The utility of an erythroblast scoring system and gender-independent short tandem repeat (STR) analysis for the detection of aneuploid fetal cells in maternal blood.
OBJECTIVE
The aim of this study was to determine whether fetal nucleated red blood cells (NRBCs) could be distinguished from maternal cells in peripheral blood using an erythroblast scoring system based on the unique morphological and hemoglobin staining characteristics of this cell type. Presumptive fetal NRBCs were further analyzed for the presence of paternally inherited DNA polymorphisms to prove fetal origin.


METHODS
NRBCs were isolated by density gradient separation, CD15/45 depletion, and gamma hemoglobin positive selection from peripheral blood of nine women following termination of pregnancy for trisomy 21 (n=4), 18 (n=1), 13 (n=2), and other genetic abnormalities (n=2). Candidate fetal NRBCs, based on four discrete morphological and hemoglobin staining criteria, were then subjected to fluorescent PCR (polymerase chain reaction) amplification of chromosome 21 (D21S1411, D21S11) and chromosome 18 (D18S535) short tandem repeat (STR) DNA polymorphisms.


RESULTS
In all cases, candidate fetal NRBCs were accurately identified on the basis of morphologic and hemoglobin staining characteristics and confirmed to be fetal in origin based on the presence of shared and nonshared polymorphic DNA alleles when compared to DNA isolated from maternal cells.


CONCLUSIONS
Using the erythroblast scoring system and subsequent analysis of inherited DNA polymorphisms, we were able to distinguish fetal NRBCs from maternal cells and prove fetal origin independent of gender. These results suggest that this novel combined approach to fetal cell isolation and genetic analysis is a promising method for noninvasive prenatal diagnostic applications.
Cha
Dong Hyun
DH
Department of Obstetrics and Gynecology, CHA Hospital, Pochon CHA University, College of Medicine, Seoul, South Korea.
Khosrotehrani
Kiarash
K
Bianchi
Diana W
DW
Johnson
Kirby L
KL
D000782:Aneuploidy; D004247:DNA / Q000032:analysis / Q000097:blood; D004900:Erythroblasts / Q000737:chemistry / Q000166:cytology*; D005260:Female; D005333:Fetus / Q000166:cytology*; D005434:Flow Cytometry; D005865:Gestational Age; D006801:Humans; D008297:Male; D008846:Micromanipulation; D018895:Microsatellite Repeats; D016133:Polymerase Chain Reaction; D011237:Predictive Value of Tests; D011247:Pregnancy; D011296:Prenatal Diagnosis; D012732:Sex Determination Analysis / Q000379:methods
D004247:DNA