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Preparation method and application of phosphorylated antibody product of new AKT substrate HIP-55

A HIP-55 and phosphorylation technology, which is applied in anti-animal/human immunoglobulin, biological testing, material inspection products, etc., can solve the problem of lack of HIP-55 phosphorylated antibody

Active Publication Date: 2021-03-12
PEKING UNIV THIRD HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There is no related report of HIP-55 phosphorylation antibody

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  • Preparation method and application of phosphorylated antibody product of new AKT substrate HIP-55
  • Preparation method and application of phosphorylated antibody product of new AKT substrate HIP-55
  • Preparation method and application of phosphorylated antibody product of new AKT substrate HIP-55

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Embodiment 1

[0046] Example 1. Phosphorylated antibody to AKT substrate HIP-55 and its preparation method and application

[0047] 1. Material method

[0048] 1. Preparation of HIP-55 phosphorylated antibody

[0049] (1) Immunogen preparation

[0050] A modified polypeptide KERAM(pS)TTS-C was designed and synthesized for the S269 site of HIP-55 protein, wherein the C at the C-terminus is cysteine, dissolved in PBS buffer solution, and coupled to On hemocyanin (KLH), immunization experimental grade Japanese big-eared rabbits.

[0051] A modified polypeptide C-FLQKQL(pT)QPE was designed and synthesized for the T291 site of HIP-55 protein, wherein the C at the N-terminus is cysteine, the polypeptide was dissolved in PBS buffer solution, and the polypeptide was coupled to On hemocyanin (KLH), and immunize experimental grade Japanese big-eared rabbits.

[0052] SEQ ID No. 1: KERAM(pS)TTS.

[0053] SEQ ID No. 2: FLQKQL(pT)QPE.

[0054] (2) Immunization process

[0055] As shown in Table 1...

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Abstract

The invention discloses a preparation method and application of a phosphorylated antibody product of a new AKT substrate HIP-55. The invention provides a method for preparing an HIP-55 protein S269 orT291 site phosphorylated antibody, which comprises the following steps of: immunizing an animal by taking antigen synthetic peptide as an immunogen, collecting antiserum, and purifying to obtain theHIP-55 protein S269 or T291 site phosphorylated antibody, wherein the antigen synthetic peptide is obtained by connecting a cysteine to the N end of a hapten synthetic peptide and then coupling the hapten synthetic peptide with a carrier protein. The amino acid sequence of the hapten synthetic peptide is as shown in SEQ ID No. 1 or SEQ ID No. 2, and the site S269 or T291 corresponding to the HIP-55 protein is modified with a phosphate group. The HIP-55 phosphorylated antibody provided by the invention has good sensitivity and specificity, can be used for detecting interaction between HIP-55 and AKT, and further can be used for auxiliary diagnosis of myocardial infarction.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a preparation method and application of a phosphorylated antibody product of the new AKT substrate HIP-55. Background technique [0002] Since its first discovery, the serine / threonine kinase AKT (also known as protein kinase B or PKB) as a proto-oncogene has become a major focus of medical attention because of its role in the regulation of various physiological or pathological Function (including metabolism, growth, proliferation, survival, transcription, and protein synthesis) plays an important role. AKT plays important physiological or pathological functions mainly by regulating its various substrates. For example, AKT regulates cell cycle progression by directly phosphorylating the AKT substrates p21 and p27; AKT generates anti-apoptotic signals and promotes cell survival by directly phosphorylating the pro-apoptotic protein Bad or transcription factor FoxO; AKT activates the s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/18C07K16/06G01N33/68
CPCC07K16/18C07K16/06G01N33/6893G01N2800/324
Inventor 李子健姜允奇何丹
Owner PEKING UNIV THIRD HOSPITAL