An evaluation of various double labelling and autoradiographic techniques for the measurement of DNA synthesis time in leukaemic cells*

SUMMARY Before applying the double labelling technique to the measurement of DNA synthesis time in leukaemic cells, various methods of labelling and of autoradiography were compared. The use of low and high doses of 3H‐thymidine proved to be impracticable because, owing to a wide range of uptake per cell, it was impossible to make a clear distinction between ‘lightly’ and ‘heavily’ labelled cells. Two consecutive doses of 3H‐thymidine resulted in only a small increase in labelling index because of the low percentage of cells in the S‐phase and the long duration of DNA synthesis. Consequently, it was not possible to establish the DNA synthesis time with any degree of accuracy by this method. The remaining method employs 3H‐thymidine and 14C‐thymidine. Auto‐radiographs made with a single emulsion proved difficult to interpret since some 14C‐labelled cells give the appearance of being labelled with 3H‐thymidine only. A double emulsion autoradiographic technique is to be preferred, but even then, more than one interval between the two isotopes is necessary in order to eliminate an error in the recognition of 14C‐labelled cells.