The path winds along isolation and analyses of fetal nucleated red blood cells in maternal peripheral blood: Past, present, and future toward non-invasive prenatal diagnosis

Traditional prenatal diagnosis detects fetal disorders through invading uterus to access fetal cells, which may cause maternal complications, fetal injury, or even miscarriage. Safe and convenient non-invasive prenatal testing (NIPT) by analyzing fetal materials (cell-free DNA/RNA, cells, and extracellular vesicles) that circulate in maternal peripheral blood attracts great attention and has been applied in risk evaluation of several fetal disorders. Among those fetal analytes, fetal nucleated red blood cells (fNRBCs) comprise entire fetal genome, possess distinct membrane antigens, and have a lifespan limited in every single gestation. They were once expected to be an ideal biomarker for NIPT and even definitive prenatal diagnosis. However, recent advances of fNRBC-based NIPT are limited and their applications toward clinical practices are still challenging. Herein, we comprehensively overview research on fNRBCs in maternal peripheral blood, trying to dissect current predicament and inspire potential solutions. The source and lineage of fNRBCs, their entrance into maternal peripheral blood, and their physiochemical characteristics are discussed, and various strategies of label-free or immunoaffinitive isolation and subsequential identification of fNRBCs from maternal blood cells are summarized. Although proof-of-concept analyses toward detecting a few fetal disorders are demonstrated, current fNRBCbased NIPT still suffers many challenges when applied to clinical practices. Nevertheless, via thorough investigation and new analytical technologies, it is believed fNRBC-based NIPT will provide a promising platform to supplement the insufficiency of current strategies.