title: Production of polyclonal antibody to a recombinant non-structural protein Nsp1a of human astrovirus.
abstract: Human astrovirus (HAstV) are important pathogens that cause acute viral diarrhea in infants. Little is known about the mechanisms of astrovirus-induced diarrhea. Previous studies have suggested that an apoptosis inducer may be encoded in the non-structural protein (nsP1a) of astrovirus and contribute to virus-induced diarrhea. To study the biological function of nsP1a and to gain further insight into nsP1a protein-host cell interactions, good quality antibodies must be produced. The nsP1agene of HAstV-1 was cloned into a bacterial expression vector Pgex-6P-1. The recombinant plasmid Pgex-6P-nsP1a was transformed into Escherichia coli BL21 (DE3) and expressed as a fusion protein that contains N-terminal GST tags. The expressed recombinant protein was purified and used as an antigen to produce an nsP1a antiserum in rabbits. ELISA was used to detect the titer of specific antibodies. Specificity activity was detected by Western blot and immunofluorescence analysis. The titer of specific antibodies was up to 1:30,000. Western blotting and immunofluorescence analysis indicated that the polyclonal antibody could recognize specifically the HAstV-1 nsP1a protein.
authors.affiliation: 1st affiliated hospital endocrinology, Liaoning Medical University, Jinzhou, 121000 Liaoning, China.
authors.forename: Chang
authors.initials: C
authors.lastname: Liu
authors.affiliation: Liaoning Medical University, Jinzhou, 121200 Liaoning, China.
authors.forename: Wen-Hui
authors.initials: WH
authors.lastname: Liu
authors.affiliation: Panjin Liaohe Oilfield General Hospital, Panjin, 124010 Liaoning, China.
authors.forename: Li-Li
authors.initials: LL
authors.lastname: Kan
authors.affiliation: Liaoning Medical University, Jinzhou, 121200 Liaoning, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Li
authors.affiliation: Liaoning Medical University, Jinzhou, 121200 Liaoning, China.
authors.forename: Yong-gang
authors.initials: YG
authors.lastname: Li
authors.affiliation: Liaoning Medical University, Jinzhou, 121200 Liaoning, China. Electronic address: zhaowei8072@163.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology*; D015153:Blotting, Western; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D054884:Host-Pathogen Interactions; D006801:Humans; D001257:Mamastrovirus / Q000276:immunology* / Q000502:physiology; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins
keywords: Escherichia coli expression; Human astrovirus; Non-structural protein; Polyclonal antibodies; Recombinant nsP1a protein