Optimization of nucleated red blood cell (NRBC) recovery from maternal blood collected using both layers of a double density gradient

The isolation of fetal nucleated red blood cells (NRBC) from maternal blood represents a promising approach to non-invasive prenatal diagnosis. Howeve r, the number of fetal NRBC in maternal circulation is quite low and theref ore difficult to isolate. An enrichment procedure in which both layers from a double density 1.077/1.107 g/ml gradient ale collected was optimized, fo llowed by MACS selection using noncommercial monoclonal antibodies. The inf luence of the delay in processing maternal blood on the NRBC distribution i n both interfaces of the gradient was also studied in cord blood and periph eral maternal blood samples. A significant increase in the number of NRBC i solated from maternal blood was achieved by collecting both layers of the d ouble density gradient compared with the previous protocol in which only th e lower layer was recovered. Cord blood samples showed significant differen ces in the number of NRBC recovered when processed at 23 instead of within 3 h. This effect was also observed in the number of NRBC collected only fro m the upper layer of peripheral maternal blood samples. Therefore, in order to minimize the target cell losses, it is advisable to process the materna l blood samples as soon as possible. Copyright (C) 2001 John Wiley & Sons, Ltd.