Method for preparing antibodies from human peripheral blood memory B lymphocytes

A technology of B lymphocytes and human peripheral blood, applied in chemical instruments and methods, from serum immunoglobulins, peptides, etc., can solve the problems of long cycle and high cost of antigenic preparation, and achieve the effect of low cost

Inactive Publication Date: 2016-05-25
HOSPITAL AFFILIATED TO GUANDONG MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, it can also be prepared by genetic engineering technology. At present, therapeutic antibodies such as human-mouse chimera, human

Method used

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  • Method for preparing antibodies from human peripheral blood memory B lymphocytes
  • Method for preparing antibodies from human peripheral blood memory B lymphocytes
  • Method for preparing antibodies from human peripheral blood memory B lymphocytes

Examples

Experimental program
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Embodiment 1

[0022] Embodiment 1: Preparation of anti-rotavirus antibody

[0023] (1) Using a commercial kit (HumanMemoryBCellIsolationKit, MiltenyiBiotec), isolate memory B lymphocytes from the peripheral blood of volunteers inoculated with rotavirus, and place them in RPMI1640 culture medium containing 10% bovine serum in a 37°C carbon dioxide incubator nourish;

[0024] (2) Add an activator to the culture medium of step (1), and continue culturing for 5 days to differentiate the memory B lymphocytes into plasma cells; the activator consists of: 10 μg / ml R848, 20 ng / ml IL-2 and 10 ng / mlIL-4;

[0025] (3) Resuscitating and culturing human myeloma cell lines, cultured in RPMI1640 culture medium containing 10% bovine serum in an incubator at 37°C and 5% (V / V) carbon dioxide, as figure 1 shown;

[0026] (4) The plasma cells prepared in step (2) are fused with the recovered human myeloma cell lines in step (3) to prepare hybridoma cell lines, and the hybridoma cell lines that can secrete ...

Embodiment 2

[0040] Embodiment 2: Preparation of antinuclear antibody

[0041] (1) Using a commercial kit (HumanMemoryBCellIsolationKit, MiltenyiBiotec), isolate memory B lymphocytes from the peripheral blood of patients with systemic lupus erythematosus, in RPMI1640 culture medium containing 15% bovine serum, at 37 ° C, 5% (V / V) cultivate in the incubator of carbon dioxide;

[0042] (2) Add an activator to the culture medium of step (1), and continue culturing for 5 days to make it differentiate into plasma cells; the activator consists of: 5 μg / ml R848, 10 ng / ml IL-2 and 20 ng / ml IL-4 ;

[0043] (3) Resuscitating and culturing the human myeloma cell line, in DMEM culture medium containing 10% bovine serum, culturing in an incubator at 37°C and 5% (V / V) carbon dioxide;

[0044] (4) The plasma cells prepared in step (2) are fused with the recovered human myeloma cell lines in step (3) to prepare hybridoma cell lines, and the fusion myeloma cell lines that can secrete antinuclear antibo...

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Abstract

The invention provides a method for preparing antibodies from human peripheral blood memory B lymphocytes. The method has the advantages of being convenient, rapid and low in cost. By means of the method, a large number of the antibodies can be prepared in vitro, and the prepared antibodies are very important in preventing infection and treating immunodeficiency diseases.

Description

technical field [0001] The invention relates to a method for preparing an antibody, in particular to a method for preparing an antibody from human peripheral blood memory B lymphocytes. Background technique [0002] Antigen-specific monoclonal or polyclonal immunoglobulin preparations are very important in the prevention of infection and treatment of immunodeficiency diseases (NatRevImmunol, 2010, 10(5):301-316). At present, clinically used immunoglobulin preparations are basically isolated from volunteer plasma (BioDrugs, 2010, 24(4):211-223). Its source is limited, the quality of different batches or sources is different, and there are potential infectious factors (AdvClinExpMed, 2015, 24(1):153-159). In addition, it can also be prepared by genetic engineering technology. At present, therapeutic antibodies such as human-mouse chimera, humanized antibody, and fully human antibody have shortcomings such as antigenicity, high production cost, and long cycle (MAbs, 2015, 7( ...

Claims

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Application Information

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IPC IPC(8): C07K16/06
CPCC07K16/06C07K2317/14C07K2317/21
Inventor 潘庆军刘华锋吴平谢彤廖焕金蔡珺张莉芳彭艳霞陈秋华杨陈安宁陈锦霞王淑君吴洪銮李尚妹
Owner HOSPITAL AFFILIATED TO GUANDONG MEDICAL COLLEGE
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