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Preparation method and application of magnetic electrochemical immunosensor for simultaneously detecting two tumor markers based on metal substrate sign

A tumor marker and immunosensor technology, which is applied in the preparation and application fields of a magnetic electrochemical immunosensor for simultaneous detection of two tumor markers based on metal-based labeling, can solve the problem of large sample consumption, cumbersome operation, and unsuitable for rapid detection. and other problems, to achieve the effect of large active surface area and improved sensitivity

Inactive Publication Date: 2016-07-06
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods have high sensitivity and selectivity, but the detection process requires expensive special instruments, complex pretreatment and cumbersome operations, and large sample consumption, which is not suitable for rapid detection

Method used

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  • Preparation method and application of magnetic electrochemical immunosensor for simultaneously detecting two tumor markers based on metal substrate sign
  • Preparation method and application of magnetic electrochemical immunosensor for simultaneously detecting two tumor markers based on metal substrate sign
  • Preparation method and application of magnetic electrochemical immunosensor for simultaneously detecting two tumor markers based on metal substrate sign

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036]Example 1. A method for preparing a magnetic electrochemical immunosensor for simultaneous detection of two tumor markers using a metal ion complex label:

[0037] (1) Add 100 μL anti-CEA (100 μg / ml, secondary antibody) and 100 μL anti-AFP (100 μg / ml, secondary antibody) to G-Au-Cd 2+ and G-Au-Pb 2+ , shaking overnight to obtain anti-CEA-G-Au-Cd 2+ and anti-AFP-G-Au-Pb 2+ , add 1mL of 1% BSA, shake for 2h, block the non-specific binding site, wash with PBS (0.01mol / L, pH7.4) by centrifugation, and disperse into 5mL of PBS (0.01mol / L, pH7.4);

[0038] (2) Add 150 μL anti-AFP (10 μg / mL, primary antibody) and 150 μL anti-CEA (10 μg / mL, primary antibody) to Fe 3 o 4 In PDA, incubate at 4°C for 1 h, then wash with PBS to remove non-specifically bound primary antibody; add 0.5 mL of 2% BSA, shake at 4°C for 1 h, wash with PBS 3 times to obtain Fe 3 o 4 PDA / Ab 1 ; the Fe 3 o 4 PDA / Ab 1 Incubate in 100μLAFP (10μg / mL) and CEA (10μg / mL) at 37oC for 40min, wash with PBS 3...

Embodiment 2

[0040] Embodiment 2. Preparation of Magnetic Dopamine Nanospheres

[0041] (1) Magnetic Fe 3 o 4 Preparation of nanoparticles

[0042] 2.7gFeCl 3 ·6H 2 O and 7.2g of NaAc were dissolved in 100mL of ethylene glycol under vigorous stirring to obtain a homogeneous yellow solution, and then the solution was transferred to a polytetrafluoroethylene autoclave to seal, and the reaction was heated at 180°C for 10h, and the autoclave was cooled to room temperature Finally, the black magnetic particles were separated and collected by a magnet, and washed with deionized water and absolute ethanol to obtain magnetic Fe 3 o 4 Nanoparticles;

[0043] (2) Magnetic Fe 3 o 4 Preparation of PDA

[0044] 25mg of Fe 3 o 4 The particles were added to 25 mL of 0.02 mol / L Tris-HCl (pH 8.0) solution and ultrasonicated for 30 min to make Fe 3 o 4 Particles can be fully dispersed, disperse 50mg of dopamine hydrochloride into 25mL deionized water, quickly inject the dopamine solution into F...

Embodiment 3

[0045] Embodiment 3. Preparation of graphene / gold / metal ion (G-Au-M) composite

[0046] (1) Preparation of G-Au

[0047] Weigh 4mg of graphene, disperse in 10ml of secondary water, ultrasonic 20min, 0.4mLHAuCl 4 (1.0wt%) and 0.4mgNaOH were quickly added to the above mixture, ultrasonicated for 2h, and centrifuged at 8000r / min for 10min to obtain a graphene / gold (G-Au) composite, which was redispersed in 2ml of secondary water;

[0048] (2) Preparation of G-Au-M complex

[0049] 1mL10 -4 mol / L of thioglycolic acid was added to 2mL of the above solution, and 1mL10 -3 mol / L of CdCl 2 solution, ultrasonically dispersed for 5min, stirred overnight at room temperature, washed with deionized water, and redispersed into 1mL of PBS (0.01mol / L, pH7) to obtain G-Au-Cd 2+ , Change thioglycolic acid to cysteine ​​and do G-Au-Pb in the same way 2+ .

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Abstract

The invention discloses a preparation method and application of a magnetic electrochemical immunosensor for simultaneously detecting two tumor markers based on a metal substrate sign, relates to the field of preparation of immunosensors for rapidly and sensitively detecting two tumor markers of alpha fetoprotein (AFP) and carcino-embryonic antigen (CEA) simultaneously, and in particular to a magnetic electrochemical immunosensor for simultaneously detecting two tumor markers on the basis of ionic compound signs. The advantages that magnetic dopamine nanoparticles are large in active surface and are easy to separate are taken, an appropriate end-capping reagent is selected to increase the load amount of metal ions, then an electromechancial signal can be amplified, and detection on tumor rkers can be rapid and sensitive. A modification electrode is simple and convenient to manufacture, good in stability, rapid to react, and relatively high in selectivity and sensitivity for alpha fetoprotein and carcino-embryonic antigen.

Description

technical field [0001] The invention relates to the field of preparation of an immunosensor for rapid and sensitive simultaneous detection of two tumor markers, alpha-fetoprotein (AFP) and carcinoembryonic antigen (CEA), in particular to a metal ion complex label for simultaneous detection of two tumors The magnetic electrochemical immunosensor for markers uses the advantages of large active surface area and easy separation of magnetic dopamine nanospheres, and increases the load of metal ions by selecting a suitable chelating agent, thereby amplifying the electrochemical signal and realizing the simultaneous detection of tumor markers. , Rapid and sensitive detection. Background technique [0002] Tumor markers are considered to be indicators for detecting normal life processes, physiological lesions and pharmacological responses of organisms, and their early detection is of great significance for the treatment of cancer. A cancer often has a variety of related tumor marke...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/48G01N33/68G01N33/574
CPCG01N27/48G01N33/57484G01N33/68
Inventor 花小霞周长利夏方诠
Owner UNIV OF JINAN
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