[Preparation and characterization of the polyclonal antibody against pig integrin beta6 subunit LBD as FMDV receptor].

Abstract

AIM:

To induce the expression of FMDV receptor integrin beta6 subunit ligand-binding domain in E.coli and prepare the rabbit polyclonal antibody against it.

METHODS:

The fragment coding beta6 ligand-binding domain was amplified by PCR and doubly digested with BamH Iand Xho I. Then it was cloned into expression vector pGEX-4T-1 to obtain recombinant plasmid pGEX-4T-1-beta6LBD. After pGEX-4T-1-beta6LBD was transformed into E.coli BL21(DE3) and induced by IPTG, the expression of fusion proteins was identified by SDS-PAGE, with inclusion body prepared and fusion protein purified. Then new Zealand rabbits were immunized to prepare polyclonal antibody against beta6LBD. GST-beta6LBD antiserum was obtained and the specificity of polyclonal antibody was detected by Western blot.

RESULTS:

SDS-PAGE demonstrated that the fusion protein GST-beta6LBD was expressed with the expected molecular weight at 42 000. A single clear band of GST-beta6LBD fusion protein appeared in SDS-PAGE gel after purification. The titer of the polyclonal antibody was above 1:12 800 and it is of high specificity.

CONCLUSION:

The successful preparation of rabbit anti pig beta6LBD polyclonal antibody with high affinity and specificity will lay a foundation for further research into the function of integrin beta6 in FMDV infection.

• Authors: Shan-Dian Gao, Hui-Yun Chang, Jun-Zheng DU, Jing-Feng Wang, Jian-Hua Zhou, Jian-Yong Zhao, Qing-Ge Xie
• Journal: Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology (Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi) Vol. 24 Issue 10 Pg. 975-8 (Oct 2008) ISSN: 1007-8738 [Print] China
• PMID: 18845083 (Publication Type: English Abstract, Journal Article)

Chemical References

• Antibodies
• Integrin beta Chains
• Protein Subunits
• Receptors, Virus
• integrin beta6

Topics

• Animals
• Antibodies (immunology, metabolism)
• Blotting, Western
• Electrophoresis, Polyacrylamide Gel
• Foot-and-Mouth Disease Virus (metabolism, pathogenicity)
• Integrin beta Chains (genetics, immunology, metabolism)
• Male
• Polymerase Chain Reaction
• Protein Subunits (genetics, immunology, metabolism)
• Rabbits
• Receptors, Virus (genetics, immunology, metabolism)
• Swine