Human Legumain chemiluminiscence detection kit and application thereof

A chemiluminescence detection and kit technology, which is applied in the field of immunodiagnosis, can solve the problems that the detection results are easily affected by the detection conditions, the detection of the kit is not sensitive, and the accuracy is not high, so as to improve the specificity and detection speed, improve the sensitivity and Applicability, effect of increasing luminosity

Pending Publication Date: 2020-05-15
南京拂晓生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a kind of application of human Legumain chemiluminescence detection kit, to solve the technical problem of insensitive detection of the kit of the prior art; The problem

Method used

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  • Human Legumain chemiluminiscence detection kit and application thereof
  • Human Legumain chemiluminiscence detection kit and application thereof
  • Human Legumain chemiluminiscence detection kit and application thereof

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preparation example Construction

[0036] A method for preparing a human Legumain chemiluminescence detection kit, the method comprising:

[0037] Step 1: Preparation of streptavidin-based magnetic bead suspension

[0038] 1) After dissolving and mixing ferric chloride and ferrous chloride in water, add ammonia water and react at 50-70°C for 3-5 hours to obtain magnetic microspheres; the molar ratio of ferric chloride and ferrous chloride The ratio is preferably 2:1;

[0039] 2) Wash the magnetic microspheres obtained in step 1) and redisperse them in 50% ethanol, and then add 0.5% PEG4000, 0.1% ammonia and 0.1% ethyl orthosilicate in sequence at 60-65°C and stir for 14h-18h to obtain Modified magnetic microspheres; the volume ratio of PEG4000, ammonia and ethyl orthosilicate is preferably 1:5:1;

[0040] 3) Wash the modified magnetic microspheres obtained in step 2) alternately with ultrapure water and absolute ethanol, then resuspend in 50% ethanol and add 0.05% ammonia and 0.5% tetramethylammonium hydroxide respect...

Embodiment 1

[0050] (1) Preparation of streptavidin-based magnetic bead suspension

[0051] 1. Weigh ferric chloride and ferrous chloride at a molar ratio of 2:1, dissolve and mix them in ultrapure water, add ammonia to a final concentration of 0.5%, stir and react at 60°C for 4 hours to obtain magnetic microspheres;

[0052] 2. Wash the obtained magnetic microspheres with ultrapure water and redisperse them in 50% ethanol, then add 1mL 0.5% PEG4000, 5mL 0.1% ammonia water and 1mL 0.1% ethyl orthosilicate, and stir at 60℃. React for 16 hours to obtain modified magnetic microspheres;

[0053] 3. Wash the modified magnetic microspheres with ultrapure water and absolute ethanol alternately, then resuspend them in 50% ethanol, and then add 5mL 0.05% ammonia, 2mL 0.5% tetramethylammonium hydroxide TMA and 4mL 2.5% 3-aminopropyltrimethoxysilane, stirred and reacted at room temperature for 16h, to obtain magnetic microspheres with surface modified amino groups;

[0054] Take 8mg of the amino magnetic mi...

Embodiment 2

[0063] (1) Preparation of streptavidin-based magnetic bead suspension

[0064] Weigh ferric chloride and ferrous chloride at a molar ratio of 2:1, dissolve and mix them in ultrapure water, add ammonia water to a final concentration of 0.5%, stir and react at 60°C for 3 hours to obtain magnetic microspheres;

[0065] The obtained magnetic microspheres were washed with ultrapure water and re-dispersed in 50% ethanol, and then 1mL 0.5% PEG4000, 5mL 0.1% ammonia water and 1mL 0.1% ethyl orthosilicate were added successively, and the reaction was stirred at 60°C for 14h. Obtain modified magnetic microspheres;

[0066] The modified magnetic microspheres were washed alternately with ultrapure water and absolute ethanol, then resuspended in 50% ethanol, and then added 5mL 0.05% ammonia, 2mL 0.5% tetramethylammonium hydroxide TMA and 4mL 2.5 % 3-aminopropyltrimethoxysilane, stirred and reacted at room temperature for 14 hours, to obtain magnetic microspheres with surface modified amino group...

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Abstract

The invention discloses a human Legumain chemiluminiscence detection kit and application thereof. The kit comprises the following components: a) streptavidin magnetic bead suspension; b) a Legumain antibody labeled by acridinium ester; c) a Legumain antibody labeled by using biotin; d) a chemiluminescent substrate solution comprising a solution A and a solution B, the solution A being a nitric acid solution and the solution B being a sodium hydroxide solution; e) calibrators: Legumain antigens with the concentrations of 0 pg / mL, 10 pg / mL, 50 pg / mL, 100 pg / mL, 500 pg / mL, 1000 pg / mL and 2500 pg / mL; f) a quality control product; g) a cleanout fluid. According to the kit, streptavidin magnetic bead suspension liquid serves as a solid-phase carrier for immune reaction and separation, avidin magnetic particles are mainly used for being connected with biotin labeled antibodies, and target substances can be captured by the avidin magnetic particles through the immune reaction to form immune complexes.

Description

Technical field [0001] The invention belongs to the technical field of immunodiagnosis, and specifically relates to the application of a human Legumain chemiluminescence detection kit. Background technique [0002] Ovarian cancer is a common gynecological malignant tumor. Its mortality rate ranks the first among all gynecological malignancies, which seriously harms women’s health. Because ovarian cancer usually has no obvious symptoms in its early stage and lacks simple and effective early diagnosis methods, it is about appointment when symptoms appear. 70% of patients have advanced stage, the 5-year survival rate of advanced patients is 20%-30%, and the 5-year survival rate of early stage patients is 70-90%. Therefore, the early detection of ovarian cancer plays an important role in the selection of treatment options and the improvement of patient prognosis. [0003] At present, the clinically commonly used early detection methods for ovarian cancer mainly include gynecological e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/543
CPCG01N33/57449G01N33/54326
Inventor 李伟赵树杰
Owner 南京拂晓生物科技有限公司
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