INTRODUCTION It is a more or less conspicuous fact that, although a very great number of experiments have been performed on tissues growing in vitro, relatively few of them deal with the significance of chemically well defined substances for the life, differentiation, and growth of cells and tissues. M. R. Lewis and W. H. Lewis (25, 26) were among the first to study the growth of tissues in media of known composition. As early as 1911 they published the results of their work on the cultivation of chick embryo tissues in solutions of various salts and salt combinations. In the same article they stated that an excellent growth could be obtained in combinations of amino-acids and polypeptids of known composition and also that media containing dextrose or maltose were better than pure salt solutions. The importance of glucose in the morphology of the cells was studied afterwards, especially by M. R. Lewis (23, 24). She found not only that the addition of dextrose improved the growth in those media which were not favorable (e.g. amnion liquid) but, also, that in the absence of carbohydrate, the cells became vacuolated and died within a few days. When dextrose was added to the media, vacuolation was retarded for several days. The fact that growth could be obtained with concentrations of glucose as high as 2 to 5 per cent is interesting. The significance, from a biochemical point of view, of carbohydrates in the culture medium has been studied by several workers, notably Krontowski and his collaborators (16, 17, 18, 19, 20). It is certain that glucose in a culture medium is very important. For a general discussion of the function of carbohydrates, the reader is referred to the well known book of Fischer (12) on tissue cultures. Much less is known about the activity of insulin, a substance closely related to carbohydrate metabolism. Krontowski, Bronstein, and Jazimirska-Krontowska (19) used it on rabbit tissue cultures and found that its addition augmented the depletion of sugar by spleen cultures. The spleen tissue also grew better in plasma from animals which had been injected with this substance (cited after Fischer). Friedheim and Roukhelman (14) concluded from their experiments that addition of insulin is unfavorable for growth, whereas Kuczynski, Tenenbaum and Werthemann (21) found that addition of insulin neutralized the “unfavorable action of glucose” in their medium. Roffo (28) observed that the addition of insulin generally causes a decrease in the growth of fibroblasts from chick embryo heart and rat fibroblasts. He could obtain growth only in very low concentrations of insulin. The very contradictory statements of sundry workers on the influence of insulin on tissue growth can be explained partly by the different sources of insulin and the manifold tissues used. That various organs react differently towards this important substance was found by Krontowski and his co-workers (19).