In a previous study, we demonstrated that apoptosis increased according to 
gestational age, accounting partly for the presence of free fetal DNA in 
maternal plasma and serum. Using simultaneous TUNEL assay and FISH analysis, we 
identified the fetal origin of part of the apoptotic cell population, but very 
few TUNEL-positive cells showed hybridization signals since they were in a late 
apoptosis stage and nuclei were destroyed. In the present study, the apoptotic 
cell population was identified immunocytochemically using Annexin V, a marker of 
cells in an early stage of apoptosis. The mean apoptosis rate in mononuclear 
cells isolated from the peripheral blood of 20 pregnant women in the 16th to 
19th week of pregnancy with Annexin V was 6.8 +/- 0.5% (range: 4.2-8.1%) 
compared to 6.14 +/- 0.5% (range: 3.7-6.9%) obtained with ethidium bromide 
staining. FISH using X and Y chromosome-specific probes was applied in 11 cases 
known to be carrying male fetuses. Eighty percent of Annexin V+ cells showed 
hybridization signals, while the proportion of apoptotic cells showing X/Y 
signals was 7.8% (range: 5-12%). Although our results are still preliminary, it 
seems that use of Annexin V antibody to detect the apoptotic cell population 
improves FISH analysis and allows a more accurate estimate of the proportion of 
fetal cells among the apoptotic cell population.