Ultrastructure and molecular cytogenetics of human metaphase II oocytes with granular vacuoles

Abstract

Studies demonstrated poor fertilization rates and compromised early embryo development in cases of oocytes with dark and granular ooplasm, or with central granular ooplasm and presence of a refractile body. Many other structural abnormalities of human mature oocytes do exist, but remain to be evaluated. In the present study we characterized granular vacuoles (GVa) of mature metaphase II human oocytes. Surplus abnormal MII oocytes (n=6) with GVa were used under informed consent, after controlled superovulation during IVF treatment cycles. In all cases, patients had normal karyotypes. For electron microscopy oocytes were fixed with karnovsky fixative for 2h at 4°C, post-fixed in 1-2% OsO4 in buffer containing 0,8% K3Fe3+(CN)6, dehydrated in ethanol series, equilibrated with propylene oxide and embedded in Epon. Semithin sections were stained with aqueous azur II and methylene blue (1:1). Ultrathin sections were stained with 3% aquous uranyl acetate (20 min) and Reynols lead citrate (10 min). Sections were observed with a transmission electron microscope JEOL 100CXII operated at 60kV.