Method for preparing CYP4A11 polypeptide, and antibody for anti human CYP4A11 polypeptide

A CYP4A11, CYP4A11-KLH technology, applied in the direction of anti-animal/human immunoglobulins, peptides, etc., can solve the problems of low safety, high cost of antibodies, and expensive CYP4A11

Inactive Publication Date: 2007-02-21
HARBIN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of the present invention is to provide a method for preparing a CYP4A11 polypeptide and an anti-human CYP4A11 polypeptide antibody in order to solve the problems that currently used human CYP4A11 is expensive, the cost of the prepared antibody is too high, and the safety is low

Method used

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  • Method for preparing CYP4A11 polypeptide, and antibody for anti human CYP4A11 polypeptide
  • Method for preparing CYP4A11 polypeptide, and antibody for anti human CYP4A11 polypeptide
  • Method for preparing CYP4A11 polypeptide, and antibody for anti human CYP4A11 polypeptide

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specific Embodiment approach 1

[0005] Specific embodiment 1: The amino acid sequence of the CYP4A11 polypeptide in this embodiment is:

[0006] ArgLeuArgArgLeuProAsnProCysGluAspLysAspGlnLeu.

specific Embodiment approach 2

[0007] Specific embodiment two: In this embodiment, the anti-human CYP4A11 polypeptide antibody is prepared according to the following steps:

[0008] (1) Analysis of CYP4A11 antigenic epitope: use DNAStar software analysis to determine the 15 amino acids at positions 505-519 of CYP4A11 protein as the amino acid sequence of the synthetic polypeptide; (2) Synthesis of CYP4A11 polypeptide: Synthesize and purify with an automatic polypeptide synthesizer; (3) ) CYP4A11 polypeptide and carrier protein cross-linking: CYP4A11 polypeptide and carrier protein KLH are linked by glutaraldehyde, and then dialyzed with boric acid buffer solution with a pH value of 8.5 for 8-12 hours to form CYP4A11-KLH by cross-linking; (4) Preparation of rabbit antibody CYP4A11 polypeptide antibody: inject the emulsified CYP4A11-KLH subcutaneously on the back of the rabbit, and repeatedly boost the immunization until the antibody titer is equal to or higher than 1:10000; (5) Collect and separate the serum ...

specific Embodiment approach 3

[0010] Specific embodiment three: the difference between this embodiment and specific embodiment two is: in step (two), a conventional C18 RP-HPLC column is used for purification. Other steps are the same as the second embodiment.

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Abstract

This invention relates to a peptide CYP4A11 and method for preparing antibody against human CYP4A11. The peptide CYP4A11 has an amino acid sequence of RLRRLPNPCEDKDQL. The method comprises: (1) analyzing the epitopes of peptide CYP4A11; (2) synthesizing peptide CYP4A11; (3) crosslinking the peptide with vector protein; (4) preparing antibody against the peptide in rabbit; (5) collecting antiserum, and purifying the antibody. The obtained antibody has high titer and low cost, and is capable of coupling with natural human CYP4A11 molecules, thus can satisfy the requirements of immunohistochemical assay, immunoblotting and ELISA experiments.

Description

technical field [0001] The invention relates to a polypeptide and a preparation method of the polypeptide antibody. Background technique [0002] CYP4A11 is one of the subtypes of cytochrome P450 4A, which is mainly distributed in the liver and kidney. It can catalyze the metabolism of various fatty acids (lauric acid, etc.) and regulate the balance of lipid metabolism in the body. Studies have found that CYP4A11 catalyzes the metabolism of endogenous arachidonic acid to generate 20-hydroxyarachidonic acid (20-HETE), and 20-HETE is related to the formation of essential hypertension. In addition, other studies have shown that CYP4A11 is also involved in the mechanism of defense against ultraviolet oxidative damage. Due to the current use of genetically recombinant human CYP4A11 (#RDI-CYP4A11abr: http: / / www.researchd.com / cyp450 / cyp4alab.htm provides rabbit anti-human CYP4A11 antibody, using purified human liver CYP4A11 protein as antigen for western blot experiments .) The p...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/00C07K7/08C07K16/18
Inventor 朱大岭唐晓波杨微刘海英武正华吴红
Owner HARBIN MEDICAL UNIVERSITY
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