Cell loss in bisected mouse, sheep and cow embryos

The purpose of this experiment was to determine the loss of cells during the splitting of embryos. The experiment was carried out on early, middle and late stage mouse blastocysts as well as on late stage sheep and cow blastocysts. The embryos were bisected with a metal mioroscalpel. Manipulation of the embryos was done in phosphate buffered saline (PBS) solution with 20% fetal calf serum (FCS) at +37 degrees C or in PBS solution free of Ca(++) and Mg(++) ions. To determine the number of cells present, intact embryos and demi-embryos were placed in acridine orange solution and examined under a fluorescent microscope. The average number of cells in early, middle and late stage mouse blastocysts was 39.4, 54.1 and 71.9, respectively. The loss of cells caused by splitting was 6.4, 7.6 and 9.9, respectively. The average number of cells in late stage sheep blastocysts was 115.3 and in late stage cow blastocysts it was 138.5. In sheep and cows, the loss of cells due to splitting the embryos in a PBS solution free of Ca(++) and Mg(++) ions did not differ from that observed in a PBS solution with 20% FCS. Similarly, bisection carried out at +37 degrees C did not result in lower cell losses, than that done at room temperature.