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A kind of anti-human cardiac troponin I antibody and its application

A technology of cardiac troponin and binding protein, which is applied in the fields of biotechnology and medicine, and can solve the problems of poor affinity and low activity

Active Publication Date: 2021-12-03
DONGGUAN PENGZHI BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Due to low activity and poor affinity, the existing cTnI antibodies cannot be well applied to the detection of cTnI protein. Therefore, there is a strong demand in this field for antibodies that can effectively and specifically bind to and detect cTnI

Method used

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  • A kind of anti-human cardiac troponin I antibody and its application
  • A kind of anti-human cardiac troponin I antibody and its application
  • A kind of anti-human cardiac troponin I antibody and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0132] 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. SMARTERTM 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. The hybridoma cell line secreting Anti-cTnI 3C7 monoclonal antibody is the existing hybridoma cell line in the applicant's laboratory, and it is recovered for use.

[0133] 1. Primers

[0134] Amplify Heavy Chain and Light Chain 5'RACE Primers:

[0135] SMARTER II A Oligonucleotide:

[0136] 5'-AAGCAGTGGTATCAACGCAGAGTACXXXXX-3';

[0137] 5'-RACE CDS Primer (5'-CDS): 5'-(T) 25 VN-3'(N=A,C,G,orT; V=A,G,orC);

[0138] Universal Primer A Mix (UPM): 5'-CTAATACGACTCACTATAGGGCAAGCAGTGGTATCAACGCAGAGT-3';

[0139] Nested Universal Primer A (NUP)...

Embodiment 2

[0167] Although the antibody of sample 1 obtained in Example 1 (with sequences such as light chain and heavy chain shown in SEQ ID NO: 11 and 12) has the ability to bind cTnI protein, the affinity and antibody activity are not ideal, so the applicant passed The antibody's light chain CDRs and heavy chain CDRs were mutated.

[0168] After analysis, the complementarity determining region (WT) of the heavy chain:

[0169] CDR-VH1 is G-A(X1)-T-F-S-G(X2)-Y-W-M-Q(X3);

[0170] CDR-VH2 is Q-L(X1)-R-N-G(X2)-P-Y-D-F-D(X3)-T-F-Y-S;

[0171] CDR-VH3 is T-P(X1)-H-F-V(X2)-H;

[0172] Complementarity-determining regions of the light chain:

[0173] CDR-VL1 is S-N(X1)-S-L-F-N(X2)-S-G-N-Q-R(X3)-N-Y-L-A;

[0174] CDR-VL2 is G-P(X1)-S-T-R(X2)-E-F;

[0175] CDR-VL3 is Q-T(X1)-D-R(X2)-T-S-P(X3)-Y-T;

[0176] Among them, X1, X2, and X3 are mutation sites.

[0177] Table 1 Mutation sites related to antibody activity

[0178]

[0179] After the mutation, the antibody activity was detected, ...

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Abstract

The present invention relates to an isolated binding protein comprising cTnI antigen binding domain, and researches on the preparation, application and other aspects of the binding protein. The binding protein has strong activity and high affinity with human cTnI protein, and can be widely used in the detection field of cTnI protein.

Description

technical field [0001] The invention relates to the fields of biotechnology and medical technology, in particular to an anti-human cardiac troponin I antibody and its application. Background technique [0002] Before the 1980s, the World Health Organization (WHO) has always regarded myocardial enzyme activity as one of the diagnostic criteria for acute myocardial infarction (AMI). In the late 1980s, researchers discovered that the sensitivity and specificity of troponin (Tn) was higher than that of phosphocreatine kinase (CK), phosphocreatine kinase isoenzyme (CK-MB), lactate dehydrogenation enzymes and biomarkers such as aspartate aminotransferase. Cardiac troponin (cTnI) exists only in the myocardium and is a marker of cardiomyocytes. Its abnormal changes can affect the systolic function of the heart, and can be used to diagnose myocardial necrosis and judge myocardial injury. It has become a sensitive and specific marker of myocardial cell injury. It is recognized as th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/18C12N15/13C12N15/85G01N33/68
CPCC07K16/18C07K2317/31C07K2317/55C07K2317/565C07K2317/569C07K2317/622C12N15/85G01N33/6893G01N2800/12G01N2800/324G01N2800/325
Inventor 崔鹏何志强孟媛钟冬梅
Owner DONGGUAN PENGZHI BIOTECH CO LTD
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