Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay

VP1 gene is an important gene responsible for the expression of a protein that may facilitate the diagnosis of chicken anaemia virus (CAV) infection.The VP1 gene was cloned and expressed in prokaryotic expression vector pRSET–B.The VP1 protein expression in E. coli was achieved as a fusion protein...

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Main Author:	Mahgoub, Elhem Omer
Format:	Thesis
Language:	English English
Published:	2004
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Online Access:	http://psasir.upm.edu.my/id/eprint/189/1/570123_t_fpv_2005_14.pdf
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spelling	my-upm-ir.1892013-05-27T06:46:22Z Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay 2004-11 Mahgoub, Elhem Omer VP1 gene is an important gene responsible for the expression of a protein that may facilitate the diagnosis of chicken anaemia virus (CAV) infection.The VP1 gene was cloned and expressed in prokaryotic expression vector pRSET–B.The VP1 protein expression in E. coli was achieved as a fusion protein containing an immunogenic epitope. The protein expressed was detected by anti-VP1 monoclonal antibody with an apparent molecular weight of 50 kDa. Batch fermentation was used to scale up production of the protein in E. coli.The recombinant VP1 protein was successfully expressed during high cell density culture. The use of tangential flow filtration (TFF) filtration step for dialysis and desalting increased both the specific activity and the final yield of the purified fraction.The protein expressed has been tested as an antigen for detection of antibody to CAV in infected chicken.An enzyme-linked immunosorbent assay (ELISA) applied for the detection of serum antibody to CAV.This test depends on the availability of CAV polyclonal antibodies present in convalescent chicken serum to react with the VP1 antigen adsorbed to the ELISA plate.When serum samples from infected chicken were tested,the sensitivity of the assay was found to be greater than 93.3%.Furthermore, the ELISA test requires serum samples to be diluted at 1:100 compared to 1:50 for commercial ELISA,an increase in the potential of non-specific background that could be contributed by proteins available in serum samples.In conclusion the indirect ELISA approach using VP1 fusion protein has many advantages. The results of the present study can be used as a basis for the development of a reliable ELISA assay for detection of CAV 2004-11 Thesis http://psasir.upm.edu.my/id/eprint/189/ http://psasir.upm.edu.my/id/eprint/189/1/570123_t_fpv_2005_14.pdf application/pdf en public masters Universiti Putra Malaysia Faculty of Veterinary Medicine English
institution	Universiti Putra Malaysia
collection	PSAS Institutional Repository
language	English English
topic
spellingShingle	Mahgoub, Elhem Omer Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
description	VP1 gene is an important gene responsible for the expression of a protein that may facilitate the diagnosis of chicken anaemia virus (CAV) infection.The VP1 gene was cloned and expressed in prokaryotic expression vector pRSET–B.The VP1 protein expression in E. coli was achieved as a fusion protein containing an immunogenic epitope. The protein expressed was detected by anti-VP1 monoclonal antibody with an apparent molecular weight of 50 kDa. Batch fermentation was used to scale up production of the protein in E. coli.The recombinant VP1 protein was successfully expressed during high cell density culture. The use of tangential flow filtration (TFF) filtration step for dialysis and desalting increased both the specific activity and the final yield of the purified fraction.The protein expressed has been tested as an antigen for detection of antibody to CAV in infected chicken.An enzyme-linked immunosorbent assay (ELISA) applied for the detection of serum antibody to CAV.This test depends on the availability of CAV polyclonal antibodies present in convalescent chicken serum to react with the VP1 antigen adsorbed to the ELISA plate.When serum samples from infected chicken were tested,the sensitivity of the assay was found to be greater than 93.3%.Furthermore, the ELISA test requires serum samples to be diluted at 1:100 compared to 1:50 for commercial ELISA,an increase in the potential of non-specific background that could be contributed by proteins available in serum samples.In conclusion the indirect ELISA approach using VP1 fusion protein has many advantages. The results of the present study can be used as a basis for the development of a reliable ELISA assay for detection of CAV
format	Thesis
qualification_level	Master's degree
author	Mahgoub, Elhem Omer
author_facet	Mahgoub, Elhem Omer
author_sort	Mahgoub, Elhem Omer
title	Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
title_short	Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
title_full	Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
title_fullStr	Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
title_full_unstemmed	Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay
title_sort	production of a recombinant vp1 protein of the chicken anaemiavirus for the development of an enzyme-linked immunosorbent assay
granting_institution	Universiti Putra Malaysia
granting_department	Faculty of Veterinary Medicine
publishDate	2004
url	http://psasir.upm.edu.my/id/eprint/189/1/570123_t_fpv_2005_14.pdf
_version_	1747810169804292096

Production of a Recombinant Vp1 Protein of the Chicken Anaemiavirus for the Development of an Enzyme-Linked Immunosorbent Assay

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