protein A antibodies as well as preparation method and application of immunofluorescence test reagent strip
  • Immunofluorescence test reagent strip for measuring cytotoxin-related protein A antibodies as well as preparation method and application of immunofluorescence test reagent strip
  • Immunofluorescence test reagent strip for measuring cytotoxin-related protein A antibodies as well as preparation method and application of immunofluorescence test reagent strip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1: Preparation of specific monoclonal antibody against CagA

[0051] 1. Animal immunity:

[0052] Female Balb / c mice aged 6-8 weeks were selected as immunization objects, blood was collected from the tail vein of each mouse before the initial immunization, and 200 μl of emulsified immunogen (antigen plus complete adjuvant, antigen CagA) was injected intraperitoneally during the initial immunization; On the 14th day and 28th day after the primary immunization, the second and third immunizations (antigen plus incomplete adjuvant) were carried out in the same way, and the orbital blood was collected from each mouse on the 5th day after the third immunization. Serum was used to evaluate the antibody titer. Booster immunization was carried out on the seventh day after the third immunization, and 100 μl of immunogen was injected into the tail vein. On the third day after the booster immunization, splenic lymphocytes and SP2 / 0 (mixing ratio of 5:1) were collected for c...

Embodiment 2

[0059] Example 2: A standard curve of an immunofluorescence reagent strip for measuring cytotoxin-associated protein A antibody

[0060] Prepare a series of concentration gradients of cytotoxin-related protein A antibody standard solution: 1RU / mL, 2RU / mL, 4RU / mL, 8RU / mL, 16RU / mL, 32RU / mL, 64RU / mL, take 50μl dropwise to immunofluorescence Then use a detector to read the fluorescence intensity of the T line and the C line, and make a fitting curve for the fluorescence intensity ratio y of T / C and the corresponding antibody concentration x ( figure 2 ), obtain the formula corresponding to the fluorescence intensity and the antibody concentration, the formula is: y=0.1678x 0.4335 , correlation coefficient: R2=0.981.

Embodiment 3

[0061] Example 3: Clinical Application of an Immunofluorescence Reagent Strip for Measuring Cytotoxin-Associated Protein A

[0062] According to the atherosclerosis index AI, (AI = [total cholesterol (TC) - high-density lipoprotein (HDL)] ÷ high-density lipoprotein (HDL), AI < 4 reflects that the degree of arteriosclerosis is not serious or is reducing, The smaller the value, the lighter the degree of arteriosclerosis, and the lower the risk of cardiovascular and cerebrovascular diseases; AI ≥ 4 indicates that arteriosclerosis has already occurred, and the larger the value, the more serious the degree of arteriosclerosis, and the risk of cardiovascular and cerebrovascular diseases The higher the sex.

[0063] 20 serum samples with AI≥4 and 30 serum samples with AI image 3 ).

[0064] The experimental results showed that the concentration of cytotoxin-associated protein A antibody was higher in most cases in the case of arteriosclerosis.

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Abstract

The invention discloses an immunofluorescence test reagent strip for measuring cytotoxin-related protein A antibodies as well as a preparation method and application of the immunofluorescence test reagent strip. The immunofluorescence test reagent strip provided by the invention has three areas, i.e., a sample adding area, a binding area and a detection area. A water absorbing pad, a chromatographic membrane, a binding pad and a sample pad are separately arranged on the reagent strip from top to bottom. The reagent strip is characterized in that the binding pad is coated with a CagA antigen, labeled with a fluorescent substance CY3, in a membrane-spraying way, and the C line of the chromatographic membrane is coated with a monoclonal antibody coated with against CagA, so that the reagent strip has high specificity and can be used for detecting a CagA antibody in serum. The immunofluorescence test reagent strip can realize quantitative detection when being used together with a fluorescence analyzer, and is simple and convenient to operate. The immunofluorescence test reagent strip can be both used for laboratory detection and on-site quick testing, and only takes about 10min for detection.

Description

technical field [0001] The invention relates to an immunofluorescent reagent strip for measuring cytotoxin-associated protein A antibody and its preparation method and application, which is used for detecting the concentration of cytotoxin-associated protein A antibody in the serum of patients with atherosclerosis, and belongs to immunology detection field. technical background [0002] Atherosclerosis (Atherosclerosis, AS) is caused by multiple factors, and its pathogenesis is complex, which has not yet been fully elucidated. AS is a chronic inflammatory disease, the inflammatory mechanism exists in the whole process of AS lesion, and the inflammatory response and immune regulation play an important role in the whole process of AS development. Lipoprotein-associated phospholipase A2 (Lp-PLA2), C-reactive protein (CRP), and various pro-inflammatory substances, such as lysolecithin and oxidized free fatty acids, produce various atherogenic effects, including endothelial cell...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/558G01N33/52
CPCG01N33/6893G01N33/52G01N33/558G01N2333/195G01N2800/323
Inventor 王毅谦高玲陈雷龙云凤邵景东姜珊李寒玲赵晓娜苏志同周萍
Owner ANIMAL AND PLANT & FOOD DETECTION CENTER JIANGSU ENTRY EXIT INSPECTION AND QUARANTINE BUREAU
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