[Preparation and characterization of monoclonal antibodies against GII.4 norovirus P domain]

Objective To prepare monoclonal antibodies (mAbs) against GII.4 norovirus P domain by multiple antigens in an immunization program. Methods BALB/c mice were immunized with the multiple GII.4 NoV P domain, namely 1996cluster (VA387), 2004cluster, 2006b cluster and 2010 cluster. The spleen cells from the immunized mice were fused with SP2/0 cells and the hybridoma cells were screened by ELISA. The supernatant of the mAbs was collected and purified by the limiting dilution assay. Its subtype was identified, and the specificity and neutralization were analyzed by indirect ELISA and HBGA blocking, respectively. Results We obtained thirteen hybridoma cell lines that stably secreted mAbs against GII.4 NoV P domain. Their titers reached above 10^-4 after purification. The subtypes of the mAbs were identified as IgG1. Indirect ELISA showed that all the mAbs specifically bound to all GII.4 norovirus variants. Five mAbs specifically bound to GII.17, GII.3 and GII.6 variants. Three mAbs specifically bound to GII.2 variants and strongly blocked NoV P particle from binding to the histo-blood group antigen (HBGA) receptors. Conclusion The mAbs against GII.4 norovirus P domain have been obtained by combined antigens immunization program. Multi-antigen immunization can enhance immune response significantly and cross-react with other GII.4 norovirus variants. The findings provide a basis for further development of novel GII.4 norovirus vaccines and for the optimization of the immunization programs of combined multi-antigen vaccine candidates.