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Chemiluminescence detection kit for big endothelin-1 and preparation method thereof

A chemiluminescence detection and chemiluminescence technology, applied in chemiluminescence/bioluminescence, analysis by chemical reaction of materials, measurement devices, etc., can solve the problems of unstable results, influence of signal-to-noise ratio, and many influencing factors, etc. Achieve the effect of no reduction in photon yield, good reagent stability and low background luminescence

Inactive Publication Date: 2018-06-12
太原瑞盛生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although radioimmunoassay has the advantages of high sensitivity and high specificity, it has many operating steps, requires special testing equipment, expensive reagents, needs to use matching instruments, and has radioactive contamination, which exposes operators to radioactive hazards; in addition, Radioactive isotopes are easy to decay, have a short validity period, and are not easy to store
ELISA uses horseradish peroxidase, and the main disadvantage of using horseradish peroxidase is that luminol will also be absorbed by H in the absence of horseradish peroxidase. 2 o 2 Oxidation is self-luminescent, the background is relatively high, which affects the signal-to-noise ratio, the reaction kinetics is complex, there are many influencing factors, the result is not stable enough, and it is not easy to obtain a substrate with high sensitivity and long plateau period

Method used

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preparation example Construction

[0025] The invention provides a large endothelin-1 chemiluminescence detection kit and preparation method, magnetic particle-coupled large endothelin-1 (ET-1) capture antibody, acridinium ester-labeled large endothelin-1 (ET-1) -1) Detection antibody. The kit also includes large endothelin-1 calibrator, chemiluminescence pre-excitation solution A, chemiluminescence excitation solution B and cleaning solution.

[0026] Specifically, in the process of preparing the magnetic particle-coupled capture antibody in the present invention, the antibody-coupled buffer is a buffer with a pH value of 5.0-6.0 and a concentration of 20-200 mmol / L MES.

[0027] Specifically, in the process of preparing the magnetic particle-coupled capture antibody in the present invention, the blocking buffer is a buffer containing 1% BSA.

[0028] Specifically, in the process of preparing the acridinium ester-labeled detection antibody in the present invention, the buffer solution of the acridinium ester-...

Embodiment 1

[0034]Example 1: The composition and preparation method of a large endothelin-1 chemiluminescent detection kit

[0035] 1. Assembly of the kit

[0036] Large endothelin-1 monoclonal antibody coupled to magnetic particles;

[0037] Acridine ester-labeled large endothelin-1 (ET-1) monoclonal antibody;

[0038] Large endothelin-1 series calibrator solution;

[0039] Chemiluminescence pre-excitation solution A and chemiluminescence excitation solution B;

[0040] Cleaning fluid.

[0041] 2. Preparation of Magnetic Microparticle Suspension Conjugated to Large Endothelin-1 Monoclonal Antibody

[0042] (1) Take 1 mg of carboxyl magnetic particles in a 0.5 mL centrifuge tube, add 200 μL of MES buffer solution with a concentration of 0.1 mol / L, vortex and mix, place on a magnetic stand, and let it stand for 5 minutes to make the magnetic particles and liquid Separate, discard the supernatant, wash 3 times, then add 200 μL of MES (pH 6.0) buffer, and vortex.

[0043] (2) Add 15 μg...

Embodiment 2

[0062] Embodiment 2: detection and result analysis with kit

[0063] (1) Add 50 μL of the sample to be tested into the cuvette, then add 150 μL of magnetic particle coupling suspension, shake and mix, and incubate at 37°C for 8 min.

[0064] (2) Separate and wash 3 times. Shake the washed reaction vessel sufficiently to disperse the magnetic particles.

[0065] (3) Add 150 μL of acridinium ester marker into the cuvette, shake to mix, and incubate at 37°C for 7 min.

[0066] (4) Separate and wash 3 times. Shake the washed reaction vessel sufficiently to disperse the magnetic particles.

[0067] (5) Add 100 μL chemiluminescence pre-excitation solution A, add 100 μL chemiluminescence excitation solution B after 1 second, and measure the relative luminous intensity. The content of large endothelin-1 in the sample is proportional to its relative luminous intensity.

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Abstract

The invention discloses a chemiluminescence detection kit for big endothelin-1 and a preparation method thereof. The kit comprises a magnetic particle coupled big endothelin-1 (ET-1) capture antibody,an acridinium ester marked big endothelin-1 (ET-1) detection antibody, a big endothelin-1 calibration product , a chemiluminescence pre-excitation liquid A, a chemiluminescence excitation liquid B and a cleaning fluid. The kit provided by the invention combines a chemiluminescence technology and immune magnetic particles, and provides a reaction system approximate to a homogeneous phase. Comparedwith the prior art, the direct chemiluminescence method built by the invention is high in sensitivity, high in specificity, accurate, fast, short in detection time, and highly accurate and repetitivein detection result; the kit can be applicable to various luminescence detection instruments.

Description

technical field [0001] The invention belongs to the field of immunological detection and analysis, in particular to a large endothelin-1 immunochemiluminescent detection kit and a preparation method thereof. Background technique [0002] Endothelin (ET) is an active peptide with various biological effects. Among the ET family, ET-1 is most closely related to tumors, which is involved in the occurrence and development of many tumors. ET-1 is an active peptide isolated from the supernatant of cultured porcine aortic endothelial cells, which has a strong vasoconstrictive effect. There are three types, namely ET-1, ET-2 and ET-3. Studies have shown that in addition to constricting blood vessels, ET also has other extensive biological activities, and is closely related to tumors. Many scholars have also found that the level of ET-1 in the plasma of patients with liver cancer, malignant hemangiocellular carcinoma, breast cancer and pancreatic cancer is higher than that of normal...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/532G01N21/76
CPCG01N33/532G01N21/76
Inventor 曹晶刘丽青胡雪婷常燕杜爱铭徐兵
Owner 太原瑞盛生物科技有限公司
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