1x71s1f fluorescence microscope

To fully characterize iPSCs, these cells were fixed with methanol for 20 min at −20 °C, and nonspecific receptors were blocked with 10% normal goat serum. Cells in culture plates or chamber slides were fixed for 20 min at −20 °C with methanol, and nonspecific receptors were blocked with 10% normal goat serum. Oct4, SSEA1, and Nanog were stained with specific antibodies (anti-Oct4, 1:500, Santa Cruz Biotechnology SC-5279; anti-Nanog, 1:500, Santa Cruz Biotechnology SC-293121; and anti-Dppa4, 1:500, R&D AF3674; anti-SSEA1, 1:1000, Invitrogen MA1-022), followed by goat anti-mouse antibody-conjugated Texas Red (1:2000, Invitrogen T-862). Nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI; Invitrogen). Immunofluorescence staining was visualized, and cells were photographed with an Olympus 1X71S1F fluorescence microscope.

Cells were cultured on coverslips, fixed with 4% (wt/vol) paraformaldehyde–PBS for 15 min, and then permeabilized with 0.2% (vol/vol) Triton X-100. Primary antibodies (anti-FOSL1; 5281) were incubated at 4°C overnight after blocking with 3% bovine serum albumin (BSA)–PBS was performed. After three washes with PBS, the cells were then incubated with fluorochrome-conjugated secondary antibodies at RT for 1 h. Following three washes, the cells were stained, sealed with Vectashield antifade mounting medium with DAPI (4′,6-diamidino-2-phenylindole; Vector Laboratories, Burlingame, CA), and imaged using an Olympus 1X71S1F fluorescence microscope.