Phenotypic Characterization Of Chinese Rhesus Macaque Plasmablasts For Cloning Antigen-specific Monoclonal Antibodies

Rhesus macaques(macaca mulatta)are used as a more human relevant animal species for evaluation of vaccines and as a source for cloning monoclonal antibodies that are highly identical to human derived antibodies.Although plasmablasts are well defined and can be easily isolated in humans,it remained unclear whether the same phenotypic markers could be applied for isolating plasmablasts in Chinese rhesus macaques.In this study,we evaluated a series of cell surface and intracellular markers and identified the phenotype of plasmablast in Chinese rhesus macaques as CD3-CD 14-CD56-CD19-CD27-CD20-/IowCD80+HLA-DR+CD95+.After vaccination with an influenza virus vaccine,the plasmablasts in peripheral blood mononuclear cell(PBMC)increased transiently,peaked at day 4-7 after a booster vaccination and returned to near undetectable by day 14.Antigen-specific Elispot assays confirmed the secretion of influenza specific antibodies from these plasmablasts.These plasmablasts have transcriptional characteristics similar to human plasmablasts.Using single cell PCR with immunoglobulin heavy chain and light chain primers,influenza specific monoclonal antibodies could be cloned from CD3-CD14-CD56-CD19-CD27-CD20-/lowCD80+HLA-DR+CD95+ plasmablasts at day 4-7 after vaccination.Our work defined the phenotypic markers for isolating antibody secreting plasmablasts and has great implication for evaluation of vaccination-induced plasmablast response and for rapid and efficient cloning of monoclonal antibodies using Chinese rhesus macaques.