FIG. 4. .

TPEN inhibits the ability of EMI2 to maintain metaphase arrest in oocytes. A) Cumulus-denuded GV oocytes were injected with Emi2 cRNA, held in IBMX for 2–3 h, and transferred to IVM medium for 14 h using control medium or medium containing 10 μM TPEN. B–D) Oocytes were fixed; stained for actin (red), tubulin (green), and DNA (blue); imaged by confocal microscopy; and scored for spindle stage. In all, 100% of Emi2 cRNA-injected oocytes cultured in control medium arrested at MI, whereas EMI2 expression caused MI arrest in only 44% of oocytes cultured in TPEN-containing medium. This experiment was repeated four times; data from one representative experiment are shown. Projections of confocal Z stacks are shown for Emi2 cRNA-injected oocytes cultured in TPEN containing medium arrested at MI (B), TI (C), and MII (D). Bar = 20 μm.