In vitro functional activity of IgG1 and IgG3 polyclonal and monoclonal anti-D

IgG anti-D is generally restricted to IgG1 and IgG3; it mediates red cell destruction through interactions with IgG Fc receptors (Fc gamma R) on effector cells. The relative ability of these two IgG subclasses of anti-D to mediate haemolysis in vitro by monocytes and K cells was investigated.

Anti-D was affinity purified from 5 preparations of prophylactic anti-D immunoglobulin, and IgG subclasses quantified by ELISA; mean levels were 86.5% IgG1, 1.4% IgG2, 11.6% IgG3 and 0.4% IgG4. IgG1 and IgG3 polyclonal anti-D were further purified separately from some of the anti-D by removal of either IgG3 using magnetic beads coated with anti-IgG3, or of IgG1 using protein A. These preparations were compared with monoclonal anti-D (BRAD-3 and BRAD-5) for their ability to lyse red cells in antibody-dependent cell-mediated cytotoxicity (ADCC) assays.

Monocyte-mediated lysis of red cells coated with IgG3 anti-D was approximately twice that of cells coated with IgG1 anti-D at similar sensitization levels, and anti-D preparations containing 10% or more IgG3 gave similar lysis. By contrast, in the K cell ADCC, IgG1 anti-D was 2-4 times more haemolytic than IgG3 anti-D. Polyclonal IgG1 and IgG3 anti-D promoted about 20% more lysis than BRAD-5 (IgG1) and BRAD-3 (IgG3), respectively, in the K cell ADCC, although no difference was observed between polyclonal and monoclonal anti-D in the monocyte ADCC.

These experiments demonstrated a functional dichotomy between these two subclasses of anti-D; IgG3-coated red cells were lysed preferentially by monocytes mediated predominantly through Fc gamma R1 interactions, whereas haemolysis of IgG1-sensitized cells was mediated mainly by Fc gamma RIII on K cells.