ABSTRACT
Single-pronuclear haploid parthenogenetically activated mouse embryos were transferred to the oviducts of suitable recipients. One group of embryos was isolated at the morula stage and subsequently allowed to develop to the expanded blastocyst stage in vitro. Intact embryos were either analysed by the air-drying technique at that stage to determine their total cell number and ploidy, or treated by immunosurgery to isolate their inner cell mass. These were either analysed to establish their total cell number and ploidy, or retained in culture for an additional 24 h or 72 h. The inner cell mass derivatives were then analysed to establish the total cell number and ploidy. A second group of recipients was ovariectomized on the 4th day of pseudopregnancy, treated with Depo-Provera and blastocysts recovered 5 or 6 days later. The ‘delayed’ blastocysts recovered were treated by immunosurgery, and the inner cell masses isolated and either analysed at this time or transferred to culture for 72 h, 96 h or 144h. As in the previous groups, the inner cell mass derivatives were analysed to establish the total cell population present and their ploidy. The analysis of this material was found to be technically particularly difficult, though in general the non-‘delayed’ embryos and their inner cell mass derivatives yielded higher success rates than the ‘delayed’ inner cell mass derivatives. The .‘delayed’ inner cell masses initially contained on average about twice the number of cells compared to the number present in those isolated from the non-‘delayed’ expanded blastocysts. Cellular profiferation occurred in all the groups retained in culture, though only a small proportion of the cells analysed gave ‘scorable’ mitotic cells in which the ploidy could be unequivocally determined. In general, in both the non-‘delayed’ and ‘delayed’ groups, the proportion of diploid mitotic cells observed increased with their duration in culture, though this effect was clearly more marked in the ‘delayed’ series. The present study indicated that the chance of obtaining haploid mouse cell lines in the future might be increased by using inner cell masses derived from non-‘delayed’ rather than ‘delayed’ blastocysts despite their initial reduced cell number at the time of explantation into tissue culture..