Figure 2.

Oct-4 protein expression in in-vivo-derived rabbit embryos. Embryos were fixed for double staining of DNA with DAPI (blue) and Oct-4 with antibody (green) and the intensity of Oct-4 signals were quantitated. (A–A4) Day-3 compact morulae had an intense Oct-4 signal in inside cells in contrast to the mild intensity in outside cells (n = 11). (B–B4) Strong Oct-4 fluorescence was still present in the ICM of day-4 early blastocysts (n = 17). (C–C4) In day-4 expanded blastocysts, obvious Oct-4 diminishment was observed in the ICM cells rather than in the TE cells (n = 17). (D–D4) In day-5 blastocysts (n = 13), a wave of Oct-4 elevation was observed manifestly in some specified cells of ICM region, while weak Oct-4 expression was still detectable in TE. (E–F4) Clear Oct-4 expression was observed in the epiblast of day-6 blastocysts (E–E4, n = 11) and day-7 blastocysts (F–F4, n = 16). (G) Comparison of the Oct-4 signal intensity of different regions in embryos collected at different stages. The Oct-4 images of in-vivo-derived embryos were used for intensity analysis and different regions of embryo were manually selected for comparison: in merged figures (A3–F3), the TE regions in rectangles were magnified and the arrows (A4–F4) indicate the representative TE cells with Oct-4 signals. Embryos at all stages examined, with the exception of day-3 compact morulae and day-4 expanded blastocysts, revealed a significant difference in Oct-4 intensity, showing stronger Oct-4 intensity in ICM and epiblast regions rather than in TE. (H) Two groups of cells (central and surrounding cells) in the ICM area of day-5 blastocysts showed different Oct-4 intensity (P < 0.05). D3CM = day-3 compact morulae; D4EBL = day-4 early blastocysts; D4EXPBL = day-4 expanded blastocysts; D5BL = day-5 blastocysts; D6BL = day-6 blastocysts; D7BL = day-7 blastocysts; Epi = epiblast; ICM = inner cell mass; TE = trophectoderm. Asterisks indicate statistically significant differences (P < 0.05). Bars = 200 µm (A3–F3) and 50 µm (A4–F4).