It has been our rcutine procedure to sacrifice animals four hours after the in of tritiated thymidine. Pilot experiments had shown that the labeling satisfactory in four hours. was quite At this interval of time occasional labeled bai: Cells were found showing that division of the cell (with labeling in both) taken place. nuclei) hac It was known that the availability of labeled thymidine aftih injection into the circulaticn was very short, owing to the rapid disappearance by Peeradation or uptake by nuclei in the process of DNA synthesis. Since only those ce which arm in the stage of DNA duplication at the time the thymidine i: s available willl these nuclei are identifiable. be laelen Several series of young, identical growing rats were injected with tphitiated thymidine and sacrificed at 15, 30, and 45 minutes and 1, 2, 3, 4, 6, 8, LO, 12, iw, = 20 and 2% hours. Three rats were sacrificed at each time interval. The Incideme of labeled nuclei was determined by counting the number of labeled nuclei infa larse Am of high power fields in microscopic sections of uniform thickness. Paired labeled ceils among this large population of unlabeled cells were also recorded. Within 30 minutes following the injection of tritiated thymidine, cells were found to contain the tritium label. # There was a sequential reacies of = ride in te incidence of these labeled miclei. This reached a plateau at about 10 holrs. In oce to determine the duration of available tritiated thymidine in the circulation, the animals were sacrificed by exsanguination,and the radioactivity in the plasma determi= In order to find out how mich of the radioactivity had been reduced th tritiated water in the plasma, samples were evaporated to dryness and counted. The fraciioactivi— in the solid fraction of the plasma was found to be only about 15% of the Jtota radioactivity witnin 2 hours. A great deal of time was then devoted to t ying to identify and quantitate by chromatography the tritium labeled organic metabolic produc as well as the ine remaining in the blood in order to better assess 2 availability of the thymidine after an hour or mcre. These attempts were Bbandore because others had shown rapid disappearance of available material and thej amounts hem in these experiments were nct paramount to our objectives. A continuing rise in the cccurrence of labeled meclei for as long as [0 hours in ~ face of rapid disappearance of tritiated thymidine from the circulation wab sound. Th increasing number of labeled nuclei up to 10 hours was very clearly demonsfrate ia two complete series of animal experiments. Pairs of labeled nuclei were f sean with certainty at 4 hours, although sane very lightly labeled pairs of nuche? were Suspected somewhat earlier. It was assumed that the lightly labeled pair @f muciei hac been in the very late stage of DNA synthesis and therefore had an opportunity to <=ke in up very little of the thymidine before DNA synthesis terminated and actual|divisim of the cell took place. The occurrence of labeled pairs increased with time dnd reacted a plateau at about 10 hours. Since a thin microscopic section represents 4 thickess tissue little wider than a cell, a pair of labeled miclei would be demonst§ated miy ithe cuts in the tissue happened to pass through both of the daughter mucleq. It can be assumed that random cutting would only reveal an occasional pair where the piain ct the : cut corresponded with the plain of the pair. It seems likely that the conqi increase of labeled cells, well beyond the time when tritiated thymidine was avail=le in the circulation, can be explained on the contimued division of cells and cuts that might catch one or the other or both of a pair.