Fig. 3. Time course of appearance of P2 and P4 prominin-1-containing membrane particles in the ventricular fluid during embryonic mouse brain development. (a) Ventricular fluid of E10.5-E13.5 mouse embryos was subjected to differential centrifugation as in Fig. 2a followed by immunoblotting of the pellets for prominin-1. Total prominin-1 immunoreactivity (sum of P1-P4 pellets, triangles) and prominin-1 immunoreactivity in the P2 pellet (squares) and the P4 pellet (circles) at the various developmental stages was determined. Data are the mean of three independent experiments; bars indicate s.d. For any given experiment, the same volume of ventricular fluid was analysed at the various developmental stages. (b) Cryosections of E7-E12.5 mouse embryos were subjected to immunofluorescence for prominin-1. The number of the ring-like prominin-1-containing particles (see inset in Fig. 1f) in the ventricular fluid was quantified per unit area in single optical sections at the level of the glass slide. E7, amniotic fluid near the neural plate (19x10⁴ µm²); E9.5-E12.5, mean of forebrain and hindbrain ventricular fluids, bars indicate the variation of the forebrain/hindbrain values from the mean (E9.5, 66x10⁴ µm²; E10.5, 139x10⁴ µm²; E12.5, 134x10⁴ µm²).