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Magnetic particle chemiluminescence assay kit and method for detecting staphylococcal enterotoxin A (SEA)

A chemiluminescence immunoassay, kit technology, applied in chemiluminescence/bioluminescence, analysis by chemical reaction of materials, measurement devices, etc. problems such as the use of radioimmunoassay to achieve the effects of high precision, high sensitivity and high accuracy

Active Publication Date: 2013-11-06
北京泽诚生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Radioactive waste treatment systems and institutions are required, as well as complex radioactive counting systems
The staff engaged in radioimmunization must undergo special training, be familiar with the technology, and must have a license to do the work, which limits the use of radioimmunoassays
However, the chromogenic enzyme-linked immunosorbent assay has disadvantages such as complex operation, long detection time, and poor stability.

Method used

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  • Magnetic particle chemiluminescence assay kit and method for detecting staphylococcal enterotoxin A (SEA)

Examples

Experimental program
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Effect test

Embodiment 1

[0024] Example 1 Construction of Magnetic Particle Chemiluminescence Immunoassay Kit for Quantitative Detection of Enterotoxin SEA

[0025] A magnetic particle chemiluminescent immunoassay kit for the quantitative detection of enterotoxin SEA was constructed to include the following components:

[0026] (1) Alkaline phosphatase (ALP) labeled mouse anti-Staphylococcus aureus enterotoxin SEA monoclonal antibody marker;

[0027] (2) Mouse anti-Staphylococcus aureus enterotoxin SEA monoclonal antibody labeled with fluorescein isothiocyanate;

[0028] (3) The concentrations of SEA standard solutions are: 25ng / ml, 10ng / ml, 3ng / ml, 0.9ng / ml, 0.3ng / ml, 0ng / ml;

[0029] (4) Rat anti-FITC monoclonal antibody labeled magnetic microsphere separation reagent;

[0030] (5) The washing solution is phosphate buffer containing 0.5% Tween-20 and 0.15M pH7.4;

[0031] (6) The sample diluent is 0.05% Tween-20 and 1% BSA 20mM pH7.2 phosphate buffer;

[0032] (7) Alkaline phosphatase chemilum...

Embodiment 2

[0033] Example 2 Preparation of Staphylococcus aureus enterotoxin SEA monoclonal antibody labeled with fluorescein isothiocyanate

[0034] 2.1 Preparation of reagents required for SEA monoclonal antibody labeling with fluorescein isothiocyanate

[0035] Coating buffer: pH 9.5, 0.1mol / L carbonate buffer;

[0036] 2.2 Preparation of SEA monoclonal antibody labeled fluorescein isothiocyanate

[0037] Dilute the SEA monoclonal antibody to 3-5mg / ml with pH 9.5, 0.1mol / L carbonate buffer, add 0.1-0.3ml of 1mg / ml FITC solution to each mg of antibody, and incubate at 37°C for 2-4h; After the reaction, put it into a dialysis bag and dialyze overnight at 2-8°C in pH 9.5, 0.1mol / L carbonate buffer solution. During this process, replace the buffer solution 1-2 times to make it 0.1-5ug / ml , and then stored at 2-8°C.

Embodiment 3

[0038] Example 3 Preparation of Staphylococcus aureus enterotoxin SEA monoclonal antibody-labeled alkaline phosphatase (ALP) label

[0039] 3.1 Reagents required for the activation of Staphylococcus aureus enterotoxin SEA monoclonal antibody

[0040] Reaction buffer: 0.1mol / L phosphate, 0.15mol / L sodium chloride, pH 7.3;

[0041] Stop buffer: 0.1mol / L phosphate, 0.5mol / L hydroxylamine, 25mM EDTA, pH 7.4

[0042] 3.2 Preparation of activated SEA monoclonal antibody

[0043] Dialyze the SEA monoclonal antibody in the reaction solution at room temperature for 2-4min, replace the buffer solution 1-2 times in the middle, then dilute the SEA monoclonal antibody to 2-5mg / ml with the reaction buffer; weigh a certain amount of activator SATA , diluted to 13mg / ml with DMSO, and mixed; add 5-30ul 13mg / ml SATA solution to each ml of antibody, and react at room temperature for 10-30min. Then dialyze in the stop buffer at room temperature for 2-4 hours, and replace the buffer 1-2 times...

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Abstract

The invention discloses a magnetic particle chemiluminescence assay kit and a method for detecting SEA, and belongs to the technical field of food safety detection. The method is a magnetic particle chemiluminescence assay method for fast detecting SEA in foods and preparations. The method comprises the following steps of diluting a standard substance, adding a marker into the dilute standard substance, carrying out uniform mixing, warm bath, sedimentation and washing, adding a luminous substrate into the mixture, determining chemiluminescence intensity, drawing a standard curve, wherein an OD value of a detected sample is in the range of the standard curve. The magnetic particle chemiluminescence assay kit has the advantages of simple operation, fast speed, high sensitivity, strong singularity and low cost.

Description

technical field [0001] The invention belongs to the technical field of food safety detection, and provides a method for rapid detection of enterotoxin SEA (Staphylococcus aureus enterotoxin A) magnetic particle chemiluminescence method from food and preparations. In this method, a double monoclonal antibody sandwich method is used. Compared with traditional methods, this method has the characteristics of wide application field, short time, strong specificity and high sensitivity. It has very important significance and practical application value for the application of food and other fields. Background technique [0002] The invention relates to a specific magnetic particle chemiluminescent immunoassay kit for SEA, which detects staphylococcus enterotoxin SEA in food by double monoclonal antibody sandwich method, the sensitivity is 0.014ng / ml, the detection speed is fast, and the result can be obtained within 0.5h , the mouse anti-FITC monoclonal antibody magnetic microsphere...

Claims

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Application Information

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IPC IPC(8): G01N21/76G01N33/577
Inventor 陈海生张华英
Owner 北京泽诚生物技术有限公司
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