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A preparation method of immunomagnetic microspheres, and a Sudan red detection kit and detection method containing immunomagnetic microspheres

A detection kit and magnetic microsphere technology, which is applied in the field of analysis and detection, can solve the problems of low coupling rate and poor stability, and achieve the effect of increased coupling rate, good stability and short detection time

Active Publication Date: 2016-06-29
BEIJING PURKINJE GENERAL INSTR
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to overcome the defects of low coupling rate and poor stability in the preparation process of immunomagnetic microspheres, the purpose of the present invention is to provide a preparation method of immunomagnetic microspheres

Method used

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  • A preparation method of immunomagnetic microspheres, and a Sudan red detection kit and detection method containing immunomagnetic microspheres
  • A preparation method of immunomagnetic microspheres, and a Sudan red detection kit and detection method containing immunomagnetic microspheres
  • A preparation method of immunomagnetic microspheres, and a Sudan red detection kit and detection method containing immunomagnetic microspheres

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preparation example Construction

[0029] The preparation process of immunomagnetic microspheres is usually: the magnetic microspheres with chemical functional groups on the surface are activated by the activator in the activation buffer, and then the activated magnetic microspheres are placed in the coupling buffer, and the The antibody or antigen undergoes a coupling reaction. After the coupling reaction is completed, unreacted sites on the surface of the magnetic microspheres are sealed, and then the obtained immunomagnetic microspheres are used or dispersed in a storage solution for storage. The activation of magnetic microspheres mostly uses MES (2-morpholineethanesulfonic acid) as a buffer, but the inventors found that in some coupling reactions between magnetic microspheres and antibodies or antigens, the coupling reaction of MES buffer is used. The connection rate is poor, and the obtained immunomagnetic microspheres have poor stability and are prone to agglutination. After a large number of experiments,...

Embodiment 1

[0066] Example 1 Preparation of immunomagnetic microspheres coated with Sudan red antibody

[0067] 1. Preparation of reagents

[0068] 1) Activation / coupling buffer:

[0069] Preparation of 10mMPB buffer (pH 6.0): ①Weigh 1.56g of sodium dihydrogen phosphate and dilute to 1000mL with deionized water; ②Weigh 0.358g of dibasic sodium phosphate and dilute to 100mL with deionized water; add ①Reagent 17.54mL and ②Reagent 2.46mL, then add Tween-20 and PVP to the 10mMPB buffer solution obtained, the volume percentage of Tween-20 is 0.05%, the mass percentage of PVP is 0.05%, and the validity period is 12 months .

[0070] 2) Sealing fluid:

[0071] Weigh 0.1 g of BSA solid and dissolve it in 10 mL of coupling buffer. The reagent is a light yellow, clear and transparent liquid, and the prepared sealing solution can be stored at 2~8℃ for 30 days.

[0072] 3) Storage solution:

[0073] Weigh 0.24g potassium dihydrogen phosphate, 3.58g disodium hydrogen phosphate, 8.0g sodium chloride, 0.2g potas...

Embodiment 2

[0085] Example 2 Construction of Sudan Red Rapid Detection Kit Based on Magnetic Microspheres

[0086] A rapid detection kit for Sudan red based on magnetic microspheres was formed to include the following components:

[0087] (1) The immunomagnetic microspheres coated with Sudan Red Antibody No. Ⅰ monoclonal prepared in Example 1.

[0088] (2) Sudan Red Enzyme Conjugate, HRP-labeled Sudan Red Antigen (Sudan Red Antigen was purchased from Guangzhou Youkangduo Biotechnology Co., Ltd. (13mg / mL), and HRP-labeled Sudan Red Antigen was from Beijing Tianenze Gene Technology Limited company (1mg / mL)).

[0089] (3) Sample diluent, the sample diluent is 0.01MPBS (phosphate) buffer containing 10% methanol, 0.1% TritonX-100, and the pH is 7.4.

[0090] (4) Sudan red standard: purchased from Beijing Beina Chuanglian Biotechnology Research Institute (100mg / bottle), and the concentration of Sudan red standard solution of 0, 0.2, 0.6, 1.8, 5.4 ng / mL was prepared with sample diluent .

[0091] (5) Chr...

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Abstract

The invention provides a preparation method for an immunomagnetic microsphere. The method adopts a phosphate buffer containing 0.01%-0.1% of Tween-20 and 0.01%-0.1% of polyvinylpyrrolidone as the activating buffer and / or coupling buffer. The invention also discloses a Sudan red detection kit containing the immunomagnetic microsphere and a detection method using the kit to conduct detection. The immunomagnetic microsphere preparation method provided by the invention has obviously improved coupling rate, and the obtained immunomagnetic microsphere has good stability. The kit provided by the invention uses the Sudan red monoclonal antibody coated immunomagnetic microsphere, has high sensitivity, can be used for rapid screening of mass samples, can avoid the influence of the matrix in a to-be-detected sample, and has high accuracy. The detection method provided by the invention can play an important role in food safety detection, and has important social significance and broad market prospects.

Description

Technical field [0001] The invention relates to the field of analysis and detection, in particular to a preparation method of immunomagnetic microspheres, and a Sudan red detection kit containing immunomagnetic microspheres and a detection method. Background technique [0002] Sudan red is a kind of chemical dye, which is widely used to enhance the color of solvents, oils, waxes, gasoline, and to enhance the gloss of shoes and floors. It is not a food additive, but because of its bright coloring, it is often illegally added to chili powder and other condiments. Its chemical composition has an azo structure. Due to the nature of this chemical structure, it is carcinogenic and has obvious toxic effects on the liver and kidney organs of the human body. Therefore, it is strictly prohibited to be added to food. [0003] At present, the detection methods of Sudan red include high performance liquid chromatography, liquid chromatography mass spectrometry, etc. Although these methods are ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/531G01N33/577
CPCG01N33/531G01N33/577
Inventor 王琳田静秒王雪万渝平周浩叶敏梁恒兴张敏
Owner BEIJING PURKINJE GENERAL INSTR
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