CD16/32-specific biotinylated 2.4G2 single-chain Fv complexed with avidin-FITC enhances FITC-specific humoral immune response in vivo in a CD16-dependent manner

Fcγ receptors (FcγRs) play an essential role in the regulation of immune response due to their ability to bind immune complexes. Activating FcγRs may facilitate antigen presentation and dendritic-cell maturation, while in the late phase of the immune response, the inhibitory FcγRIIb may down-regulate B-cell activation upon cross-linking with activating receptors. In this study, we investigated the in vivo role of FcγRs on the modulation of humoral immune response. In order to get well-defined immune complexes that can bind to both the activating and the inhibitory FcγRs, we designed a mono-biotinylated single-chain fragment variable construct from the rat anti-mouse CD16/32 clone 2.4G2, linked to avidin–FITC, and tested its effect on the FITC–hapten-specific T-independent type 2 (TI-2) and T-dependent (TD) immune response. When injected intravenously in mice, the complex bound to a small portion of B220+, CD11bhigh and CD11chigh cells and was localized in the spleen on marginal zone macrophages 15 min after treatment. When applied as a booster following primary immunization with TI-2 (FITC–dextran) or TD (FITC–keyhole limpet haemocyanin) antigens, the complex elevated the number of hapten-specific IgM/IgG-producing B cells. This effect was diminished in CD16KO mice, suggesting that the activating-type FcγRIII might be a key mediator of this mechanism.