The mechanism whereby passive Rh (D) immunoglobulins suppress the fetomatemal
alloimmunization is still unclear. New in vitro tests are needed to better
characterize the functional properties of polyclonal anti-Ds. The DAF assay was
developped to monitor the antibody-dependent cell-mediated cytotoxicity
(ADCC) and the phagocytosis of anti-Rh (D)-sensitized RBCs by effector cells.
The principle of this test is based on the oxydization of the 2,7-diaminofluorene
(DAF) by the pseudoperoxidase activity of free hemoglobin. The reaction is proportional
to the hemoglobin concentration. This test was performed to determine
and emphasize the efficacy of different polyclonal anti-D immunoglobulin preparations
to mediate lysis and phagocytosis of sensitized RBCs by human peripheral
mononuclear cells. The functional properties of different human RhD monoclonal
antibodies were also analyzed and compared. The test was found to be
convenient to perform and allowed the avoidance of radioactive labelling of
RBCs for ADCC studies. It is mainly useful for the direct quantitation of phagocytosis.