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Method for establishing immortalized lymphoblastoid cell lines and kit

A technology of lymphocytes and B lymphocytes, applied in the field of immortalized lymphoblastoid cell line establishment, can solve the problems of cell apoptosis, different methods, necrosis, etc., achieve inhibition of activation and proliferation, high transformation efficiency, and improve transformation rate effect

Inactive Publication Date: 2018-12-18
YUANJIAN BIOTECH SHANGHAI CO LTD
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Problems solved by technology

However, cyclosporin A is highly toxic and can induce apoptosis and necrosis of cells, thereby reducing the transformation efficiency of B lymphocytes
[0004] At present, there is no universally recognized and unified standard for the methodology of EB virus transformation of peripheral blood B lymphocytes, resulting in different methods used by various laboratories; in addition, there is no method for EB virus transformation of peripheral blood in the domestic market. Ready-made kits for B lymphocytes

Method used

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  • Method for establishing immortalized lymphoblastoid cell lines and kit
  • Method for establishing immortalized lymphoblastoid cell lines and kit

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Experimental program
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Embodiment 1

[0049] The method that the kit is used to transform peripheral blood B lymphocytes with Epstein-Barr virus is as follows ( figure 2 ):

[0050] 1. Preparation of EB virus:

[0051] 1) Resuscitation of B-958 cells: Take out the frozen B-958 cells in the kit, quickly place them in a 37°C water bath, and shake them from time to time to melt them as soon as possible. After thawing, take out the cryopreservation tube from the 37°C water bath, suck out the cell suspension, add it to the centrifuge tube and drop more than 10 times of 1640 complete medium (containing the following components by volume percentage: 10% fetal bovine serum, 1% chain Mycin, 1% penicillin and 1% glutamine), after mixing, centrifuge (1000rpm, 5min), discard the supernatant;

[0052] 2) Add 1640 complete medium to the cells, culture in 5% CO2, 37°C, and adjust the cell state to logarithmic phase;

[0053] 3) For cell counting, use 1640 complete medium to adjust the cell concentration to 1×10 6 / ml;

[0...

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Abstract

The invention discloses a method for establishing immortalized lymphoblastoid cell lines (LCLs). Epstein-Barr (EB) virus is utilized to transform peripheral blood B lymphocytes to establish the LCLs.According to the method, the anti-IL-2RA (CD25) monoclonal antibody is added to inhibit the activation and proliferation of T lymphocyte, thus enhancing the conversion rate of EB virus on peripheral blood B lymphocytes. The invention also discloses a kit for the EB virus to transform the peripheral blood B lymphocytes into immortalized lymphoblastoid cell lines LCLs. The kit is suitable for the EBvirus to transform human peripheral blood B lymphocytes into LCLs, the operation is simple, and the conversion efficiency is high.

Description

technical field [0001] The invention relates to a method and a kit for establishing an immortalized lymphoblastoid cell line, in particular to a method and a kit for establishing an immortalized lymphoblastoid cell line by transforming peripheral blood B lymphocytes from Epstein-Barr virus, and belongs to the technical field of biomedicine. Background technique [0002] Epstein-Barr virus (Epstein-Barr virus, EBV) for short, is a herpes-like virus. Epstein-Barr virus can transform peripheral blood B lymphocytes into a kind of lymphoblastoid cell lines (B lymphoblastoid cell lines, LCLs) that can continuously divide and proliferate indefinitely. The LCLs established by this method are an immortalized cell line that can be passed on forever; this immortalized cell line can also preserve the complete genome of the original individual who provided the blood sample, and keep its biochemical and molecular biological characteristics unchanged. Therefore, transforming peripheral bl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0781C12N5/078
CPCC12N5/0635C12N2500/70C12N2501/2302
Inventor 杜飞
Owner YUANJIAN BIOTECH SHANGHAI CO LTD
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