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Anti-myoglobin antibody and kit

A myoglobin and antibody technology, applied in the field of anti-myoglobin antibodies and kits

Pending Publication Date: 2022-01-14
DONGGUAN PENGZHI BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, there are some deficiencies in the sensitivity, specificity and affinity of monoclonal antibodies used to detect myoglobin, and there is still much room for improvement. There is still a strong demand for monoclonal antibodies to detect myoglobin in this field

Method used

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  • Anti-myoglobin antibody and kit
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  • Anti-myoglobin antibody and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0112] In this example, the restriction endonuclease and Prime Star DNA polymerase were purchased from Takara Company. MagExtractor-RNA extraction kit was purchased from TOYOBO Company. BD SMART TM RACE cDNA Amplification Kit was purchased from Takara Company. The pMD-18T vector was purchased from Takara Company. Plasmid extraction kit was purchased from Tiangen Company. Primer synthesis and gene sequencing were performed by Invitrogen.

[0113] 1 Construction of recombinant plasmids

[0114] (1) Antibody gene preparation

[0115] The mRNA was extracted from the hybridoma cell line (11D2) secreting anti-myoglobin monoclonal antibody, and the DNA product was obtained by RT-PCR method, and the product was inserted into the pMD-18T vector after adding A reaction with rTaq DNA polymerase, Transform into DH5α competent cells, and after the colonies grow out, take 4 clones of Heavy Chain and Light Chain gene clones and send them to a gene sequencing company for sequencing.

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Embodiment 2

[0136] Antibody performance testing

[0137] (1) Activity detection of the antibody and its mutants in Example 1

[0138] Further analysis, the heavy chain variable region of the myoglobin monoclonal antibody (WT) of Example 1 is shown in SEQ ID NO: 12, wherein, wherein, the amino acid sequence of each complementarity determining region is as follows:

[0139] CDR1-VH: G-Y-S(X1)-F-T-S(X2)-Y-I(X3)-M-H;

[0140] CDR2-VH: Y-I(X1)-H(X2)-P-Y-Q(X3)-Y-D-T-K-Y-N-D(X4)-K-F;

[0141] CDR3-VH: A(X1)-R-G-G-N(X2)-F;

[0142] Its light chain variable region is shown in SEQ ID NO: 11, wherein the amino acid sequence of each complementarity determining region of the light chain is as follows:

[0143] CDR1-VL: S(X1)-A-S-S-S-V(X2)-Y-P-S-Y-I(X3)-Y;

[0144] CDR2-VL: T-T-S-Q(X1)-I(X2)-A-S;

[0145] CDR3-VL: H(X1)-Q-W(X2)-H-R-S-R.

[0146] On the basis of the myoglobin monoclonal antibody in Example 1, mutations were carried out at sites related to antibody activity in the complementarity d...

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Abstract

The invention discloses an anti-myoglobin antibody and a kit, and relates to the technical field of antibodies. The antibody or a functional fragment thereof disclosed by the invention comprises a heavy chain complementarity determining region and a light chain complementarity determining region. The antibody or the functional fragment thereof has better binding capacity and better affinity to myoglobin; and the kit using the antibody or the functional fragment thereof to detect the myoglobin has better specificity and sensitivity.

Description

technical field [0001] The present invention relates to the technical field of antibodies, in particular to an anti-myoglobin antibody and a kit. Background technique [0002] Myoglobin (myoglobin globin, Mb) is a protein that stores and distributes oxygen in mammalian cells (mainly muscle cells), and its oxygen saturation curve is hyperbolic. It is composed of a polypeptide chain and a prosthetic heme, with a relative molecular mass of 16700 and 153 amino acid residues. The apomyoglobin without heme is called globin (globin), which has obvious homology with the subunits of hemoglobin (a-globin chain and p-globin chain) in amino acid sequence, and their conformation and The functions are also very similar. [0003] Most of the hydrophobic side chains on the amino acid residues in the polypeptide chain of myoglobin are inside the molecule, and the hydrophilic side chains are mostly located on the surface of the molecule, forming a compact spherical three-dimensional structu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/18C12N15/13G01N33/68
CPCC07K16/18G01N33/6887C07K2317/56C07K2317/567C07K2317/565C07K2317/92C07K2317/94G01N2333/805G01N2800/324
Inventor 崔鹏何志强孟媛钟冬梅叶颖覃婷王晨
Owner DONGGUAN PENGZHI BIOTECH CO LTD