Anti hoct4

Cells on glass coverslips were fixed in 4% paraformaldehyde for 15 min. The cells were then blocked and permeabilized in PBS containing 0.1-0.2% Triton X-100 and 10% horse serum. Coverslips were then incubated with the primary antibody in the blocking solution overnight at 4 °C. The coverslips were washed in Tris-buffered saline and incubated with the secondary antibodies for 30 min at room temperature and counterstained with DAPI. The coverslips were then washed and mounted on slides using Fluoromount-G (Southern Biotech), and dried overnight in the dark. The antibodies used for pluripotency and neuronal characterization were anti-hOct4 (Cell Signaling, Danvers, MA, USA 2840 S), anti-hNanog antibody (Cell Signaling 4903 S), anti-MAP2 (Abcam 5392) anti-PAX6 (Abcam ab109233), anti-Nestin (Abcam ab105389), anti-NeuN (Abcam ab177487), and anti-PROX1(Millipore MAB5654).