ECL (electrochemiluminescence) immunosensor for detecting tumor markers and preparation method and applications thereof

A tumor marker, luminescent immune technology, applied in the field of electrochemiluminescence immunosensor and its detection, to achieve the effects of accurate results, flexible and diverse methods, and strong selectivity

Inactive Publication Date: 2013-05-22
NINGBO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to its ultra-high sensitivity, biological barcoding technology has attracted more and more attention from scientists since it came out in 2003, but so far there has been no research report on biological barcoding technology in the construction of immune sensors

Method used

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  • ECL (electrochemiluminescence) immunosensor for detecting tumor markers and preparation method and applications thereof
  • ECL (electrochemiluminescence) immunosensor for detecting tumor markers and preparation method and applications thereof
  • ECL (electrochemiluminescence) immunosensor for detecting tumor markers and preparation method and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

specific Embodiment 1

[0053] The synthesis of the composite functionalized nanospheres simultaneously labeled with the second antibody of the tumor marker and the electrochemiluminescent marker, the specific steps are as follows:

[0054] (a) Synthesis of conductive nanospheres: Add 90–100 mL of a mixture of ethanol, water, and concentrated ammonia into a clean beaker (V 乙醇 :V 水 :V 浓氨水 = 87:7:1), magnetically stirred for 30 min; after the solution was evenly mixed, slowly added 5 mL of tetraethyl orthosilicate (TEOS) dropwise while stirring magnetically; after the dropwise addition, sealed with parafilm The mouth of the beaker was reacted for 10 h; centrifuged at 4000 rpm, and then the precipitate was dispersed in ultrapure water to form a 10 mg / mL first suspension; 2 mL of the first suspension was put into a centrifuge tube, and 0.18 mL of 3- After aminopropyltriethoxysilane (APTES), mix and stir at room temperature for 7 h; centrifuge and wash at 4000 rpm, and then disperse the precipitate in ...

specific Embodiment 2

[0059] The preparation of the gold electrode assembled by cysteamine, its specific steps are as follows:

[0060] A gold electrode with a diameter of 3-5 mm was polished with 1.0 μm, 0.3 μm, and 0.05 μm Al2O3 polishing powder in sequence, and the gold electrode was rinsed with ultrapure water, ultrasonicated in ultrapure water for 2 min, and then placed in 0.5 M H 2 SO 4 In the process, the cyclic voltammetry scan was carried out in the potential range of 0-1.6 V, and the scan rate was 100 mV / s until the cyclic voltammetry curve was stable; after the gold electrode was rinsed with ultrapure water, soaked in 0.1 mol / L The cysteamine-modified gold electrode was obtained by reacting in the cysteamine solution at 4 °C for 10 h, and then rinsing with ultrapure water. This process can be monitored by AC impedance method. Such as image 3 As shown, the impedance of the bare electrode before unmodified cysteamine is very small (curve a), and the impedance of the gold electrode aft...

specific Embodiment 3

[0061] The preparation of the gold electrode assembled with glutaraldehyde and tumor marker primary antibody, its specific steps are as follows:

[0062] After rinsing the gold electrode in Example 2 with ultrapure water, soak it in an aqueous solution containing 2.5% glutaraldehyde at 4°C for 1 h; after rinsing the gold electrode with ultrapure water, soak it in 50 μg / mL of the tumor marker primary antibody was reacted at 4 °C for 12-18 h, rinsed with ultrapure water, and the gold electrode modified with glutaraldehyde and tumor marker primary antibody was obtained. This process can be monitored by AC impedance method. Such as image 3 As shown, the impedance of the gold electrode after modification of the glutaraldehyde and the primary antibody of the tumor marker increased greatly (curve c), indicating that the glutaraldehyde and the primary antibody of the tumor marker were successfully assembled on the gold electrode.

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Abstract

The invention discloses an ECL (electrochemiluminescence) immunosensor for detecting tumor markers and a preparation method and applications thereof. The immunosensor comprises a working electrode, a reference electrode and a counter electrode, wherein the working electrode is a gold electrode, cysteamine, glutaraldehyde, a tumor marker first-antibody, a tumor marker, and a composite functionalized nanosphere are sequentially decorated on the surface of the gold electrode, the composite functionalized nanosphere is simultaneously marked by a tumor marker second-antibody and an ECL marker, and non-specific active sites are closed by using bovine serum albumins. The preparation method comprises the steps of preparing the gold electrode on which the tumor marker first-antibody is fixedly carried; synthesizing the composite functionalized nanosphere simultaneously marked by the tumor marker second-antibody and the ECL marker; and finally, assembling the obtained gold electrode and the composite functionalized nanosphere into an ECL immunosensor. The ECL immunosensor disclosed by the invention is high in sensitivity, rapid in analysis speed, strong in stability and selectivity, good in reproducibility, easy to operate, and flexible in method.

Description

technical field [0001] The invention relates to an electrochemiluminescence immunosensor and a detection method thereof, in particular to an electrochemiluminescence immunosensor constructed based on a biological barcode pattern for detecting tumor markers, a preparation method and an application method thereof. Background technique [0002] Cancer is one of the diseases that seriously threaten human health, with high morbidity and high mortality. With the improvement of people's living standards, early detection and diagnosis of cancer have become the focus of attention and research. Tumor markers are substances that are produced by tumor cells themselves during the occurrence and proliferation of tumor cells, or produced by the body's response to tumor cells, secreted and released into blood, cells, and body fluids, and can be directly , Effectively reflect the generation, development and therapeutic effect of tumor cells in the body. Obviously, the detection of tumor mar...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/532G01N21/76
Inventor 杜书平郭智勇郝婷婷陈贝贝王泽波
Owner NINGBO UNIV
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