Preparation method and application of sandwich electrochemiluminescent immunosensor for detecting tumor markers

A tumor marker, luminescent immunosensor technology, applied in the field of sandwich electrochemiluminescence immunosensor preparation, can solve the problems of long response time, high technical requirements, poor selectivity, etc., and achieve the effect of high sensitivity

Active Publication Date: 2017-02-01
GUANGDONG TCRCURE BIOPHARMA TECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the immunoassay methods for detecting tumor markers mainly include: radioimmunoassay (RIA), enzyme-linked immunosorbent assay (ELISA), chemiluminescence immunoassay (CLIA), electrochemical immunoassay (ECIA) Chemiluminescence immunoassay (ECLIA) and immunoelectron microscopy, etc., these methods have certain sensitivity and accuracy, but each has certain shortcomings: some instruments are expensive, complex to operate, and require high technology; Various, prone to false positives and false negatives, some use radioactive reagents, some have low specificity, low sensitivity, poor selectivity, some have long response time, and cannot be repeatedly measured
At present, there are no published reports about sandwich-type electrochemiluminescence immunosensors for detecting tumor markers based on graphene oxide and carbon nitride nanocomposites.

Method used

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  • Preparation method and application of sandwich electrochemiluminescent immunosensor for detecting tumor markers
  • Preparation method and application of sandwich electrochemiluminescent immunosensor for detecting tumor markers
  • Preparation method and application of sandwich electrochemiluminescent immunosensor for detecting tumor markers

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specific Embodiment 1

[0040] A method for preparing a sandwich-type electrochemiluminescence immunosensor for detecting tumor markers, the specific steps are as follows:

[0041] (1) Functionalized graphene oxide (Ab 1 -nanoFe 3 o 4 Preparation of @GO)

[0042] a. Synthesis of magnetic graphene oxide: Add 30-50mL 1mg / mL graphene oxide into a clean three-necked flask, sonicate for 1h; then add 20-30mL0.05-0.07mol / L dropwise under the protection of nitrogen atmosphere ferric chloride solution and 20-30mL 0.10-0.15mol / L ferric chloride solution, stirred and refluxed at 70-90°C, and slowly added 10-15mL 20-30wt% ammonia water dropwise until the pH of the solution = 10- 11. Stir and reflux for 3 to 5 hours; after cooling, add a magnet for magnetic separation, wash with secondary water until the pH of the solution is 7, and set the volume to 50mL to obtain nanometer ferric oxide particles (nanoFe 3 o 4 ) Magnetic graphene oxide (nanoFe) uniformly covered on the surface of graphene oxide (GO) 3 o 4...

specific Embodiment 2

[0052] The sandwich-type electrochemiluminescent immunosensor for detecting tumor markers is used to detect tumor markers, and the specific steps are as follows:

[0053] Incubate the magnetic glassy carbon electrodes adsorbed with functionalized graphene oxide prepared in step (3) of the above-mentioned specific embodiment in 50-100 μL of tumor marker solutions containing different concentrations, and incubate at 4°C for 1-2 hours After that, wash; then take 5-10 μL of the functionalized carbon nitride solution prepared in the first step (2) of the above-mentioned specific example, and incubate at 4°C for 1-2 hours, then wash; use it as a working electrode; use a platinum electrode As a counter electrode, an Ag / AgCl electrode or a saturated calomel electrode is used as a reference electrode to form a three-electrode system; put the three-electrode system into the buffer solution to start the electrochemical reaction and measure the intensity of electrochemiluminescence; obtain...

specific Embodiment 3

[0058] A preparation method for a sandwich type electrochemiluminescence immunosensor for detecting squamous cell carcinoma antigen (SCCA), the specific steps are as follows:

[0059] (1) Preparation of functionalized graphene oxide

[0060] a. Synthesis of magnetic graphene oxide: Add 40mL of 1mg / mL graphene oxide in a clean three-necked flask, ultrasonic for 1h; then add 25mL of 0.06mol / L ferrous chloride solution and 25mL of 0.12mol / L ferric chloride solution, stirred and refluxed at 80°C, at the same time, slowly added 12mL of 25wt% ammonia water to the solution pH=10, stirred and refluxed for 4h; Wash until the pH of the solution is 7, and set the volume to 50mL to obtain a magnetic graphene oxide solution in which nanometer iron ferric oxide particles are evenly covered on the surface of graphene oxide, and store it at 4°C for later use;

[0061] b. Take 200 μL of the magnetic graphene oxide solution obtained in step (a) and place it in a glass bottle, sonicate for 1 h,...

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Abstract

The invention discloses a preparing method and application of a sandwich type electrochemical luminescence immunosensor for detecting a tumor marker. The method comprises the steps of synthesizing magnetic oxidized graphene, wherein a coupling reagent and a tumor marker primary antibody are immobilized onto the surface of magnetic graphite oxide in sequence to obtain the functional oxidized graphene; preparing functional carbon nitride, wherein gold nanoparticles and a tumor marker secondary antibody are immobilized onto the surface of a carbon nitride film in sequence to obtain the functional carbon nitride; finally adsorbing the functional oxidized graphene, the tumor marker to be detected and the functional carbon nitride onto the surface of a magnetic glassy carbon electrode in sequence. The obtained sandwich type electrochemical luminescence immunosensor serves as a working electrode, a platinum electrode is taken as a counter electrode and an Ag / AgCl electrode or saturated calomel electrode is taken as a reference electrode to calculate the accurate concentration of the tumor marker in a sample solution to be detected. Detection speed is high, the sensitivity and accuracy of detection results are high, and specificity is high.

Description

technical field [0001] The invention relates to an electrochemiluminescence immunosensor, in particular to a preparation method and application of a sandwich type electrochemiluminescence immunosensor for detecting tumor markers. Background technique [0002] Tumor is the killer of human health. Malignant tumors, also known as cancer, are a type of disease that seriously threatens human health and life. Under the action of various internal or external carcinogenic factors, cells in local tissues lose their ability to grow at the genetic level. Normal regulation, resulting in the formation of new organisms due to abnormal proliferation. According to WHO research, in 2012, the number of cancer patients and deaths in the world increased disturbingly. Nearly half of the new cancer cases occurred in Asia, most of which were in China. The number of new cancer cases in China ranked first in the world. WHO also pointed out that early diagnosis and treatment is one of the effective...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/574
CPCG01N33/57484
Inventor 郭智勇武琳张宁沙玉红卢静贾亚茹宋信信李温柔
Owner GUANGDONG TCRCURE BIOPHARMA TECHNOLOGY CO LTD
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