AN INVESTIGATION OF METHODS FOR ENRICHING TROPHOBLAST FROM MATERNAL BLOOD

Trophoblast deportation is known to occur in normal human pregnancy, b ut it is not yet clear whether these cells routinely enter the materna l peripheral circulation and are available as a source of fetal DNA fo r non-invasive prenatal diagnosis of genetic disorders. To resolve thi s issue requires an efficient method of enriching trophoblast from mat ernal blood combined with a means to confirm its identity. Five differ ent techniques were tested on ten retroplacental blood samples to dete rmine the most sensitive and operator-efficient method. Lysis of red c ells alone gave the best recovery of trophoblast but had to be discoun ted, together with Ficoll density gradient centrifugation, due to the very low purity and the excessive time required. Fluorescence-activate d cell sorting (FACS) of pre-enriched trophoblast resulted in the lowe st recovery rate (8 per cent) despite a 3250-fold enrichment and a ver y high purity. Immunomagnetic beads (Dynabeads) coated with anti-CD16 antibody proved to be the best method for the subsequent immunocytoche mical characterization of deported trophoblast. However, IO beads coat ed with anti-CD45 antibody may be more useful for isolating trophoblas t for prenatal diagnosis due to the high purity, enrichment (32-fold), and recovery rate (78 per cent) obtained with this method.