title: High throughput screening of scFv antibodies against viral hemorrhagic septicaemia virus by flow cytometry.
abstract: Viral hemorrhagic septicaemia (VHS) is an economically important disease that affects salmon and trout worldwide. In this study, a recombinant single chain variable fragment (scFv) antibody library derived from rainbow trout immunized with purified viral hemorrhagic septicaemia virus (VHSV) was constructed. The library was subjected to three rounds of screening by flow cytometry (FCM) against VHSV through a bacteria display technology, resulting in the enrichment of scFv. Four scFv clones with different fluorescence intensity were obtained by colony pick up at random following three rounds of screening. The isolated scFv antibodies were expressed and purified. Relative affinity assay showed the four clones had different sensitivity to VHSV, in accordance with FCM. The potential use of the selected VHSV-specific scFv antibodies was demonstrated by the successful application in Western blotting assay, ELISA and immunofluorescence antibody test (IFAT), and one of the isolated scFv molecular showed excellent in vitro and in vivo blocking activities against VHSV. scFv isolated in this study can be promising diagnostic and/or therapeutic reagents for VHS. This study provides powerful strategies for screening antibodies against new diseases.
authors: College of Life Sciences and Agro-forest, Qiqihar University, Qiqihar 161000, PR China. Electronic address: zhouyao830226@163.com.
authors: Yao
authors: Y
authors: Zhou
authors: College of Life Sciences and Agro-forest, Qiqihar University, Qiqihar 161000, PR China. Electronic address: 2731848980@qq.com.
authors: Zhi-Gang
authors: ZG
authors: Xie
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D000915:Antibody Affinity / Q000276:immunology; D005393:Fish Diseases / Q000276:immunology* / Q000821:virology*; D005434:Flow Cytometry; D015723:Gene Library; D057166:High-Throughput Screening Assays; D009500:Neutralization Tests; D029141:Novirhabdovirus / Q000276:immunology*; D018353:Rhabdoviridae Infections / Q000662:veterinary*; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification
chemical_list: D000914:Antibodies, Viral; D057127:Single-Chain Antibodies
keywords: Antibody library; Bacteria display; Fluorescence-activated cell sorting; Immunofluorescence assay; Single chain variable fragment; Viral hemorrhagic septicaemia virus