Methods for Inferring Cell Cycle Parameters Using Thymidine Analogues

Tritiated thymidine autoradiography, 5-bromo-2'-deoxyuridine (BrdU) 5-chloro-2'-deoxyuridine (CldU), 5-iodo-2'-deoxyuridine (IdU), and 5-ethynyl-2'-deoxyiridine (EdU) labeling have been used for identifying the fraction of cells undergoing the S-phase of the cell cycle and to follow the fate of these cells during the embryonic, perinatal, and adult life in several species of vertebrate. In this current review, I will discuss the dosage and times of exposition to the aforementioned thymidine analogues to label most of the cells undergoing the S-phase of the cell cycle. I will also show how to infer, in an asynchronous cell population, the duration of the G₁, S, and G₂ phases, as well as the growth fraction and the span of the whole cell cycle on the base of some labeling schemes involving a single administration, continuous nucleotide analogue delivery, and double labeling with two thymidine analogues. In this context, the choice of the optimal dose of BrdU, CldU, IdU, and EdU to label S-phase cells is a pivotal aspect to produce neither cytotoxic effects nor alter cell cycle progression. I hope that the information presented in this review can be of use as a reference for researchers involved in the genesis of tissues and organs.