Altered specificity of single-chain antibody fragments bound to pandemic H1N1-2009 influenza virus after conversion of the phage-bound to the soluble form.

摘要: In 2009, a novel influenza A/H1N1 virus (H1N1pdm) quickly spread worldwide and co-circulated with then-existing seasonal H1N1 (sH1N1). Distinguishing between these 2 viruses was necessary to better characterize the epidemiological properties of emergent virus, including transmission patterns, pathogenesis, anti-influenza drug resistance. This situation prompted us develop point-of-care differentiation system before entering 2009–2010 season. Aiming establish H1N1pdm-specific detection tools rapidly, we employed phage display libraries select single-chain variable fragments (scFvs). Human single-fold scFv (Tomlinson I + J) underwent selection for ability bind H1N1pdm particles. Three rounds panning brought 1152 phage-bound scFvs, which 58 clones reacted specifically or preferentially over sH1N1 in an enzyme-linked immunosorbent assay (ELISA). After conversion scFvs soluble form, 7 demonstrating high/stable expression were finally obtained. However, all except No. 29 found have lost their specificity/preference ELISA. The also confirmed by immunofluorescence immunoprecipitation, viral nucleoprotein identified ELISA as its target protein. change specificity associated from form could be due loss scaffold pIII protein, likely provides structural support antigen-binding site. It is possible that similar antigenic led observed alterations specificity. Using library, obtained reactive against H1N1pdm; however, only 1 showed toward H1N1pdm. Our results using highly advantageous rapid development molecules detect antigens. our indicated this strategy might not been effective selecting antibodies during 2009 pandemic, where co-circulating shared properties. suggests it advisable use synthetic library strategically considering characteristics antigens potential situations.