Method for external amplification natural killer cell

A technology of natural killer cells and in vitro amplification, applied in the field of cellular immunology, can solve the problems such as no NK cell method invention and report, and achieve the effects of low reagent cost, high amplification multiple, and less blood samples

Inactive Publication Date: 2008-12-03
BENGBU MEDICAL COLLEGE
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

After extensive review of domestic and foreign patent documents and various publications, so far, there has been no invention or report of a practical and efficient in vitro method for amplifying NK cells with a large number of advantages

Method used

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  • Method for external amplification natural killer cell
  • Method for external amplification natural killer cell
  • Method for external amplification natural killer cell

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Embodiment Construction

[0031] The present invention will be further described in detail below in conjunction with specific embodiments.

[0032] This embodiment includes the following steps:

[0033]1. Isolation of peripheral blood mononuclear cells (PBMC): In this case, 8 healthy adult volunteers (20 to 35 years old) were selected, and peripheral blood was drawn into a heparin anticoagulation tube, diluted with an equal amount of culture medium, and placed in Ficoll- On Hypaque Lymphocyte Separation Solution, conventional density gradient centrifugation is used to obtain PBMC. After washing twice, adjust the cell concentration to 2×10 with RPMI1640 culture medium containing 10% autologous serum. 6 / ml.

[0034] 2. MβCD pretreatment and cell expansion: put the PBMC suspension into a 24-well culture plate, 1ml / well; add MβCD (RPMI1640 configuration containing 10% autologous serum) to a final concentration of 0-4mmol / L. In addition, set up a LAK cell control group (add rhIL-21000U / ml), 48h later, add rhIL...

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Abstract

The invention belongs to the technology field of the cellular immunology, and particularly relates to a method for the dominant amplification of in vitro peripheral blood nature killer cells with a large amount. The method comprises the steps as follows: (1) separating single nucleus cell from human or animal peripheral blood; (2) suspending the single nucleus cell in an RPMI 1640 culture solution containing 5%-15% autologous serum and pretreating with methyl-Beta-cyclodextrin with an effective concentration of 1-4mmol/L for 36-60 hours; and (3) adding recombined interleukin 2, and amplification-culturing in an RPMI 1640 culture solution containing 5% of new-born calf serum and 5% of autologous serum for above 10 days. The method has the advantages that the blood sample amount is small; the cost is low; the operation is convenient; and the amplification multiple is high, and the nature killer cells can be amplified by 392-1752 times in a short term.

Description

Technical field [0001] The invention belongs to the technical field of cellular immunology, and specifically is a method for a large number of advantages in vitro to amplify natural killer cells in human or animal peripheral blood. Background technique [0002] Natural killer cells (NK cells) are a type of lymphocytes that do not depend on antigen stimulation and can spontaneously dissolve a variety of tumor cells and virus-infected cells. The marker molecule on the cell surface is CD3 - , CD56 + . NK cells exist in peripheral blood and spleen, and account for 15%-20% of lymphocytes in peripheral blood. NK cells have the functions of anti-tumor, anti-infection and immune regulation. They are the first line of defense of the body's immune surveillance and play an important role in the body's resistance to tumors and infections. Studying the method of rapid expansion of NK cells is a very important subject in clinical immunology practice. Because the content of NK cells in peripher...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/06C12N5/08C12N5/0783
Inventor 吕合作李柏青
Owner BENGBU MEDICAL COLLEGE
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