CELLULAR COMPOSITION AND VIABILITY OF DEMI-EMBRYOS AND QUARTER-EMBRYOS MADE FROM BISECTED BOVINE MORULAE AND BLASTOCYSTS PRODUCED IN-VITRO

The cellular composition and viability of intact, IVP embryos were com pared with those of demi- and quarter-embryos produced by bisection of IVP morulae and blastocysts. Embryos were produced by established tec hniques from oocytes harvested from slaughterhouse ovaries. In Experim ent 1, morulae at Day 6 or blastocysts at Day 7 were bisected on an in verted microscope using a microsurgical steel blade. Demi-embryos were then cultured without a zona pellucida until Day 8, when they were mo rphologically assessed for quality (viability). A higher proportion of demi-embryos made from blastocysts than from morulae were classified as viable (381/420, 91% vs 164/267, 61%; P<0.001). In Experiment 2, on ly Day 7 blastocysts were bisected, and some of the resulting demi-emb ryos were bisected a second time 24 h later to produce quarter-embryos . The remaining demi-embryos, the quarter-embryos, and control intact embryos were cultured until Day 9, at which time they were assessed fo r quality and subjected to immunosurgery and differential staining to count inner cell mass (ICM) and trophectoderm cells. A higher proporti on of demi-embryos than quarter-embryos was classified as viable (408/ 459, 89% vs 223/319, 70%, respectively; P<0.001). Total cell numbers d ecreased with successive bisections, but the proportion of surviving c ells found in the ICM was significantly (P<0.05) higher in the best qu ality demi- and quarter-embryos (35 and 32%, respectively) than in the controls (22%). Transfer of all 12 quarter-embryos derived from 3 bla stocysts, in pairs, into 6 recipient heifers resulted in 2 pregnancies , each with a single viable fetus at 90 d of gestation. The fetuses or iginated from 2 different blastocysts. The results suggest that bisect ion of intact IVP embryos into demi-embryos and bisection of those int o quarter-embryos can increase the number of transferable embryos by a s much as 178 and 235%, respectively. (C) 1998 by Elsevier Science Inc .