Method for evaluating DNA (Deoxyribose Nucleic Acid) damages of peripheral blood lymphocytes caused by ionizing radiation

A technology for DNA damage and lymphocytes, applied in measuring devices, individual particle analysis, scientific instruments, etc., can solve problems such as poor repeatability of experiments and large differences between different individuals

Active Publication Date: 2013-02-06
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The purpose of the present invention is to solve the problems of poor experimental repeatability and large differences between different individuals in the method of analyzing the γ-H2AX protein content in peripheral blood lymphocytes by flow cytometry

Method used

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  • Method for evaluating DNA (Deoxyribose Nucleic Acid) damages of peripheral blood lymphocytes caused by ionizing radiation
  • Method for evaluating DNA (Deoxyribose Nucleic Acid) damages of peripheral blood lymphocytes caused by ionizing radiation
  • Method for evaluating DNA (Deoxyribose Nucleic Acid) damages of peripheral blood lymphocytes caused by ionizing radiation

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Experimental program
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Embodiment

[0020] 1) Take 0.9ml of peripheral blood from a healthy person using a heparin anticoagulant tube, and divide it into 3 parts, each 0.3ml, of which the first part is not irradiated, the second part is irradiated with 2Gy gamma rays, and the third part is irradiated with 6Gy gamma rays;

[0021] 2) Add 1.5ml of fixative solution (2% paraformaldehyde solution) to the blood sample and let it react for 30 minutes at room temperature;

[0022] 3) Centrifuge at room temperature (500g) for 5 minutes, remove the supernatant;

[0023] 4) Wash the cell pellet with 1.5ml PBS buffer, centrifuge (500g) for 5 minutes at room temperature, remove the supernatant, and keep the pellet;

[0024] 5) Add 1.5ml Triton-100 solution with a concentration of 0.4%, mix the cell pellet, and let it react at room temperature for 15 minutes;

[0025] 6) Centrifuge (500g) for 5 minutes at room temperature, and remove the supernatant;

[0026] 7) Add 1.5ml PBS buffer solution to wash the cells, centrifuge a...

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Abstract

The invention relates to a method for evaluating DNA (Deoxyribose Nucleic Acid) damages of peripheral blood lymphocytes caused by ionizing radiation. The method mainly comprises the following steps: irradiating peripheral blood through ionizing radiation; marking gamma-H2AX proteins in a peripheral blood lymphocyte and a granulocyte by using specific antibodies; analyzing contents of the gamma-H2AX proteins in the two cells through the flow cytometry; by taking the granulocyte as reference, figuring out relative content of the gamma-H2AX proteins in the lymphocyte; and evaluating the DNA damage degree of the ionizing radiated peripheral blood lymphocyte by using RL / G value of the gamma-H2AX (ratio between the content of the gamma-H2AX proteins in the lymphocyte and the content of the gamma-H2AX proteins in the granulocyte). The method for evaluating the DNA damages of the peripheral blood lymphocytes caused by the ionizing radiation, provided by the invention, has the advantages of simplicity in operation and stable result and can be widely applied to the field of diagnosis, cell biology research and so on.

Description

technical field [0001] The invention relates to a cell detection method, in particular to a method for evaluating DNA damage of peripheral blood lymphocytes caused by ionizing radiation. technical background [0002] Peripheral blood lymphocytes are sensitive cells to ionizing radiation and are one of the cell models for studying DNA damage caused by ionizing radiation. γ-H2AX protein is a molecule that changes after DNA damage and is a specific molecule for DNA damage. The detection of γ-H2AX protein can evaluate DNA damage, and it is widely used for studying the molecular mechanism of DNA damage. Flow cytometry has the characteristics of rapidity and high throughput, and can detect the content of γ-H2AX protein in cells. The current method of analyzing the content of γ-H2AX protein in peripheral blood lymphocytes by flow cytometry has problems such as poor experimental repeatability and large differences between peripheral blood samples from different individuals. These ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N15/14
Inventor 王治东陈英张学清
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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