Fluorescence microscopy of nuclear DNA in oocytes and zygotes during in vitro fertilization and development of early embryos in mice.

Fluorescence microscopic investigations of nuclear DNA stained with mithramycin or 4',6-diamidino-2-phenylindole (DAPI) were undertaken in oocytes and zygotes during in vitro fertilization (IVF) and subsequent early embryo development in mice. Mithramycin and DAPI bind stoichiometrically with G-C and A-T pairs of DNA, respectively. In the present study, it was possible to visualize the germinal vesicle, the nucleolus, and the chromosome in the oocyte. After IVF, the nuclei, male and female pronuclei, polar bodies, nucleolus-associated chromatins, and chromosomes were successively observed in the zygote, the blastomere during cleavage, and the blastocyst until the hatching stage. The formation of male and female pronuclei, the process of their fusion, the appearance of the chromosome, and the commencement of the first cleavage of the zygote to form two blastomeres were all clearly demonstrated. The number of blastomeres was easily counted in the morula in spite of the compaction of the blastomeres. The present study indicates that the methods reported herein are useful for the determination of exact functional stages of the oocyte and the zygote during IVF and early embryo development, and thus contributes to the advancement of reproductive and developmental biomedical research.