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Detection method for inhibiting lymphocyte proliferation in vitro by mesenchymal stem cells

A lymphocyte proliferation and mesenchymal stem cell technology, applied in the field of detection of mesenchymal stem cells inhibiting lymphocyte proliferation in vitro, can solve problems such as instability, large differences in immune regulation effects, and differences in the quality of mesenchymal stem cells

Pending Publication Date: 2021-04-27
华夏源细胞工程集团股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, there are still some problems to be solved urgently in the application of mesenchymal stem cells in the field of immune regulation
First of all, due to the differences in the methods of cell culture proliferation and de-proliferation, the quality of mesenchymal stem cells is also very different, which leads to great differences in the immune regulation effect of mesenchymal stem cells in different laboratories
In addition, the current detection method for the inhibition of lymphocyte proliferation by mesenchymal stem cells in vitro has the characteristics of large fluctuations in data and instability.

Method used

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  • Detection method for inhibiting lymphocyte proliferation in vitro by mesenchymal stem cells
  • Detection method for inhibiting lymphocyte proliferation in vitro by mesenchymal stem cells
  • Detection method for inhibiting lymphocyte proliferation in vitro by mesenchymal stem cells

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Embodiment 1

[0059] Embodiment 1 of the present invention provides a detection method for mesenchymal stem cells inhibiting lymphocyte proliferation in vitro, specifically as follows:

[0060] (1) MSC cell inoculation: take 400g cell suspension, centrifuge for 5min, remove the supernatant, add Gibco 1640 medium containing 10wt% fetal bovine serum to resuspend, sample and count; according to the counting result, adjust the cell concentration to 2×10 5 / mL; Inoculate the adjusted cell suspension in three 24-well culture plates, inoculate 4 wells in each plate, 500 μL in each well, and inoculate 12 wells in total. Place the inoculated cell plates in a carbon dioxide incubator (CO 2 The concentration is set to: 5.0vol%, the temperature is set to: 37.0°C), and cultured for 20 hours;

[0061] (2) MSC cell deproliferation treatment: discard the medium in each well in step (1), add 500 μl of Gibco 1640 medium containing 8 μg / mL mitomycin C to each well and contain 10wt% fetal bovine serum, and pl...

Embodiment 2

[0065] Embodiment 2 of the present invention provides a detection method for mesenchymal stem cells inhibiting lymphocyte proliferation in vitro, specifically as follows:

[0066] (1) MSC cell inoculation: take 400g cell suspension, centrifuge for 5min, remove the supernatant, add Gibco 1640 medium containing 10wt% fetal bovine serum to resuspend, sample and count; according to the counting result, adjust the cell concentration to 2×10 5 / mL; Inoculate the adjusted cell suspension in three 24-well culture plates, inoculate 4 wells in each plate, 500 μL in each well, and inoculate 12 wells in total. Place the inoculated cell plates in a carbon dioxide incubator (CO 2 The concentration is set to: 5.0vol%, the temperature is set to: 37.0°C), and cultivated for 24 hours;

[0067] (2) MSC cell deproliferation treatment: discard the medium in each well in step (1), add 500 μl of Gibco 1640 medium containing 12 μg / mL mitomycin C to each well and contain 10wt% fetal bovine serum, and...

Embodiment 3

[0071] Embodiment 3 of the present invention provides a detection method for mesenchymal stem cells inhibiting lymphocyte proliferation in vitro, specifically as follows:

[0072] (1) MSC cell inoculation: take 400g cell suspension, centrifuge for 5min, remove the supernatant, add Gibco 1640 medium containing 10wt% fetal bovine serum to resuspend, sample and count; according to the counting result, adjust the cell concentration to 2×10 5 / mL; Inoculate the adjusted cell suspension in three 24-well culture plates, inoculate 4 wells in each plate, 500 μL in each well, and inoculate 12 wells in total. Place the inoculated cell plates in a carbon dioxide incubator (CO 2 The concentration is set to: 5.0vol%, the temperature is set to: 37.0°C), and cultured for 22 hours;

[0073] (2) MSC cell deproliferation treatment: suck and discard the medium in each well in step (1), add 500 μl of Gibco 1640 medium containing 10wt% fetal bovine serum containing 10 μg / mL mitomycin C to each wel...

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Abstract

The invention relates to the field of cell culture and detection, in particular to a detection method for inhibiting lymphocyte proliferation in vitro by mesenchymal stem cells. The detection method comprises the following steps: (1) MSC (mesenchymal stem cell) inoculation: taking an MSC suspension, centrifuging the MSC suspension for n times to remove supernate, carrying out resuspending by using a complete culture medium, and then conducting inoculated culture, wherein n is larger than or equal to 1; (2) proliferation removal treatment of the MSCs: conducting proliferation removal on the MSCs finally obtained in the step (1), and performing continuous culture; and (3) lymphocyte proliferation inhibition detection: carrying out co-culture on the MSCs finally obtained in the step (2) and the stained PBMCs (peripheral blood mononuclear cells), and recording the lymphocyte proliferation ability. The detection method for inhibiting lymphocyte proliferation in vitro by the MSCs is simple to operate, stable in detection data and high in lymphocyte proliferation inhibition rate.

Description

technical field [0001] The present invention relates to the fields of cell culture and detection, and more specifically, the present invention relates to a detection method for mesenchymal stem cells inhibiting lymphocyte proliferation in vitro. Background technique [0002] Mesenchymal stem cells are an important class of adult stem cells derived from the mesoderm in early development. They have a variety of tissue repair capabilities and have been widely used in clinical treatment research. Mesenchymal stem cells were originally non-hematopoietic pluripotent stem cells isolated from bone marrow stroma. It has been confirmed that mesenchymal stem cells also exist in various tissues such as adipose tissue and umbilical cord. Studies have shown that mesenchymal stem cells are an important application in the field of immune regulation. A large number of studies have confirmed that mesenchymal stem cells can induce immunosuppression in vivo, reduce GVHD and rejection during he...

Claims

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Application Information

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IPC IPC(8): C12N5/078C12Q1/02
CPCC12N5/0634C12N2502/1352G01N33/5005
Inventor 朱灏
Owner 华夏源细胞工程集团股份有限公司
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