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Platinum-based alloy structured nanorod simulation enzyme solution and application thereof in ELISA (Enzyme-Linked Immunosorbent Assay)

A platinum-based alloy, gold nanorod technology, applied in material inspection products, measuring devices, instruments, etc., can solve the problems of lack of in-depth research and reporting, and achieve the effect of simple and easy operation, easy preparation and high repeatability.

Active Publication Date: 2014-03-19
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, there is still a lack of in-depth research and reports on the detection of platinum-based alloy-structured nanorod-mimicking enzyme solutions as peroxidase-like substitutions for HRP in ELISA.

Method used

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  • Platinum-based alloy structured nanorod simulation enzyme solution and application thereof in ELISA (Enzyme-Linked Immunosorbent Assay)
  • Platinum-based alloy structured nanorod simulation enzyme solution and application thereof in ELISA (Enzyme-Linked Immunosorbent Assay)
  • Platinum-based alloy structured nanorod simulation enzyme solution and application thereof in ELISA (Enzyme-Linked Immunosorbent Assay)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1: Preparation and purification of nanorods with gold core / (copper) platinum alloy nano-island shell structure

[0050] The nanorods of the gold core / (copper) platinum alloy nano-island shell structure described in the present invention can be obtained by methods well known to those skilled in the art. In the present invention, we use gold nanorods as seeds to realize the preparation of nanostructures by reducing Pt with a reducing agent or co-deposition of Pt and Cu.

[0051] Preparation of gold core / platinum nano-island shell structure nanorods (AuPt): Take 1 mL of purified gold nanorod solution, add 75 μL of 2 mM K 2 PtCl 4 Solution, 22.5 μL of 0.1M AA solution, mixed evenly, placed in a 30°C water bath for half an hour, then added 0.5mL of 0.1M CTAB solution, centrifuged at 12000rpm for 5 minutes once, and the precipitate was dispersed in deionized water for later use .

[0052] Preparation of nanorods (AuPtCu) with gold core / copper-platinum alloy nano-is...

Embodiment 2

[0054] Embodiment 2: Enzyme kinetic parameter analysis

[0055] The reaction kinetics was investigated by monitoring the change of the absorbance of the TMB oxidation product. Using Varian Cary50 kinetic mode, every 0.1min interval measurement. The test conditions are: the concentration of AuPt or AuPtCu nanorods is 15pM, 2.4mL of 0.1M pH4.5 phosphate buffer solution, and the reaction temperature is set at 30°C. When using TMB as a substrate, H 2 o 2 The concentration is fixed at 2mM; when H 2 o 2 As the substrate, the concentration of TMB was fixed at 0.13mM. The apparent kinetic parameters are from the Lineweaver-Burk equation: 1 / V=(K m / V max )(1 / [C])+1 / V max get. where V is the reaction velocity, V max is the maximum reaction velocity, [C] is the substrate concentration, K m is the Michaelis constant.

[0056] The kinetic parameters obtained are shown in Table 1 and figure 2 shown.

[0057] Table 1 AuPt and AuPtCu nanorods as peroxidase-like apparent kinetic...

Embodiment 3

[0065] Example 3: Goat anti-human IgG antibody modification on the surface of nanorods with gold core / (copper) platinum alloy nano-island shell structure

[0066] Take 1 mL of the original CTAB-coated gold core / (copper) platinum alloy shell nanostructure solution that was purified once, add 50 μL of 20 mg / mL PSS solution, mix well, place it at room temperature for more than 3 hours, and then centrifuge at 12000 rpm for 5 minutes Once, the precipitate was dispersed in 200 μL deionized water for later use, and the concentration of the purified alloy nanorods was about 2 nM.

[0067] Take 1 mL of purified PSS-coated 2 nM alloy nanorod solution and disperse it in 1 mL of Tris buffer solution (0.05 M, pH 8) containing 0.05% BSA. Add 5 μL of 1 mg / mL goat anti-human IgG solution, mix well and put it in a 37°C incubator. After incubation for 30 minutes, centrifuge at 8000 rpm for 5 minutes to remove free goat anti-human IgG molecules. Pour off the supernatant, and disperse the precip...

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Abstract

The invention relates to a platinum-based alloy structured nanorod simulation enzyme solution and application thereof in ELISA (Enzyme-Linked Immunosorbent Assay). The platinum-based alloy structured nanorod simulation enzyme solution contains a gold kernel-platinum-based alloy-antibody or antigen composite structure which consists of a cylindrical gold nanorod kernel, an island-shaped platinum-based alloy structure and an antibody or antigen, wherein the outside surface of the cylindrical gold nanorod kernel is coated with the island-shaped platinum-based alloy structure, and the outside surface of the cylindrical gold nanorod kernel is coated with the antibody or antigen. When applied to the ELISA, the platinum-based alloy structured nanorod simulation enzyme solution disclosed by the invention has the advantages that the response range is wide in detection on the antibody or antigen, the sensitivity is high, and the detection limit can reach the level of pg / mL. Particularly, when a gold kernel / copper-platinum alloy shell nanorod is used, the separation of binding sites and the catalysis sites can be realized, and coexisting protein has not obvious influence on the catalytic activity of nanostructures.

Description

technical field [0001] The invention relates to a platinum-based alloy structure nanorod and its application, in particular to a platinum-based alloy structure nanorod simulated enzyme solution and its application in ELISA. Background technique [0002] Enzyme-linked immunoassay (ELISA) is a widely recognized and powerful method for detecting proteins. The method has high sensitivity and standard operating procedures, and usually uses horseradish peroxidase (HRP)-labeled immunoreagents to generate signals to detect target molecules. Although it has the advantages of various methods and high sensitivity, it also has some disadvantages. For example, large amounts of expensive proteins are required, many incubation steps are required for diffusion-controlled reactions, and many washing steps are required. In the past few years, people have mainly devoted themselves to the research on how to improve the detection range, sensitivity and specificity recognition of ELISA. [000...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/558G01N33/531
CPCG01N33/531G01N33/54306G01N33/558G01N33/68
Inventor 吴晓春阿迪蒂亚萨兰胡晓娜张会
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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