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Composition for separating and detecting exotic protein of carbohydrate chain, kit, as well as method and application of composition

A technology for detecting kits and abnormal proteins, applied in biological testing, material inspection products, etc., can solve the problems of high price, limited application and false positives of mass spectrometry, improve separation efficiency and accuracy, ensure accuracy, Accurate results

Active Publication Date: 2018-11-06
BEIJING HOTGEN BIOTECH CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] 1. Mass spectrometry: the method of liquid chromatography-electrospray mass spectrometry combined with proteolysis and glycoside hydrolysis provides a fast and sensitive detection method for the analysis of glycoproteins, and can obtain the structural information of related sugar chains; mass spectrometry is expensive And the corresponding data processing after detection still needs to be improved, which limits the application
[0014] 2. Nuclear Magnetic Resonance Spectroscopy (NMR) is a method for qualitative analysis of various material components and structures by utilizing the different absorption of radio frequency radiation by atomic nuclei. It can provide information on the structure of sugar chains, but NMR has very high requirements for samples. limited clinical application
As described in Chinese patent CN207067147U, a detection device for abnormal sugar chain protein TAP is a qualitative detection, which cannot effectively prompt the relationship between tumor burden and detection results, and there are false positives
[0016] However, there is no public record about the automatic detection method of the sugar chain abnormal protein with simple operation, high specificity and high sensitivity

Method used

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  • Composition for separating and detecting exotic protein of carbohydrate chain, kit, as well as method and application of composition
  • Composition for separating and detecting exotic protein of carbohydrate chain, kit, as well as method and application of composition
  • Composition for separating and detecting exotic protein of carbohydrate chain, kit, as well as method and application of composition

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] This example is an exemplary preparation method of the composition for separating abnormal sugar chain proteins. In order to illustrate the present invention, the following examples use silicide-embedded magnetic beads for separating abnormal sugar chain proteins and lentil agglutinin (LCA) for illustration.

[0053] 1. Preparation of magnetic bead-lectin coupling complex (particle size 20μm-300μm)

[0054] Silicide-embedded separation magnetic beads and lentil agglutinin (LCA) are commercial raw materials, which can be purchased from well-known raw and auxiliary material supply companies such as sigma, or prepared by methods disclosed in literature.

[0055] Silicide-embedded separation beads and lentil agglutinin (LCA) form a magnetic bead-lectin conjugate complex by the following steps:

[0056] (1) Separate the silicide-embedded separation magnetic beads with a magnetic separation rack, and then wash with purified water or 1mM HCl solution to remove the residual sol...

Embodiment 2

[0067] This example is an exemplary preparation method of the composition for detecting abnormal sugar chain proteins. This example uses carboxyl magnetic beads, glycoprotein-selected alpha-fetoprotein (AFP), and glycoprotein-selected alpha-fetoprotein heterogeneous (AFP-L3) for illustration.

[0068] 1. Preparation of magnetic bead-antibody complex (particle size 0.1μm-20μm)

[0069] The magnetic bead-antibody complex is composed of carboxyl magnetic beads combined with anti-AFP antibody 1 to form a magnetic bead-antibody complex, which is used to detect AFP-L3 separated by the separation magnetic beads. The preparation process is as follows:

[0070] Mix the carboxyl magnetic beads, add a magnetic field to remove the supernatant, wash the magnetic beads once with coating buffer, then add the activator EDC to 10-50mg / ml, shake and react at room temperature for 0.5-2h, then remove the supernatant, and add the coating Buffer and an appropriate amount of anti-glycoprotein anti...

Embodiment 3

[0082] This example is an exemplary preparation method of a glycan protein detection composition. This example uses carboxyl magnetic beads and glycan protein to select alpha-fetoprotein (AFP) for illustration.

[0083] In order to ensure the same background in the detection process of glycoproteins and abnormal glycoproteins, and ensure the accuracy of the ratio of glycoprotein abnormalities, the exemplary preparation method of the glycoprotein detection composition is the same as that of the glycoprotein abnormality in Example 2. The exemplary preparation method of the detection composition is the same, and the raw and auxiliary materials used in the preparation process are the same, and the detailed description is as follows:

[0084] 1. Preparation of magnetic bead-antibody complex (particle size 0.1μm-20μm)

[0085] The magnetic bead-antibody complex is composed of carboxyl magnetic beads combined with anti-AFP antibody 1 to form a magnetic bead-antibody complex, which is...

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Abstract

The invention relates to a composition for separating and detecting an exotic protein of a carbohydrate chain, a kit, as well as a method and application of the composition. The diagnosis conditions of the tumor caused by the exotic protein of the carbohydrate chain and the other related diseases are indicated according to the method, and the method is highly sensitively, and is rapid, convenientand automatic.

Description

technical field [0001] The invention relates to a composition, a kit, a method and an application for the separation and detection of sugar chain abnormal disease marker protein (hereinafter referred to as: sugar chain abnormal protein), and belongs to the field of medical equipment and in vitro diagnosis. Background technique [0002] Glycoproteins are binding proteins composed of one or more oligosaccharide chains covalently linked to the polypeptide backbone of the protein. Glycoproteins are formed by glycosylation modification of proteins, and the sugar chains bind to specific sites of the peptide chains along with the extension of the newly synthesized peptide chains, prompting the peptide chains to fold and aggregate into glycoproteins with the correct spatial conformation. [0003] Abnormal sugar chain proteins are mainly formed by incomplete glycosylation or changes in the activity of glycosyltransferases. Abnormal sugar chain proteins have various manifestations: di...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 乔雍李艳召闵微林长松高琦李永胜林长青余韶华
Owner BEIJING HOTGEN BIOTECH CO LTD
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