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Method for separating out and purifying Kupffer cells in livers

A separation and purification, liver technology, applied in the medical field, can solve the problems of complex operation, difficult technical mastery, cell activity and the impact of subsequent experiments.

Inactive Publication Date: 2016-05-18
NANJING AIRUI BIOLOGICAL TECH CO LTD
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  • Abstract
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Problems solved by technology

In summary, although mechanical perfusion and density gradient centrifugation can obtain Kupffer cells with higher purity and activity, they have the following disadvantages: 1. The equipment of density gradient centrifugation is relatively expensive, and some laboratories cannot meet the experimental conditions; Second, the sorting time is long, which may have an impact on cell viability and subsequent experiments; third, the operation is more complicated and the technology is more difficult to master

Method used

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  • Method for separating out and purifying Kupffer cells in livers
  • Method for separating out and purifying Kupffer cells in livers
  • Method for separating out and purifying Kupffer cells in livers

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Experimental program
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Embodiment

[0016] Experimental Materials

[0017] 1.1 Mouse selection

[0018] C57BL / 6 mice were obtained from the Animal Experiment Center of Nanjing Medical University.

[0019] 1.2 Required reagents and experimental equipment

[0020] DMEM / Ham'sF-121:11×medium (Hyclonecat#SH30023.01), Pronase (BoehringerMannheimcat#1459643), Collagenase (typeIV, sigmacat#C3155), DNase (sigmacat#:D4527), CD11cPE flow antibody (MiltenyiBiotecOrderNO130 -097-982), Anti-PEMicrobeads magnetic beads (MiltenyiBiotecMat120-000-294), Anti-FITCMicrobeads magnetic beads (MiltenyiBiotecMat120-000-742), F4 / 80APC flow cytometry antibody (MiltenyiBiotecOrderNO130-099-435), Anti-APCMicrobeads ( MiltenyiBiotecOrderNO130-048-801), medium (RPMI1640, 10% FBS, 100mg / mlstreptomycin, 10000U / mlpenicilin, Gibco), Peristalticpump (Masterflex), Shaker shaker (GuohuaTHZ-82), LS / MScolumn magnetic column (MiltenyiBiotec), MidiMACS (MACS), Flowcytometer (BD), Cell Incubator (Thermo), Microscope (Zeiss Axiovert).

[0021] 2 meth...

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Abstract

The invention relates to a method for separating out and purifying cells, in particular to a method for separating out and purifying Kupffer cells in livers, and belongs to the technical field of medicine. By means of the method, Kupffer cells in livers are successfully separated out through mouse liver mechanical circulation pouring and immunomagnetic beads for the first time, the Kupffer cells obtained through the method are high in purity, good in activity, easy to operate and high in generalizability, the specific F4 / 80+CD11c- phenotype is achieved, and stronger macrophage characteristics are achieved. By means of the successful establishment of the cell sorting technology, a new experimental method is provided for further function research on Kupffer cells.

Description

technical field [0001] The invention relates to a method for separating and purifying cells, in particular to a method for separating and purifying liver Kupffer cells, and belongs to the technical field of medicine. Background technique [0002] The immune response of the liver is related to various liver diseases such as liver cancer, hepatitis virus infection and liver injury in chronic liver disease, and has received more and more attention and research. Macrophages are important immune cells in the human body, and Kupffer cells in the liver are extremely important for the immune response of the liver itself and the whole body. Liver Kupffer cells account for about 15% of the total cells in the liver and 50% of the macrophages in the whole body. They are mainly located in the sinusoidal wall of the liver. They are the cells that enterogenous pathogens first contact in the liver and are one of the important innate immune cells in the liver. , participate in the innate im...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/067C12N2509/10
Inventor 吕凌陆云杰古鉴
Owner NANJING AIRUI BIOLOGICAL TECH CO LTD
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