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Anti-SIRP alpha rabbit recombinant monoclonal antibody as well as preparation method and application thereof

A monoclonal antibody and heavy chain technology, applied in the biological field, can solve problems such as anemia and accidental injury of red blood cells

Active Publication Date: 2022-05-31
缔码生物科技(武汉)有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, CD47 is also highly expressed on the surface of red blood cell membranes. While CD47 therapeutic drugs kill tumor cells, they will accidentally injure red blood cells, resulting in anemia
In addition, due to the widespread expression of CD47, clinically high doses of antibodies are required to be effective, which are two major bottlenecks in the clinical development of CD47

Method used

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  • Anti-SIRP alpha rabbit recombinant monoclonal antibody as well as preparation method and application thereof
  • Anti-SIRP alpha rabbit recombinant monoclonal antibody as well as preparation method and application thereof
  • Anti-SIRP alpha rabbit recombinant monoclonal antibody as well as preparation method and application thereof

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Experimental program
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Embodiment

[0032] (1) Preparation of SIRPα extracellular region protein: the present invention constructed human SIRPα extracellular region (Glu31-Arg370) FC fusion protein eukaryotic expression plasmid, transfected 293F cells for 5 days, collected cell culture supernatant, and used proteinA resin Purification was performed and the concentration of the protein was identified.

[0033] (2) Acquisition of peripheral blood mononuclear cells (ie, PBMC cells) from immunized animals: New Zealand white rabbits were selected as immunized animals. For the first immunization, 250 μg of SIRPα extracellular domain protein was emulsified with an equal volume of complete Freund’s adjuvant, and injected into the subcutaneous back of New Zealand white rabbits at multiple points. The second immunization was performed 21 days later, and 120 μg of protein was mixed with an equal volume of The incomplete Freund's adjuvant was emulsified, and the New Zealand white rabbit was subcutaneously injected on the ba...

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Abstract

The invention relates to an anti-SIRP alpha rabbit recombinant monoclonal antibody as well as a preparation method and application thereof, amino acid sequences of heavy chain complementary determining regions HCDR1, HCDR2 and HCDR3 of the anti-SIRP alpha rabbit recombinant monoclonal antibody are amino acid sequences as shown in SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3 respectively, and amino acid sequences of light chain complementary determining regions LCDR1, LCDR2 and LCDR3 are amino acid sequences as shown in SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6 respectively. The anti-SIRP alpha rabbit recombinant monoclonal antibody has good specificity, can be used in ELISA (Enzyme-Linked Immunosorbent Assay) and a common flow detection method, most importantly, the antibody can block the interaction between CD47 protein and SIRP alpha protein, and has potential tumor immunotherapy value.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to an anti-SIRPα rabbit recombinant monoclonal antibody with blocking activity, a preparation method and application thereof. Background technique [0002] Signal regulatory protein α (Signal reg μ latory protein alpha, SIRPα) is a transmembrane protein expressed on the surface of macrophages and bone marrow cells, which can bind to its ligand CD47 protein to generate a "don't eat me" signal , thereby preventing the phagocytosis of healthy cells by macrophages. However, tumor cells are very cunning, and they can use this mechanism to escape the phagocytosis of macrophages. Therefore, blocking the binding of CD47 to SIRPα by CD47 antibody can activate the phagocytosis of tumor by macrophages. However, CD47 is also highly expressed on the surface of red blood cell membranes. While CD47 therapeutic drugs kill tumor cells, they will accidentally injure red blood cells, resulti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C12N15/13C12N15/85C12N5/10A61K39/395A61P35/00G01N33/574G01N33/577
CPCC07K16/2896C12N15/85A61P35/00G01N33/577G01N33/574G01N33/57492C07K2317/565C07K2317/56C07K2317/14C07K2317/76C12N2800/107A61K2039/505G01N2333/70596
Inventor 马东晖黄少梅洪淑娟蔡萌李沛梅芬
Owner 缔码生物科技(武汉)有限公司
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