title: Generating and characterizing monoclonal and polyclonal antibodies against avian H5N1 hemagglutinin protein.
abstract: Accurate and timely diagnoses are central to H5N1 infection control. Here we describe the cloning and expression of the HA1 protein of the A/Vietnam/1203/04 strain in a bacterial system to generate mono-/polyclonal antibodies. All of the eight generated monoclonal antibodies recognized the same linear epitope on the top globular region of the HA structure -- a highly conserved epitope among all circulating H5N1 clades identified by amino acid alignment. Results from immunofluorescence staining and Western blotting indicate that all monoclonal antibodies interacted with a denatured form of HA proteins, while the resultant polyclonal antibodies recognized both denatured and native HA proteins on H5N1 reverse-genetics (RG) viruses. Results from flow cytometry and microneutralization assays indicate that the polyclonal antibodies blocked viral binding and neutralized H5N1-RG viruses. Our results may prove useful to establishing future H5N1 mono-and polyclonal antibodies, and perhaps contribute to the development of an alternative H5N1 vaccine.
authors.affiliation: Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan.
authors.forename: Sheng-Fan
authors.initials: SF
authors.lastname: Wang
authors.forename: Kuan-Hsuan
authors.initials: KH
authors.lastname: Chen
authors.forename: Arunee
authors.initials: A
authors.lastname: Thitithanyanont
authors.forename: Ling
authors.initials: L
authors.lastname: Yao
authors.forename: Yuan-Ming
authors.initials: YM
authors.lastname: Lee
authors.forename: Yu-Jiun
authors.initials: YJ
authors.lastname: Chan
authors.forename: Shih-Jen
authors.initials: SJ
authors.lastname: Liu
authors.forename: Pele
authors.initials: P
authors.lastname: Chong
authors.forename: Wu-Tse
authors.initials: WT
authors.lastname: Liu
authors.forename: Jason C
authors.initials: JC
authors.lastname: Huang
authors.forename: Yi-Ming Arthur
authors.initials: YM
authors.lastname: Chen