Long-term culture and characterization of goat primordial germ cells

While the culture and identification of primordial germ cells (PGCs) in mic e is established, only limited investigations on PGCs in livestock have bee n reported. This study was performed to characterize goat PGCs after cultur e and cryopreservation. Goat PGCs were isolated from Day 32 fetuses and cul tured on a continuous cell line of murine embryonal fibroblasts (STO) as fe eder-cells in the presence of leukemia inhibitory factor (LIF). The PGCs pr oliferated slowly and showed colony formation in early passages. Frozen-tha wed PGCs continued to proliferate when stem cell factor (SCF) was added to the culture medium. However, differentiation into epitheliallike polygonal cells or neuronal cells was observed after 1 or 2 passages. The PGCs of 1 f emale and 1 male cell line were characterized by immunocytochemistry. The P GCs showed positive staining for anti stage-specific embryonic antigen-1 (S SEA-1) and FMA-1 (monoclonal antibody produced against a glycoprotein cell surface antigen of the embryonal carcinoma Nulli SCC1), whereas the reactiv ity to alkaline phosphatase (AP), an established marker for PGCs in mice, w as inconsistent. After differentiation, PGCs lost their positive reaction t o SSEA-1, EMA-1 and AP. In conclusion, SSEA-1 and EMA-1 can be used as reli able markers for identifying goat PGCs in addition to morphological criteri a. The results indicate that goat PGCs can be kept in long-term culture wit hout losing their morphological characteristics and their positive reaction to SSEA-1 and EMA-1, thus providing a promising source of donor-karyoplast s for nuclear transfer procedures. (C) 2000 by Elsevier Science Inc.