Neutral granulocyte elastase chemiluminescence detection kit and preparation method thereof
A technology of chemiluminescence detection and elastase, which is applied in chemiluminescence/bioluminescence, biological testing, and analysis by making materials undergo chemical reactions, can solve problems such as complex reaction kinetics, unstable results, and difficult substrates , to achieve the effect of low cost, good reagent stability and rapid response
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[0025] The invention provides a chemiluminescent detection kit and preparation method for neutrophil elastase, including magnetic particle-coupled neutrophil elastase (NE) capture antibody, acridinium ester-labeled neutrophil elastase ( NE) detection antibody. The kit also includes neutrophil elastase 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 acridin...
Embodiment 1
[0034] Example 1: Construction and preparation method of a chemiluminescence detection kit for neutrophil elastase
[0035] 1. Assembly of the kit
[0036] Magnetic particle-conjugated monoclonal antibody to neutrophil elastase;
[0037] Acridinium ester-labeled neutrophil elastase (NE) monoclonal antibody;
[0038] Neutrophil elastase 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 Neutrophil Elastase 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 μ...
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 neutrophil elastase in the sample is proportional to its relative luminous intensity.
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