Anti-mESOP-1 antibodies and mESOP-1 protein expression.

(A) Analysis of the specificity of anti-mESOP-1 antibodies. The coding region of mESOP-1 with FLAG tag at the 3′ end was recloned into the expression vector pEF-BOS (pEF-BOS-mESOP-1-FL). 293T cells were transfected with pEF-BOS-mESOP-1-FL (lanes 2, 4, 6) or vacant pEF-BOS vector (lanes 1, 3, 5) as a negative control using lipofectamine. Lysates of 293T cells were probed by either anti-FLAG antibodies (lanes 1, 2) or anti-mESOP-1 antibodies AK612 (lanes 3, 4) or AK641 (lanes 5, 6). (B) Immunohistochemical detection of mESOP-1 protein expression in yolk sac (i-vi), developing nervous system (vii, viii), and adult genital organs (ix, x). Both AK612 and AK641 gave the same results. Whole-mount staining of 7.5 dpc (i, ×30), 8.5 dpc (ii, ×25) embryo, and 9.0 dpc yolk sac (iii, ×12) . Magnified view (iv, ×80) and flat-mount view (v, ×200) of 9.0 dpc yolk sac, Cross-section of ii (vi, ×600). Arrowheads in a show blood islands. Arrowheads in iv show ESOP-1–positive endothelial cells in yolk sac. Spaces between the pairs of facing arrowheads indicate lumen of vascular plexus. Arrows in e show endothelial cells that expressed ESOP-1 protein. Transverse (vii, ×45) and parasagittal (viii, ×12) slices of 12.5 dpc embryos show ESOP-1 expression in the nervous system. Arrowheads show the hair follicles of vibrissae. Staining of adult ovary (ix, ×45) and testis (x, ×30) shows that all follicles, from small follicles to large, maturated follicles, oocytes (arrowheads), and granulosa cells (arrows) are ESOP-1 positive, and the signals become stronger as follicles develop. In testis, the cells in seminiferous tubules (asterisks) are stained. They may be spermatogonia or spermatocytes. VV, vitelline vessel; Br, brain; NT, neural tube; DRG, dorsal root ganglia; SN, sympathetic nerve; TN, trigeminal nerve; SF, small follicles; MF, mature follicles.