title: Production and characterization of mouse monoclonal antibodies against lysis protein E of phiX174.
abstract: Bacterial ghosts offer a new avenue for the study of inactivated vaccines. However, for many years the mechanism of genetic inactivation was controversial. To obtain mouse monoclonal antibodies (mAbs) against protein E will allow the observation of its location and dynamic expression to expand understanding of the lysis mechanism. In this study, a His-tagged ΔE fusion protein expressed in bacteria was used as the immunogen, and mAbs targeting protein E were produced by the hybridoma technique and selected by enzyme-linked immunosorbent assay using GST-E and GST-ΔE as coating proteins. Purified mAbs from mouse ascites were screened against a phage-displayed random dodecapeptide library (Ph.D.-12). After three rounds of biopanning, 30 phage clones were randomly selected and sequenced, and their amino acids were deduced. One epitope showed a good match with protein E at 57-63aa (KPLN--R) and the synthetic decapeptide Epep (VRLKPLNCSR) strongly inhibited combination of E-A5 with E fusion proteins. Immunofluorescence microscopy indicated specific immunoreactivity of E-A5 with Escherichia coli-expressed native protein E. The novel mAbs may be of great potential value in analysis of the function and lysis pathway of protein E and other relative phages and in evaluation of E as potential marker of bacterial ghosts.
authors.affiliation: State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Science, Harbin 150001, China. yushenye@163.com
authors.forename: Shenye
authors.initials: S
authors.lastname: Yu
authors.forename: Hailing
authors.initials: H
authors.lastname: Zhao
authors.forename: Huanan
authors.initials: H
authors.lastname: Wang
authors.forename: Xiumei
authors.initials: X
authors.lastname: Wang
authors.forename: Guangxi
authors.initials: G
authors.lastname: Shao
authors.forename: Lili
authors.initials: L
authors.lastname: Xu
authors.forename: Wei
authors.initials: W
authors.lastname: Si
authors.forename: Liping
authors.initials: L
authors.lastname: Chen
authors.forename: Wanjiang
authors.initials: W
authors.lastname: Zhang
authors.forename: Siguo
authors.initials: S
authors.lastname: Liu
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000302:isolation & purification*; D000914:Antibodies, Viral / Q000276:immunology* / Q000302:isolation & purification*; D010584:Bacteriophage phi X 174 / Q000276:immunology*; D018604:Epitope Mapping; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D014764:Viral Proteins / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; C029555:E protein, bacteriophage X174; D014764:Viral Proteins