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Linda R. Mohr and A. O. Trounson

Summary. A hatched human blastocyst obtained after in-vitro fertilization and culture was examined by transmission electron microscopy and the ultrastructural features compared with hatched mouse and bovine blastocysts. The human blastocyst contained a continuous layer of trophoblast cells with apical junctional complexes, an inner cell mass and the beginning of a primitive endoderm layer. Certain ultrastructural features were common to the blastocysts of all 3 species; these included characteristic junction regions between adjacent trophoblast cells, an abundance of microvilli on the external surfaces of the blastocysts and the presence of well developed mitochondria and numerous ribosomes in the trophoblast cells. The features that were dissimilar included the extent of development of the endoderm layer, the appearance of the inner cell mass and the nature and extent of vesicular inclusions in the trophoblast cells.

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Linda R. Mohr and A. O. Trounson

Summary. Preimplantation mouse embryos that were exposed to fluorescein diacetate (FDA) accumulated intracellular fluorescein and fluoresced brightly under ultraviolet (u.v.) light. The rate at which intracellular fluorescein was lost from the cells was measured at 37, 28 and 4°C and the rate decreased as the storage temperature decreased. The rate at which intracellular fluorescein accumulated increased as FDA concentration increased until a maximum rate was attained. The ability to accumulate intracellular fluorescein could be removed by heating embryos at 56°C for 30 min or by damaging the cell membrane. Cells grown under inadequate culture conditions lost the ability to accumulate intracellular fluorescein. Exposure of 2-cell mouse embryos to FDA and u.v. light did not alter the rate of blastocyst formation in vitro, and exposure of blastocysts to FDA and u.v. light did not alter the rate of implantation or post-implantation development in vivo.

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A. O. Trounson, Linda R. Mohr, C. Wood and J. F. Leeton

Summary. Oocytes were obtained from patients with tubal infertility at fixed times after the onset of the endogenous LH rise or hCG injection, and were inseminated immediately after recovery or after periods of 4–4½, 5–5½ and 6–6½ h in culture in vitro. Delayed insemination resulted in a marked increase in the proportion of oocytes that were fertilized and developed to normal embryos and maximum rates occurred after 5–5½ h in culture (0–½ h, 26%; 4–4½ h, 50%; 5–5½ h, 89%; 6–6½ h, 69%). The range and mean (± s.d.) intervals from insemination for the pronuclear and early cleavage stages were 27–43 (35·6 ± 4·4) h for 2-cell stages, 36–65 (45·7 ± 8·3) h for 4-cell stages, 45–73 (54·3 ± 12·6) h for 8-cell stages and 68–85 h for the 16-cell stage. In 7/50 patients receiving 1 or 2 embryos at the 2-, 4- and 8-cell stages, fetal development was normal and 2 women had twin pregnancies (36% success compared with 8% for single embryos). All pregnancies were from the groups in which insemination was delayed for 5–6½ h. It is concluded that a short period of culture in vitro may allow the completion of oocyte maturation, and improve the results of in-vitro fertilization.