Basophilic granulocyte activation and degranulation identification method

A basophil identification method technology, applied in the field of allergic disease diagnosis, can solve the problems of basophil activation, allergen-specific basophil stimulation test cannot be carried out, etc., to achieve repeatable good sex effect

Active Publication Date: 2015-07-15
HPY BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Unfortunately, to date there is no reliable and reproducible method for specifically identifying the activated, degranulated state of basophils in whole blood, leading to allergen-specific basophil priming. The test cannot be carried out
Our recent study found that if CD123, H...

Method used

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  • Basophilic granulocyte activation and degranulation identification method
  • Basophilic granulocyte activation and degranulation identification method
  • Basophilic granulocyte activation and degranulation identification method

Examples

Experimental program
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Embodiment 1

[0053] A method for identification of basophil activation and degranulation, such as Figure 1.1 , 1.2 , 1.3, 1.4, and 1.5, including the following steps:

[0054] (1) First, according to the ratio of 9:1, use 9 times of ultrapure water to dilute the concentrated erythrocyte lysate stock solution in the kit to the working state; Dilute the stock solution of phosphate buffered saline to working condition. Note: Red blood cell lysate is recommended to be prepared immediately;

[0055] (2) Number the test samples in sequence, and mark the flow sample loading tubes required for the test;

[0056] (3) Add the required antibody combination into each flow cytometry tube: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD203c and anti-human CD63 fluorescently labeled flow antibody combinations, such as FITC-labeled Anti-human CD123, APC-labeled anti-human CCR3, APC / Cy7-labeled anti-human HLA-DR, PE-labeled anti-human CD203c, PE / Cy7-labeled anti-human CD63, the amou...

Embodiment 2

[0066] A method for the identification of basophil activation, such as Figure 2.1 , 2.2 , 2.3, 2.4, including the following steps:

[0067] (1) First, according to the ratio of 9:1, use 9 times of ultrapure water to dilute the concentrated erythrocyte lysate stock solution in the kit to the working state; Dilute the stock solution of phosphate buffered saline to working condition. Note: Red blood cell lysate is recommended to be prepared immediately;

[0068] (2) Number the test samples in sequence, and mark the flow sample loading tubes required for the test;

[0069] (3) Add the required antibody combination into each flow cytometry sample tube: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD203c fluorescently labeled flow antibody combination, such as FITC-labeled anti-human CD123, APC-labeled anti-human CCR3, APC / Cy7-labeled anti-human HLA-DR, PE-labeled anti-human CD203c, the amount of each of the four antibodies is 5 μL per person to identify the...

Embodiment 3

[0079] A method for the identification of basophil degranulation, such as Figure 3.1 , 3.2 , 3.3, 3.4, including the following steps:

[0080] (1) First, according to the ratio of 9:1, use 9 times of ultrapure water to dilute the concentrated erythrocyte lysate stock solution in the kit to the working state; Dilute the stock solution of phosphate buffered saline to working condition. Note: It is recommended to prepare red blood cell lysate immediately after use.

[0081] (2) Number the test samples in a certain order, and mark the flow sample loading tubes required for the test;

[0082] (3) Add the required antibody combination into each flow cytometry sample tube: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 fluorescently labeled flow antibody combination, such as FITC-labeled anti-human CD123, APC-labeled anti-human CCR3, APC / Cy7-labeled anti-human HLA-DR, PE-labeled anti-human CD63, the amount of each of the four antibodies is 5 μL per person ...

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Abstract

The invention provides a basophilic granulocyte activation and degranulation identification method. The basophilic granulocyte activation and degranulation identification method comprises the following steps that test samples are numbered in sequence, and flow type sample feeding pipes required by testing are marked; a required anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD203c and/or anti-human CD63 fluorescence labeling flow type antibody combination is added into each flow type sample feeding pipe; mixed whole blood is added into the marked flow type pipes, and shading incubation is conducted at the indoor temperature; red blood cell lysate is added into each sample pipe, and shading incubation is conducted at the indoor temperature again; supernatant is removed in a centrifugal mode after incubation; detection is conducted by means of a flow cytometry, and the number of basophilic granulocytes and the average fluorescence intensity are analyzed. According to the basophilic granulocyte activation and degranulation identification method, 80%-100% of the basophilic granulocytes in peripheral blood can be distinguished from other types of cells under the condition that the basophilic granulocytes do not need to be separated or purified, the basophilic granulocyte activation and/or degranulation state can be identified, whole blood of no more than 100 microlitres is needed by each sample to be tested, and the repeatability is good.

Description

technical field [0001] The invention relates to the technical field of allergic disease diagnosis, in particular to an identification method for basophil activation and degranulation. Background technique [0002] The incidence of allergic diseases accounts for more than 30% of the world's total population, and is listed by the World Health Organization as one of the four major non-infectious diseases in the 21st century. With the development of industrial economy and the change of ecological environment, such diseases have been increasing in recent years, becoming common and frequently-occurring diseases, which is a major problem that needs to be solved in the field of health and economic development in our country. [0003] Historically, most of the definitions and diagnostic schemes of diseases were proposed by developed countries. In 2011, He Shaoheng and Zhang Huiyun took the lead in revising and discussing the definition of allergic diseases that had been used for nea...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N15/14G01N33/53G01N33/56966G01N2015/149G01N2800/24
Inventor 何韶衡
Owner HPY BIOTECH CO LTD
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