Authors: I. Hromadníková 1; Lenka Sedláčková 2; M. Mrštinová 3; K. Štechová 1; KaramanovS. 5; J. Kofer 5; M. Macek 4 Authors‘ workplace: II. dětská klinika 2. LF UK, FN Motol, Praha, přednosta doc. MUDr. J. Vavřinec, CSc. 2 Laboratoř imunologie, KlinLab, Praha 3 Gynek. -porod. klinika FN Motol, Praha, přednosta doc. MUDr. K. Citterbart, CSc. 4 Oddělení lékařské genetiky, FN Motol, Praha, p 1 Published in: Ceska Gynekol 2000; (1): 33-37 Category:
Objective: Enrichment of nucleated red blood cells (NRBCs) from maternal blood for non-invasiveprenatal diagnosis.Design: Pilot study.Setting: 2 nd Clinic of Paediatrics, University Hospital Motol, Prague, Czech Republic.Methods: Mononuclear cells were isolated from 13 - 28 ml of peripheral maternal blood between 13and 37 weeks of gestation. Leukocytes from maternal peripheral blood were depleted from mono-nuclear cells by treatment with anti-CD14 and anti-CD45 microbeads and high-gradient magneticcell separation (MACS) on VarioMACS. NRBCs were sorted from CD14 - /CD45 - fraction by positiveselection using anti-CD71 microbeads on MiniMACS. All sorting steps were analysed by three-co-lour cytometric analysis with FACScan flow cytometer.Results: In 68 out of 78 pregnant woman (87 %) NRBCs were found in range 2x10 5 - 1,02 x 10 6.NRBC were enriched with an average enrichment rate of 138-fold ranging from 4 - 526 fold. In ourcohort of pregnant woman the number of isolated NRBCs was individual. We identified NRBCsfrom the 13 th week of gestation.Conclusion: The aim of the study is to establish and standardise the method of enrichment ofNRBCs from maternal blood samples and verify the applicability of this alternative source fornon-invasive prenatal diagnosis.
Key words: nucleated red blood cells, high-gradient magnetic cell separation (MACS), flow cyto-metry (FACS), prenatal diagnosis, fetal cells.
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