The human SSC line was established by overexpressing human SV40 large T antigen in human primary GPR125-positive undifferentiated spermatogonia in our lab (Hou et al., 2015 (link)), the cell line was positive for a series of SSC markers including GPR125, GFRA1, PLZF, UCHL1, and THY1, and it can be expanded in vitro for a long time.
The human SSC line cells were cultured in DMEM/F12 (Gibco, Grand Island, NY, United States) with 10% FBS (Gibco) and 100 unit/mL streptomycin/penicillin (Invitrogen, CA, United States) at 34°C in 5% CO2 incubator. The cells were passaged every 4 days using 0.05% trypsin and 0.53 mM EDTA (Invitrogen). In order to seek which growth factor mediates TCF3, the cells were incubated in the DMEM/F12 with the addition of 10 ng/mL GDNF (R&D Systems, MN, United States), 10 ng/mL FGF2 (R&D Systems), 10 ng/mL epidermal growth factor (EGF) (Sigma, MO, United States), or 1,000 IU/mL LIF (Cyagen, Suzhou, China).
The human SSC line cells were cultured in DMEM/F12 (Gibco, Grand Island, NY, United States) with 10% FBS (Gibco) and 100 unit/mL streptomycin/penicillin (Invitrogen, CA, United States) at 34°C in 5% CO2 incubator. The cells were passaged every 4 days using 0.05% trypsin and 0.53 mM EDTA (Invitrogen). In order to seek which growth factor mediates TCF3, the cells were incubated in the DMEM/F12 with the addition of 10 ng/mL GDNF (R&D Systems, MN, United States), 10 ng/mL FGF2 (R&D Systems), 10 ng/mL epidermal growth factor (EGF) (Sigma, MO, United States), or 1,000 IU/mL LIF (Cyagen, Suzhou, China).
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