A protocol to detect fetal nucleated red blood cells (NRBCs) was tested in 217 pregnant women and in 19 nonpregnant controls. All the pregnant women were sampled after chorionic villus sampling (CVS); 20 were also sampled pre-CVS. NRBC recognition was based upon morphology by using staining of hemoglobin with 3,3-diaminobenzidin (DAB) or by immunocytochemical staining for fetal hemoglobin (HbF), This was combined with FISH analysis for both the X- and Y-chromosomes on the same cells. Progressive refinement of the methods increased the number of cases where NRBCs were detected from 53% (DAB) to 75% and 78% for DAB and HbF staining, respectively, with on average 43 NRBCs (range, 1-220), DAB gave a slightly higher yield than HbF in the lower cell count range (
