Goat anti rabbit igg rbitc

After 4 d, spheroid formation or monolayer on slides was washed with PBS and fixed in 4% paraformaldehyde at room temperature for 10 min, after three times washed with PBST, and permeabilised with 0.1% Triton X-100 solution for 15 min at room temperature. Then, washed with PBST, primary antibodies were then incubated with samples overnight at 4°C: anti-Oct4 (1:200, Cell Signaling Technology), anti-Sox 2(1:200, Cell Signaling Technology), anti-Nanog (1:200, Cell Signaling Technology), anti-CK14 (1:200, ZSGB-BIO, Beijing, China), anti-Vimentin (1:200, ZSGB-BIO), anti-DSPP (1:100, Santa Cruz Biotechnology) and anti-DMP-1 (1:100, Abcam). After incubation with primary antibodies, samples were washed with PBST and then incubated with goat anti-rabbit IgG/RBITC or goat anti-mouse IgG/RBITC (1:100, Solarbio Life Science, Beijing, China) for 1 h at room temperature. For nuclear DNA dye, the samples were mounted with DAPI. Images were captured using the Leica TCS SP8 laser scanning confocal microscopy system.