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Monoclonal antibody for anti pig growth hormone, preparation process and application

A monoclonal antibody, growth hormone technology, applied in biochemical equipment and methods, botanical equipment and methods, anti-hormone immunoglobulin and other directions, can solve the problems of accumulation residues, human health hazards, etc. Market application prospects, good social and economic benefits

Inactive Publication Date: 2005-10-26
JILIN AGRICULTURAL UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The invention provides an anti-porcine growth hormone monoclonal antibody to solve the problem that porcine growth hormone drugs are used in production to accumulate residues in animals and cause serious harm to human health

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 Preparation of anti-porcine growth hormone monoclonal antibody

[0028] 1. Instruments and equipment

[0029] CO 2 Incubator, Japan SANYO product; Inverted microscope, Chongqing photon instrument factory product; Ultra-clean bench, Suzhou Purification Equipment Factory, W-CT-1F; Microplate reader, American Bio-Rad550 type; Ultrapure water meter, millipore product .

[0030] 2. Animals and Cells

[0031] Experimental animals Balb / c mice were purchased from Changchun Biological Experimental Animal Center, and SP2 / 0 cells were purchased.

[0032] 3. Main medicines and culture medium

[0033] 3.1 Antigen

[0034] Porcine Growth hormone (Porcine Growth hormone) is a product of Sigma Company. PEG4000 is a product of Promega; HT, HAT and enzyme-labeled secondary antibodies are all products of Sigma; RPMI-1640, Freund's complete adjuvant and Freund's incomplete adjuvant are products of Gibco.

[0035] 3.2 Basal medium:

[0036] Prepare according to the instruc...

Embodiment 2

[0050] Example 2 Cloning and DNA sequencing of anti-porcine growth hormone monoclonal antibody gene

[0051] 1. Cell strains, strains and vectors The porcine growth hormone-resistant hybridoma cell line was obtained from Example 1; Escherichia coli DH5α was purchased and stored; pMD18-T vector was purchased from TaKaRa Company.

[0052] 2. Reagents Restriction enzymes Sfi I, Not I and DL2000 DNA Markers were purchased from TaKaRa Company; TaqDNA polymerase, dNTP, RNAgent Tptal RNA Isolation System, PoluATract Mrna Isolation SystemIV, Wizard PCR Preps DNA Purification System were purchased from Promega Company ; Mouse ScFvMoudle / Recombinant Phage Antibody System, purchased from Pharmacia.

[0053] 3. Extraction of total RNA from hybridoma cells According to the methods provided by RNAgent Total RNA Isolation System and PolyATractmRNA Isolation System IV, total cellular RNA was extracted and mRNA was isolated and purified.

[0054] 4. Synthesis of the first strand of cDNA Accor...

Embodiment 3

[0060] Example 3 Subclass identification of anti-porcine growth hormone monoclonal antibody immunoglobulin Ig

[0061] The type and subclass of the monoclonal antibody obtained in Example 1 were identified. Immuno PureMonoclonal antibody istyping kit from Pierce was used to detect the obtained monoclonal antibody hybridoma cell culture supernatant, and it was found that the detected antibody belonged to the IgG1 subclass, and the light chain was of the κ chain type.

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Abstract

The present invention relates to a monoclonal antibody for resisting swine growth hormone, its preparation method and application. It includes heavy chain variable region and light chain variable region, every variable region has respective amino acid sequence. Said invention utilizes the screening process to obtain the hybridoma strain with stable secretion swine growth hormone monoclonal antibody, and can prepare monoclonal antibody with secretion specificity for resisting swine growth hormone, so that it can lay the foundation for further creating quick, flexible and specific swine growth hormone detection method and developing new-type growth promoter.

Description

technical field [0001] The present invention relates to the field of biotechnology. The invention utilizes technologies such as animal immunization, cell fusion and cloning to construct a hybridoma strain stably secreting a mouse anti-pig growth hormone monoclonal antibody, and prepare a specific monoclonal antibody secreting the porcine growth hormone. The present invention also relates to the preparation method of the monoclonal antibody and the application of the monoclonal antibody. Background technique [0002] Growth hormone (GH) is an important hormone that promotes animal growth and regulates metabolism. It can promote glucose absorption, nucleic acid and protein synthesis, and fat breakdown. Porcine growth hormone is a single-chain polypeptide hormone composed of 190 amino acids secreted by the anterior lobe of the pig pituitary gland, with a molecular weight of 22kD. Hormone drugs were once widely used in modern animal production. However, its use has been banne...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/26
Inventor 郑鑫朱世成李月红王哲
Owner JILIN AGRICULTURAL UNIV
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