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Kit for detecting anti-MCV (mutation citrullinated vimentin) antibody and preparation method of kit

A technology of citrullination and vimentin, which is applied in the field of clinical immunoassay, can solve the problems of colloidal gold method such as low sensitivity and specificity, complicated detection steps, long reaction time, etc., to achieve improved sensitivity and specificity, narrow emission, affinity and high adsorption effect

Inactive Publication Date: 2019-01-25
AUTOBIO DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The ELISA method requires special equipment, the reaction time is relatively long, and the detection steps are complicated; the colloidal gold method has low sensitivity and specificity, and its application is limited
Up to now, kits for rapid quantitative detection of anti-MCV antibodies using fluorescent microsphere labeling technology are still blank in my country

Method used

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  • Kit for detecting anti-MCV (mutation citrullinated vimentin) antibody and preparation method of kit
  • Kit for detecting anti-MCV (mutation citrullinated vimentin) antibody and preparation method of kit
  • Kit for detecting anti-MCV (mutation citrullinated vimentin) antibody and preparation method of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Preparation of a kit for detecting anti-mutant citrullinated vimentin antibodies

[0029] In the first step, a nitrocellulose membrane coated with anti-mutant citrullinated vimentin (MCV) was prepared

[0030] Take 30cm of nitrocellulose membrane and paste it on the self-adhesive liner, place it on the film scratcher, and coat the film with a spray volume of 1ul / cm according to the operating rules of the film scratcher; wherein the detection line is coated with a concentration of 0.5-1mg / ml MCV protein, the quality control line is coated with goat anti-mouse IgG antibody at a concentration of 1-2mg / ml, and dried at 37°C for 5 hours after coating;

[0031] The second step is to prepare a fluorescent microsphere conjugate pad coupled with mouse anti-human IgG antibody (quantum dot microspheres are used for fluorescent microspheres)

[0032] Take 2ml 0.5M PH8.0 Tris buffer in a clean glass bottle, add 100-200ug quantum dot microspheres and ultrasonically dispers...

Embodiment 2

[0039] The usage method of the kit prepared in embodiment 2 embodiment 1

[0040] 1. Reagent preparation and preparation:

[0041] 1. Place the kit and samples at room temperature (18-25°C) to equilibrate for at least 30 minutes.

[0042] 2. Quality control: Confirm that the chip card matches the batch number of the test card, and insert the fluorescence measuring instrument for quality control.

[0043] 2. Experimental operation:

[0044] 1. Open the test reagent and package, take out the test card and sample diluent and number them.

[0045] 2. Add sample

[0046] Take 20 μl of serum / plasma / whole blood sample, add it to the sample diluent, mix well, use a matching pipette to draw 60 μl into the sample hole of the test card, and start timing.

[0047] 3. Detection

[0048] Insert the test card into the instrument immediately after adding the test card for 10 minutes, click the "Test" button, the system will automatically read the card and display the test result. When the ...

Embodiment 3

[0049] Embodiment 3 The performance test of kit of the present invention

[0050] 1. Analytical sensitivity: the lowest detection limit is not higher than 0.5U / mL;

[0051] Analytical sensitivity is expressed in terms of detection limit, and the definition of analytical sensitivity is: it refers to the amount that can be distinguished from zero dose in a statistical sense. Repeat 20 times to measure the 0-value calibrator, calculate the mean (X) and standard deviation (SD), and the calculated concentration value of X+2SD is the analytical sensitivity of the kit. The analytical sensitivity of the kit of the present invention is 0.5U / mL.

[0052] 2. Linear range: 0-1000 U / mL;

[0053] Dilute the high-value samples close to the upper limit of the linear range to at least 5 concentrations in a certain proportion, and the samples with low-value concentrations must be close to the lower limit of the linear range. The samples of each concentration were tested twice, the average va...

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Abstract

The invention discloses a kit for detecting an anti-MCV (mutation citrullinated vimentin) antibody. The kit comprises a nitrocellulose membrane, a fluorescent microsphere conjugate, anti-MCV antibodyseries standard products and sample diluting liquid, wherein the nitrocellulose membrane is coated with MCV; the fluorescent microsphere conjugateis coupled with a mouse anti-human IgG antibody. The kit has the advantages that by combining an immunochromatography method and a fluorescent microsphere coupling technique, a fluorescent microsphere amplification technique is combined on the basis of immunochromatography analysis, and the quantum yield of luminescence can be measured by a fluorescent signal measuring instrument; the quantum yield of luminiscence can form a ratio relationship with the content of a to-be-measured substance in a sample, so as to establish a calibrating curve and calculate the content of the to-be-measured substance in the sample; the sensitivity and specificity ofthe kit are higher, the reaction time is only 10 to 15min, the detection is convenient and rapid, and the kit is suitable for large hospitals to detect, and is also suitable for basis medical organizations, community medical service stations, clinical departments, outpatient departments and the like to instantly detect.

Description

technical field [0001] The invention relates to clinical immunological detection technology, in particular to a kit for detecting anti-mutant citrullinated vimentin antibody by immunofluorescence method, and also relates to a preparation method of the kit. Background technique [0002] Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease and one of the most common autoimmune diseases, with a worldwide incidence of 1% and an incidence of about 0.3% to 0.6% in my country. The main manifestation of RA is joint damage and loss of function caused by joint inflammation. Early diagnosis of RA and immediate appropriate treatment are essential to prevent total joint damage. So far, the physical damage caused by arthritis to the body cannot be repaired, so it is necessary to intervene and treat the disease as early as possible to reduce the degree of irreparable damage. This means that early diagnosis and early treatment of RA are crucial to the quality of life of pa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/558G01N33/543G01N21/64
CPCG01N21/6428G01N33/54313G01N33/558G01N33/6893G01N2021/6439
Inventor 李奎于林李双法付清山程云龙渠海刘功成付光宇吴学炜
Owner AUTOBIO DIAGNOSTICS CO LTD
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