In vitro differentiation of human oocyte-like cells from oogonial stem cells: single-cell isolation and molecular characterization

人类卵母细胞样细胞从卵原干细胞体外分化:单细胞分离和分子表征

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作者:Erica Silvestris, Paola Cafforio, Stella D'Oronzo, Claudia Felici, Franco Silvestris, Giuseppe Loverro

Methods

Ovarian cortex fragments were processed by immuno-magnetic separation using a rabbit anti-human DDX4 antibody and the positive populations were measured by assessing both FRAGILIS and stage-specific embryonic antigen 4 (SSEA-4) expression. After 3 weeks in culture, large oocyte-like cells were individually isolated by DEPArray based on PKH26 red staining and cell size determination. GDF-9 and SYCP3 as final, and developmental pluripotency-associated protein 3 (DPPA3) as primordial, germline markers were measured by droplet digital PCR. The haploid versus diploid chromosomal content of chromosomes X and 5 was investigated using fluorescence in situ hybridization (FISH). Main

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