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Determination reagent, kit and quantitative method for human asialoglycoprotein receptor

The technology of a sialoglycoprotein and a kit is applied in the field of assay reagents for human asialoglycoprotein receptors, which can solve the problems of cumbersome production and preparation, difficulty in obtaining effective antibodies, and the lack of a homogeneous detection kit.

Pending Publication Date: 2022-07-22
江西乐成生物医疗有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above-mentioned method kits all use the solid-phase method of monoclonal antibody, and need to label the detection antibody. The production and preparation are cumbersome, and they are not suitable for the application of high-throughput automatic biochemical analyzers.
[0008] Due to the simple structure of sH2a protein, strong homology and similar structure with H1 and H2b, it is difficult to obtain effective antibodies suitable for immunoturbidimetric platforms, so no homogeneous detection kits for automatic biochemical analyzers have been found yet

Method used

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  • Determination reagent, kit and quantitative method for human asialoglycoprotein receptor
  • Determination reagent, kit and quantitative method for human asialoglycoprotein receptor
  • Determination reagent, kit and quantitative method for human asialoglycoprotein receptor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] A quantitative method of human asialoglycoprotein receptor for non-disease diagnostic and therapeutic purposes, comprising the following steps:

[0041] (1) Mix and react the sample with a first reagent containing buffer solution, anti-interference agent A, dispersant and coagulant to obtain a first reaction solution, and the first reagent is used to exclude high-concentration chyle in the sample and lipid interference, and disperse exposure of the tested protein in the sample;

[0042](2) mixing and reacting the first reaction solution with a second reagent containing a polyclonal antibody linked to an insoluble carrier and a monoclonal antibody against the sH2a specific domain EGHRG pentapeptide to obtain a second reaction solution;

[0043] (3) Read the absorbance value of the second reaction solution at a wavelength of 600 nm, and calculate the content of human asialoglycoprotein receptor.

[0044] It should be noted that the instrument used in the above quantitati...

Embodiment 2

[0096] 1. Standard sample

[0097] The following buffer components were used to prepare standard samples as calibration solution, one of the same three parts was sealed and placed at 2-8 °C as a control, and the other two were opened and placed at 2-10 °C and one was sealed at 37 °C. One copy, the results are shown in Table 1, the stability of the calibration solution meets the requirements of daily testing.

[0098] Calibration solution components:

[0099] 50mmol / L, pH7.4 glycine peptide buffer;

[0100] 50.0g / L bovine serum albumin;

[0101] 35mmol / L glutamic acid;

[0102] 5.0mmol / L EDTA.2Na;

[0103] 0.9% sodium chloride;

[0104] 200g / L sucrose;

[0105] 10.0g / L polyethylene glycol 6000;

[0106] 1.0L pure water;

[0107] 0.1% sodium azide;

[0108] Pure sH2a was added to the above buffer.

[0109] Table 1

[0110] 2-8°C airtight 2-10℃ open bottle for 1 day 37°C airtight for 7 days 37°C airtight for 15 days 57ng / mL 55ng / mL 60ng / mL 62...

Embodiment 3

[0138] 1. Reagents

[0139] Reagent 1 as first reagent (R1)

[0140] 70mmol / L, pH7.50 Tris buffer;

[0141] 30mmol / L succinic acid;

[0142] 100mmol / L sodium bromide;

[0143] 35mmol / L sodium glutamate;

[0144] 15.0g / L PEG 20000;

[0145] 0.3% Tween-20;

[0146] 0.02% BSA;

[0147] 0.03% sodium azide.

[0148] Reagent 2 as second reagent (R2)

[0149](1) Anti-sH2a antibody-BSA linking process steps:

[0150] (1.1) Mix the anti-sH2a polyclonal antibody with an antibody concentration of 6.0 mg / mL with BSA in a 20 mmol / L, pH 6.10 MES buffer according to a mass ratio of 2:3;

[0151] (1.2) Add EDC dissolved in the same MES buffer, stir at room temperature for 20 minutes, and the final concentration of EDC is 50% of the BSA concentration;

[0152] (1.3) Mix the anti-sH2a monoclonal antibody with an antibody concentration of 1.0 mg / mL with BSA in a 20 mmol / L, pH 6.10 MES buffer according to a mass ratio of 2:3;

[0153] (1.4) Add EDC dissolved in the same MES buffer, sti...

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Abstract

The present invention provides a human asialoglycoprotein receptor determination reagent, which comprises the following components: a first reagent, which comprises a buffer solution, an anti-interference agent A, a dispersant and a coagulation accelerator; and the second reagent comprises a polyclonal antibody connected with an insoluble carrier and a monoclonal antibody of an anti-sH2a specific structural domain EGHRG pentapeptide. A buffer solution, an anti-interference agent A, a dispersing agent and a coagulant are combined for use, and are matched with a polyclonal antibody connected with an insoluble carrier and a monoclonal antibody of anti-sH2a specific structural domain EGHRG pentapeptide, so that the rapid detection of the human asialoglycoprotein receptor in a sample is realized, and meanwhile, the precision of a determination result is ensured.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a determination reagent, a kit and a quantitative method for a human asialoglycoprotein receptor. Background technique [0002] The human asialoglycoprotein receptor (ASGPR) is a membrane heteropolymer that is only expressed in hepatocytes. The soluble secreted form sH2a is not produced by shedding at the cell surface, but by intracellular cleavage of its membrane-bound precursor, which is encoded by an alternatively spliced ​​form of the receptor H2 subunit. [0003] ASGR2 (asialoglycoprotein receptor 2) is one of the two subunits of hetero-oligomeric ASGPR, which is specifically expressed on hepatocytes. It is also called H2 and has high homology with another subunit called H1. ASGPR is present on the plasma membrane of hepatocytes, towards the sinusoidal side, and undergoes endocytic cycles. The receptor is a type II membrane protein, therefore, ASGR2 has an N-...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/536G01N33/577G01N21/31
CPCG01N33/536G01N33/577G01N21/31
Inventor 万蒙王学忠
Owner 江西乐成生物医疗有限公司
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