With the object of discovering the significance of the "opaqueness" surrounding so-called lipolytic colonies on fat-emulsion agar plates, detailed observations have been made on uninoculated globules of butterfat, pure triglycerides, fatty acids, and mixtures of fats and fatty acids. The following observations have been made:At room temperature, sooner or later, fat crystals form in all butterfat globules. The rapidity with which crystals form and their general conformation and texture are capable of considerable variation. This depends upon the previous history of the butterfat, temperature, and the physical and chemical characteristics of the agar medium in which the fat is suspended.Sodium chloride, among other substances, brings about rapid changes in the form of the globules and the texture of the crystals.The crystals of pure fats suspended in agar are quite unlike the crystals in globules of butterfat. When these same fats are suspended in some form of oily matrix this difference disappears.The chief difference between pure fats and fatty acids, when suspended in oil, is that there is a tendency for the fatty acids to crystallize at the fat-agar interface, while the fats do not. The lower, solid fatty acids often protrude out into the agar medium.As well as copper sulphate and Nile blue sulphate, a large number of other dyes have been used to differentiate fatty acids from fats.It appears probable that the phenomenon of opaqueness in the fat globules surrounding lipolytic colonies in fat-emulsion agar plates is owing, at least in part, to the rapid formation of fat crystals, as well as to the formation of fatty acids.