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Preparation and identification method for aldose reductase-like protein (ARL-1) monoclonal antibody

A monoclonal antibody, reductase technology, applied in the fields of botanical equipment and methods, biochemical equipment and methods, applications, etc., can solve problems such as poor specificity

Inactive Publication Date: 2004-11-10
SOUTHEAST UNIV
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Problems solved by technology

However, due to the poor specificity of polyclonal antibodies, there is cross-reactivity, so it is necessary to prepare anti-ARL-1 monoclonal antibodies to detect ARL-1 antigen in peripheral blood of patients for early diagnosis of liver cancer

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Embodiment Construction

[0019] Below are embodiments of the present invention:

[0020] Aldose reductase-like gene (ARL-1) is a liver cancer-related gene, and the ARL-GST recombinant protein is expressed and purified by gene recombination. Splenocyte suspension was prepared from ARL-GST immunized mice, fused with Sp2 / 0 myeloma cells in the logarithmic growth phase under the action of polyethylene glycol (PEG), and hybridoma antibody detection and several times of cloning were performed to obtain A cell line stably secreting anti-ARL-1 monoclonal antibody was named F9. It was determined that the anti-ARL-1 monoclonal antibody secreted by F9 belonged to IgG2b subtype, the light chain was κ type, and the ascites titer was 1:4×10 -5 . It has been identified that the monoclonal antibody secreted by F9 only reacts to the ARL-1 protein, but does not have any effect on AR that has a high homology with the ARL-1 protein, indicating that the above-mentioned monoclonal antibody is specifically anti-ARL-1 prot...

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Abstract

The invention is a method of preparing and identifying anti-aldose reductase similar protein monoclonal antibody, and its preparing method: using gene reformation to express and purify ARL-GST reformed protein, using ARL-GST immune mice to prepare splenic cell suspension liquor, under the action of polyglycol, fusing with Sp / 2 / 0 medullary cancer cell in logarithmic growing period, by hybridoma antibody detection and several times of clone, obtaining a cell strain of stably secreting anti-ARL-1 monoclonal antibody, named F9. The anti-ARL-1 monoclonal antibody secreted by F9 is IgG2b subtype, its light chain is k-type, and its ascite valence is 1 : 4X10 to the power -5. By identification, the monoclonal antibody secreted by F9 only reacts with ARL-1 protein and have no effect on all aldose reductases having higher isogeny with the ARL-1 protein, thus showing the above monoclonal antibody is a specific anti-ARL-l protein monoclonal antibody.

Description

technical field [0001] The invention relates to a preparation method of a monoclonal antibody, which belongs to the technical field of bioengineering development of new diagnostic reagents for liver cancer. Background technique [0002] Primary liver cancer is a common malignant tumor in my country, with a high mortality rate and a five-year survival rate of 2.2-2.3%, which seriously threatens the health of our people. [0003] Liver cancer has a relatively long natural development process, and its natural course is at least 24 months: it usually takes 10 months from the appearance of low concentration of AFP to the establishment of the diagnosis of subclinical liver cancer as the early stage; from the subclinical stage to the appearance of clinical symptoms is the middle stage. The period is 8 months; from the onset of clinical symptoms to the appearance of jaundice, ascites, distant metastasis or dyscrasia is called the late stage, and the time is ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12P21/08G01N33/577
Inventor 谢维金俊飞张建琼单军
Owner SOUTHEAST UNIV
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