At the beginning of the 1990s, co-culture of cattle and sheep embryos was the most favoured method to support embryo development, but the use of this system has hampered progress in raising the efficiency of embryo production. Furthermore, little was known of the requirements of embryos and the biochemistry of early embryo development. As the decade progressed, energy metabolism studies improved our understanding of the energy substrate requirements for embryo development. Furthermore, an appreciation of the reproductive tract environment increased. This resulted in more "defined" systems, which have evolved further in the development of "sequential" media systems, where components change in accordance to the needs of the embryo. Nevertheless, wholly defined systems, such as the replacement of albumin with PVA, are less able to support similar levels of development as protein-containing medium, and the resulting embryos are metabolically compromised. This highlights the nutritive role of albumin. One area in which much work has been conducted, but yet no unifying theory has emerged, is that of the interactive roles of growth factors (including autocrine/paracrine), cytokines and extra-cellular matrix molecules in the development of a viable embryo. A new concept is that of regulation of energy metabolism. Compounds such as ethylenediamine tetraacetic acid (EDTA), NaN(3) and 2,4-dinitrophenol have been shown to increase embryo development and quality of resulting embryos. This demonstrates that the process of ATP production is a key regulator of in vitro embryo development.