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Fluorescence-based alpha-fetoprotein detection kit and preparation thereof

A detection kit, a technology for alpha-fetoprotein, which is applied in the fields of biotechnology and clinical medical diagnosis, can solve the problems such as the inability to meet the requirements of less than 1ng/mL, the small particle size of CdTe quantum dots, and the inability to achieve rapid detection, etc. Market competitiveness, high sensitivity and good stability

Inactive Publication Date: 2018-07-31
NANJING UNIVERSITY OF TRADITIONAL CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The study of the present invention shows that many nanoparticles do not meet the requirements of the test within this range, and the simple CdTe quantum dots have a small particle size and limited antibody connection, which cannot meet the clinical requirements for sensitivity less than 1ng / mL and the minimum detection limit. The requirement of 5ng / mL, and the linear range does not cover the concentration range of normal physiological and pathological conditions in clinical practice, so false negative results are often caused; at the same time, there is also a long reaction time (equilibrium time 10-30min), which cannot meet the requirements of rapid testing

Method used

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  • Fluorescence-based alpha-fetoprotein detection kit and preparation thereof
  • Fluorescence-based alpha-fetoprotein detection kit and preparation thereof
  • Fluorescence-based alpha-fetoprotein detection kit and preparation thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0029] Embodiment 1: the preparation of fluorescent ZnS-CdTe nano microsphere

[0030] Experimental materials: zinc nitrate hexahydrate, batch number: B1224002, purchased from Shanghai Aladdin Reagent Co., Ltd; gum arabic powder, batch number 20120825, purchased from Shanghai Aladdin Reagent Co., Ltd; thioacetamide, batch number: 20010619, purchased from From Shanghai Aladdin Reagent Co., Ltd.; β-mercaptoethylamine, batch number: 11325054, purchased from Shanghai Aladdin Reagent Co., Ltd.; dimethylformamide, batch number: F20061110, purchased from Nanjing Chemical Reagent Co., Ltd.; polyacrylic acid, Batch number: J1225008, purchased from Nanjing Chemical Reagent Co., Ltd.; ethanol, batch number: 14021710247, purchased from Nanjing Chemical Reagent Co., Ltd. DL-dithiothreitol, batch number MFCD00004877, was purchased from Shanghai Sangong Reagent Co., Ltd.

[0031]Experimental instruments and equipment: round bottom flask, KQ-500DE ultrasonic cleaner, PTFE reaction kettle, DH...

Embodiment 2

[0039] Example 2: Conjugation of antibodies to ZnS-CdTe nanoparticles

[0040] Experimental materials: bovine serum albumin, batch number 140318, Shanghai Aladdin Reagent Co., Ltd.; 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), batch number 090M14531V, Shanghai Aladdin Reagent Co., Ltd. Latin Reagent Co., Ltd.; succinimide (NHS), lot number MKBG7914V, Shanghai Aladdin Reagent Co., Ltd.; ethanol, lot number: 14021710247, purchased from Nanjing Chemical Reagent Co., Ltd.

[0041] Experimental instruments and equipment: KQ-500DE ultrasonic cleaner, DHG-9143BS electric blast drying oven, AY120 electronic analytical balance, TGL-16C centrifuge.

[0042] Add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) 2mg and 1mg of N- Hydroxysuccinimide (NHS), shake at room temperature for 60 minutes, then add 200 μL of alpha-fetoprotein antibody solution to the above solution, and react at room temperature for 24 hours. After the reaction was completed, ...

Embodiment 3

[0043] Embodiment 3: the configuration of R2 reagent in the kit

[0044] Experimental materials: PEG 8000, batch number MFCD00081839, Shanghai Sangon Biological Reagent Co., Ltd.; glycerol, batch number MFCD00004722, Shanghai Sangon Biological Reagent Co., Ltd.

[0045] Experimental instruments and equipment: KQ-500DE ultrasonic cleaner, AY120 electronic analytical balance, TGL-16C centrifuge.

[0046] A stabilizer is added to the nanometer ZnS-CdTe particle solution coupled with the alpha-fetoprotein antibody. In the ZnS-CdTe nanoparticle solution after the above-mentioned coupling, add the phosphate buffer of the pH value 7.2 of weight volume ratio 10% sucrose, weight volume ratio 0.2% PEG 8000, weight volume ratio 0.5% BSA and weight volume ratio 5% glycerol The solution was resuspended and stirred for 10 minutes to obtain the R2 reagent, which was divided and used.

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Abstract

The invention belongs to the field of biotechnology and clinical medical diagnosis, and relates to an alpha-fetoprotein detection kit based on fluorescent CdTe quantum dot composite porous ZnS nanospheres (ZnS-CdTe) and a preparation thereof, and early diagnosis of cancer and tumor diseases can be realized. The kit has the characteristics of high sensitivity, strong specificity and good stability,can be used for detecting the alpha-fetoprotein content in serum or plasma, and is suitable for a multi-function microplate reader with a fluorescent function, the alpha-fetoprotein detection sensitivity of the kit can reach 0.02ng / mL, and the detection linear range is 0.1-100ng / mL, and the kit has a good application prospect.

Description

technical field [0001] The invention belongs to the field of biotechnology and clinical medical diagnosis, and relates to an alpha-fetoprotein detection kit based on fluorescent CdTe quantum dot composite porous ZnS nanospheres (ZnS-CdTe) and its preparation, so as to realize early diagnosis of cancer and tumor diseases . Background technique [0002] In 1944, the existence of alpha-fetoprotein was studied, but it was not until 1965 that American scientists Bergstyandh and Zar observed that the serum of two patients with primary liver cancer could react with the monovalent antiserum of anti-fetal serum, indicating that the primary There is a common special component in the serum of liver cancer patients and human fetal serum, and this special protein component is called a-fetoprotein (AFP). Alpha-fetoprotein, an oncogenic glycoprotein, serves as a "molecular marker" of physiological status in adults and is often used as a suitable "biomarker" for the diagnosis of hepatocell...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/543G01N33/533G01N21/64
CPCG01N21/6428G01N33/533G01N33/54346G01N33/6893G01N2021/6439
Inventor 朱栋缪赵怡胡玥
Owner NANJING UNIVERSITY OF TRADITIONAL CHINESE MEDICINE
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