Detection method of human peripheral blood lymphocytes

A technology of lymphocytes and human peripheral blood, applied in the detection field of human peripheral blood lymphocytes, can solve the problems of long process of sample cell damage and apoptosis, unfavorable absolute count of lymphocytes, long operation time of lymphocytes, etc., so as to maintain the activity. , the effect of reducing manual operation and improving sensitivity

Inactive Publication Date: 2019-11-22
UB BIOTECHNOLOGY ZHEJIANG CO LTD
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  • Abstract
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Problems solved by technology

However, the method disclosed in the above-mentioned patent can only output the positive percentage of each subgroup, and the result is relatively single, while the simple positive percentage is only a relative count, such as the overall cell number of lymphocytes has decreased, that is, the absolute number has decreased, but the relative Group percentages likely to remain unchanged
In addition, the detection method of the above-mentioned patents takes a long time to extract lymphocytes, which is not conducive to the absolute count of lymphocytes. At the same time, it will lead to a long process of damage and apoptosis of sample cells, and the detection sensitivity is low, which affects the detection results.

Method used

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  • Detection method of human peripheral blood lymphocytes
  • Detection method of human peripheral blood lymphocytes
  • Detection method of human peripheral blood lymphocytes

Examples

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Embodiment 1

[0041] Example 1: Detection of T lymphocyte subsets expressed in human peripheral blood mitochondria

[0042] The detection reagent of the T lymphocyte subpopulation expressed by the human peripheral blood mitochondria of the present invention comprises: hemolysin (10×), mitochondrial fluorescent dye, CD3APC, CD4PE-Cy7, CD8PE, CD45APC-Cy7 monoclonal antibody.

[0043] (1) Preparation of reagents for T lymphocyte subsets expressed in human peripheral blood mitochondria

[0044] 1. Hemolysin

[0045] Dilute 10× hemolysin with deionized water to form 1× working solution (1× hemolysin), and prepare according to the amount calculated by adding 0.5 mL of working solution to 50 μL of human peripheral blood.

[0046] To prepare 1L of 10×hemolysin, 82.9g of ammonium chloride, 10g of potassium bicarbonate, 372mg of disodium EDTA, and 20mL of fetal calf serum (FBS) are required.

[0047] 2. Mitochondrial Fluorescent Dye and Monoclonal Antibody

[0048] Mitochondrial fluorescent dye pr...

Embodiment 2

[0058] Example 2: Detection of NK lymphocyte subsets expressed in human peripheral blood lysosomes

[0059] The NK lymphocyte subset detection reagent expressed in human peripheral blood lysosomes of the present invention includes: hemolysin (10×), lysosome fluorescent dye, CD3APC, CD56PE, CD16-PE / Cy7, CD45-APC-Cy7 monoclonal Antibody.

[0060] (1) Preparation of reagents for NK lymphocyte subsets expressed in human peripheral blood lysosomes

[0061] 1. Hemolysin

[0062] Dilute 10× hemolysin with deionized water to form 1× working solution (1× hemolysin), and prepare according to the amount calculated by adding 2 mL of working solution to 100 μL of human peripheral blood.

[0063] To prepare 1L of 10×hemolysin, 82.9g of ammonium chloride, 10g of potassium bicarbonate, and 372mg of disodium EDTA are required.

[0064] 2. Lysosomal Fluorescent Dye and Monoclonal Antibody

[0065] Preparation of lysosomal fluorescent dye (for 1 person): add 1 μL of lysosomal fluorescent dye...

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Abstract

The invention discloses a detection method of human peripheral blood lymphocytes. The preparation method comprises the following steps of: taking human anticoagulation peripheral blood; adding anti-human CD3, CD4, CD8 and CD45 monoclonal antibodies and anti-human CD3, CD56, CD16 and CD45 monoclonal antibodies, then adding a hemolysin working solution containing a fluorescent probe to respectivelyobtain the percentage and absolute count value of a human peripheral blood T lymphocyte subset and an NK lymphocyte subset and achieve the detection of human peripheral blood lymphocytes. The method has the advantages of simplicity and convenience in operation, long sample stable preservation time, high accuracy, multi-index output and the like.

Description

technical field [0001] The invention belongs to the technical field of cell detection, in particular to a detection method of human peripheral blood lymphocytes. Background technique [0002] Flow cytometry (Flow Cytometry, FCM) is a high-tech technology developed in the 1970s. It integrates computer technology, laser technology, fluid mechanics, cytochemistry, and cellular immunology. It also has the functions of analyzing and sorting cells. The detection principle is to fluorescently label the cells to be tested and make a single-cell suspension, and analyze the physical and chemical characteristics of the cells according to the scattered light signals and fluorescence signals of the cells in the liquid flow after receiving laser irradiation, such as cell size and the information of the particles inside the tested cells Wait. It can not only measure cell size and internal particle properties, but also detect cell surface and cytoplasmic antigens, intracellular DNA, RNA co...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N15/14G01N33/577
CPCG01N15/14G01N33/577
Inventor 何淼李国平张涛
Owner UB BIOTECHNOLOGY ZHEJIANG CO LTD
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