The hPASCs were cultured in a poly-D-lysine-coated 6-well plate for attachment, then washed with 1X PBS and fixed with 4% paraformaldehyde for 30 min at room temperature. The fixed cells were incubated in 0.3% Triton™ X-100 (Sigma-Aldrich, USA) for 15 min and washed with 1X PBS three times. The cells were blocked with 5% bovine serum albumin (BSA) for 1 h and incubated with primary antibody at 4 °C overnight. The cells were then washed with 1X PBS and incubated with secondary antibody conjugated to Alexa-Fluor 488 or Alexa-Fluor 647 (abcam, USA) at 1:1000 dilution for 1 h at room temperature. The images were captured using a Zeiss microscopic imaging system with fluorescence emission system at different magnifications. For tissue staining, the samples were sectioned at a thickness of 10 μm per slice and subjected to the above procedures.
All antibodies were purchased from Abcam unless otherwise stated. The antibodies used in the experiments were as follows: rabbit anti-Sox2 (1:800), rabbit anti-Oct4 (1:500), rabbit anti-Nestin (1:200), rabbit anti-CD133 (1:500), rabbit anti-ANXA2(1:1000), and mouse anti-FSHβ (1:800, Santa Cruz Biotechnology).
All antibodies were purchased from Abcam unless otherwise stated. The antibodies used in the experiments were as follows: rabbit anti-Sox2 (1:800), rabbit anti-Oct4 (1:500), rabbit anti-Nestin (1:200), rabbit anti-CD133 (1:500), rabbit anti-ANXA2(1:1000), and mouse anti-FSHβ (1:800, Santa Cruz Biotechnology).