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Multi-index parallel-detection protein chip detection kit, preparation method and detection method

A detection kit, protein chip technology, applied in biological testing, measurement devices, material inspection products, etc., can solve problems such as quantitative inaccuracy, and achieve the effect of easy detection process, high design and manufacturing requirements, and easy automatic detection.

Inactive Publication Date: 2014-10-29
上海铭源数康生物芯片有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, chemiluminescence is a dynamic signal, and the signal changes with time, resulting in inaccurate quantification

Method used

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  • Multi-index parallel-detection protein chip detection kit, preparation method and detection method
  • Multi-index parallel-detection protein chip detection kit, preparation method and detection method
  • Multi-index parallel-detection protein chip detection kit, preparation method and detection method

Examples

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Effect test

Embodiment 1

[0036] Detection of Multiple Tumor Markers by Double Antibody Sandwich Method

[0037] Detection of multiple tumor markers (including CA72-4, SCC, c-PSA, NSE, CA125, CK19, CA19-9, AFP, CEA, CA242, β-HCG, CA15-3 and other 12 tumor markers)

[0038] 1. Infrared fluorescent labeling of 12 kinds of monoclonal antibodies

[0039] 12 kinds of monoclonal antibodies (including CA72-4, SCC, c-PSA, NSE, CA125, CK19, CA19-9, AFP, CEA, CA242, β-HCG, CA15-3) were dialyzed with PBS respectively, followed by 7μL NHS-activated Succinimidyl ester IRDye800 near-infrared fluorescent dye (LI-COR, USA) labeling ratio of 1mg monoclonal antibody Mix antibody with fluorescent dye, then add 1 / 10 volume of 1M NaHCO 3 , after mixing evenly, react at room temperature in the dark for 1 hour. After the reaction, the labeled substance was put into a dialysis bag with a pore size of 10K, and dialyzed against PBS at 4°C overnight to remove free fluorescent dye. The solution was added with a final concentra...

Embodiment 2

[0077] Example 2: Indirect method for detection of various autoantibodies

[0078] 1. Infrared fluorescence labeled goat anti-human IgG antibody

[0079] The goat anti-human IgG antibody was dialyzed with PBS, and the antibody was mixed with the fluorescent dye according to the ratio of 7 μL of NHS-activated succinimide ester IRDye800 near-infrared fluorescent dye (LI-COR, USA) to label 1 mg antibody, and then 1 / 10 volume of 1M was added. NaHCO 3 , after mixing evenly, react at room temperature in the dark for 1 hour. After the reaction, the labeled substance was put into a dialysis bag with a pore size of 10K, and dialyzed against PBS at 4°C overnight to remove free fluorescent dye. The solution was added with a final concentration of 1M BSA and 2mM sodium azide, and stored at 2-8°C in the dark.

[0080] 2. Preparation of Protein Chip

[0081] Solid phase carrier: nitrocellulose membrane. Proteins used on the solid phase carrier: 6 autoantigens with detection activity (S...

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Abstract

The invention relates to a multi-index parallel-detection protein chip detection kit based on a nitrocellulose membrane and a near-infrared fluorescent molecular marker. The multi-index parallel-detection protein chip detection kit comprises a micro-array protein chip, wherein the micro-array protein chip is formed by adopting the nitrocellulose membrane as a solid substrate and fixing different first proteins in different areas on the solid substrate; each first protein can be specifically combined with a disease marker in the body to form an immune complex, a second protein which can be specifically combined with the immune complex is marked by utilizing the near-infrared fluorescein molecules, and the second protein and the immune complex form a marker immune complex. An infrared fluorescent signal is a static signal and is not time-varied, and the signal and target protein concentration are in a proportional relation, so that the infrared fluorescent signal can be used for precise quantization; moreover, simplicity in operation is achieved, and the automation of the system can be realized. The invention also discloses a preparation method and a detection method of the protein chip detection kit.

Description

technical field [0001] The invention relates to the application of near-infrared fluorescein molecular labeling in protein chip detection. In particular, it relates to a protein chip detection kit, preparation method and detection method based on nitrocellulose membrane and near-infrared fluorescent molecular markers applied to multi-indicator parallel detection. Background technique [0002] As an emerging technology for multi-indicator parallel detection developed in recent years, protein chip technology has the characteristics of high throughput and high information content, and has broad application prospects in the fields of scientific research and clinical diagnosis. However, due to some technical factors, its practical application is limited, especially in the field of clinical diagnosis, which has not been promoted as it should. For example, the choice of chip substrates includes inorganic materials such as silicon wafers and glass sheets, and organic material subst...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/533G01N33/544
CPCG01N33/6893G01N21/6402G01N33/533G01N33/57484G01N2800/7028
Inventor 朱跃为柳飞舟季海鹏喻长杰
Owner 上海铭源数康生物芯片有限公司
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