DIFFERENTIAL DEVELOPMENT OF FETAL AND ADULT HEMOGLOBIN PROFILES IN COLONY CULTURE - ISOLATION OF FETAL NUCLEATED RED-CELLS BY 2-COLOR FLUORESCENCE LABELING

Fetal cells in maternal peripheral blood are a source of fetal DNA for prenatal genetic diagnosis, but their numbers are so small and variab le that a reliable isolation procedure has pet to be demonstrated, The problem of scarcity map be overcome by amplification of fetal progeni tor cells in cultures from maternal blood samples. One challenge is to identify post-culture fetal cells and colonies, We have found that th e progeny of fetal and adult erythroid progenitors developed different ial haemoglobin profiles in co-culture, Fetus-derived cells initially made only fetal haemoglobin (HbF) and began to express adult haemoglob in (HbA) only alter intracellular HbF had reached maximum levels, whic h occurred after c 7 d in culture. By this time the large majority of adult-derived erythroid cells contained already high levels of HbA alo ne or combined with HbF Using the HbF(+)HbA(-) criterion, we were able to flow-sort fetal cells with up to nearly 50% purity from some post- termination brood cultures, and with >90% purity in cultures from mate rnal blood spiked with 1% blood from the fetus. Fetal cell purity depe nded on culture time and serum supplement. After 7-10 d, purity was hi gher in low concentrations of human cord serum (1-3%) than in the stan dard 30% fetal calf serum. This was reversed at later times. Thus, if fetal clonogenic erythroid cells were present in maternal blood, their progeny could be isolated from most adult erythoid cells based on hae moglobin profiles. Cultures using CD34(+) cells could be performed com plementary to other methods targeting more mature fetal cells in the s ame maternal blood samples, thus increasing the overall chances of fin ding fetal cells and potentially providing clonal isolation of such ce lls.