OBJECTİVES: To evaluate the morphological characteristics, immunophenotypic features, osteogenic, adipogenic, and chondrogenic differentiation capacities of in-vitro cultured amniotic cells during the culture process based on mesenchymal stem cell (MSC). MATERİAL/METHOD: This study used waste cells remaining after fetal karyotype determination with second-trimester amniotic cell culture. Cultured cells were morphologically evaluated daily. Selective subculture (SSC) was produced by the scraping method. Immunophenotypic features of primary and subculture amniotic cells were evaluated based on specific surface antigens. Specific growth mediums were used to assess their osteogenic, adipogenic, and chondrogenic differentiation abilities. Differentiation was confirmed using specific staining. RESULTS: Fifteen cases with 46,XY fetal cytogenetic analyses at 16+3-19+5 gestational weeks were included in the study. Amniotic fluid cells were evaluated morphologically in three classes as Amniotic fluid specific cell-type(AF), Fibroblastoid cell-type(FB1, FB2), and Epithelioid cell-type(E). The first cell adhesions occurred in the first 24-48 hours(36%) of cell culture, while the first colonies within three days(82%). SSC was produced in AF-SSC(n=4) and FB2-SSC(n=6) directions by mechanical scraping. Primary amniotic fluid cells and AF/FB-SSCs were positive for MSC immunophenotypic markers CD29,CD73,CD166,CD44,CD49e,CD90, while negative for CD34,CD45, and HLA-DR. Cells were shown to have osteogenic (Alizarin Red-S) and chondrogenic (Alcian Blue) differentiation potentials based on histochemical staining, while adipogenic (Oil Red-O) differentiation was not obtained. CONCLUSION: We believe that preliminary protocols and experiences to obtain MSCs from amniotic fluid-derived cells produced under routine prenatal diagnosis cell culture conditions can enable many promising pre-clinical/clinical studies and be the pioneer of "stem cell therapy applications" in many "incurable" diseases in the clinic.