title: Characterization of first trimester fetal erythroblasts for non-invasive prenatal diagnosis.
abstract: Isolating fetal erythroblasts from first trimester maternal blood offers a promising non-invasive alternative for prenatal diagnosis. The aim of this study was to characterize the biological properties of first trimester primitive erythroblasts to facilitate their enrichment from first trimester maternal blood. Primitive erythroblasts were the predominant cell type until 12 weeks gestation, after which time their numbers declined steeply; 100% were epsilon-globin-positive versus <0.06% definitive erythroblasts. Buoyant densities of first trimester fetal erythroblasts ranged from 1.077 to 1.130 g/ml, and optimal recoveries were obtained with Percoll 1118. Although primitive erythroblasts carried a negative surface charge and were resistant to NH(4)Cl lysis, these properties had only a limited role in fetal cell enrichment. Immunophenotyping showed that primitive, like definitive, erythroblasts were GPA+, CD47+, CD45- and CD35-, whereas CD71 expression was weak/undetectable on primitive erythroblasts but strongly positive on 100% of definitive erythroblasts; primitive erythroblasts were also CD36- whereas definitive erythroblasts were CD36+. We therefore used CD45/GPA selection of Percoll 1118-separated cells to demonstrate successful enrichment of male epsilon-globin-positive fetal erythroblasts from model mixtures, and as proof of principle from some first trimester maternal blood samples. Fetal cell enrichment protocols based on first trimester epsilon-globin-positive primitive erythroblasts may allow reliable enrichment of fetal cells from maternal blood for early non-invasive prenatal diagnosis of genetic disorders.
authors.affiliation: Wolfson and Weston Research Centre for Family Health, Institute of Reproductive and Developmental Biology, Imperial College London, Queen Charlotte's and Chelsea Hospital, Hammersmith Campus, Du Cane Road, London W12 0NN, UK. obgmac@nus.edu.sg
authors.forename: Mahesh
authors.initials: M
authors.lastname: Choolani
authors.forename: Keelin
authors.initials: K
authors.lastname: O'Donoghue
authors.forename: David
authors.initials: D
authors.lastname: Talbert
authors.forename: Sailesh
authors.initials: S
authors.lastname: Kumar
authors.forename: Irene
authors.initials: I
authors.lastname: Roberts
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Letsky
authors.forename: Phillip R
authors.initials: PR
authors.lastname: Bennett
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Fisk
mesh_terms: D000643:Ammonium Chloride / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism; D000944:Antigens, Differentiation, B-Lymphocyte / Q000378:metabolism; D000954:Antigens, Surface / Q000378:metabolism; D004900:Erythroblasts / Q000166:cytology / Q000502:physiology*; D005260:Female; D005865:Gestational Age; D006021:Glycophorins / Q000378:metabolism; D006801:Humans; D018189:Immunomagnetic Separation; D017404:In Situ Hybridization, Fluorescence; D017493:Leukocyte Common Antigens / Q000378:metabolism; D008297:Male; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D011296:Prenatal Diagnosis / Q000379:methods*; D011990:Receptors, Transferrin
chemical_list: D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; D000954:Antigens, Surface; C068126:CD71 antigen; D006021:Glycophorins; D011990:Receptors, Transferrin; D000643:Ammonium Chloride; D017493:Leukocyte Common Antigens