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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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, chemiluminescence is a dynamic signal, and the sign

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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

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[0035] Example 1:

[0036] Double antibody sandwich method for detecting multiple tumor markers

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

[0038] 1. 12 monoclonal antibodies labeled with infrared fluorescence

[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 and activated according to 7μL NHS Succinimidyl ester IRDye800 near-infrared fluorescent dye (LI-COR, USA) labeled with 1mg monoclonal antibody ratio. Mix the antibody and fluorescent dye, and then add 1 / 10 volume of 1M NaHCO 3 , After mixing, react at room temperature and avoid light for 1 hour. After the reaction, the marker was placed in a 10K dialysis bag and dialyzed with PBS at 4°C overnight to remove free fluorescent dyes. Add the final concentration of 1M BSA and 2mM sodium azide to the solution, and...

Example Embodiment

[0077] Example 2: Indirect method to detect multiple autoantibodies

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

[0079] The goat anti-human IgG antibody was dialyzed with PBS, and the antibody and the fluorescent dye were mixed according to the ratio of 7μL NHS-activated succinimidyl ester IRDye800 near-infrared fluorescent dye (LI-COR company, USA) to label 1mg antibody, and then add 1 / 10 volume of 1M NaHCO 3 , After mixing, react at room temperature and avoid light for 1 hour. After the reaction, the markers were placed in a 10K dialysis bag and dialyzed overnight with PBS at 4°C to remove free fluorescent dyes. Add the final concentration of 1M BSA and 2mM sodium azide to the solution, and store 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 (SSA, SSB, Scl-70, Jo-1, Sm, RNP).

[0082] Spots are A1, A2-SSA...

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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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