Comparison of different labelling index formulae used on bromodeoxyuridine-flow cytometry data.

The essence of the bromodeoxyuridine (BrdUrd)-flow cytometry (FCM) technique is that cells are labelled with the thymidine analogue BrdUrd. They are then allowed to progress through the cell cycle in a BrdUrd-free environment during the postlabelling time period. At a postlabelling time shorter than the length of the S phase (Ts), cells are fixed and prepared for FCM-mediated analysis of BrdUrd and DNA contents. From FCM-derived data, cell cycle kinetic parameters such as labelling index (LI), Ts, and potential doubling time (Tpot) can be calculated. Tpot is believed to be important in the evaluation of tumor aggressiveness and therapy response. Since LI is most commonly used together with Ts to calculate Tpot, it is important that both LI and Ts are independent of the time when cells are sampled. Several formulae to calculate LI and Ts have been presented. In the present paper, we deal with various formulae to calculate LI. These formulae differ in how they take into account unlabelled and BrdUrd-labelled cells in various fractions of the cell cycle. We present a new formula, which takes into consideration cells in the different fractions and thus makes LI theoretically independent of postlabelling time. Our results show that different LI values are obtained when different formulae are used to calculate LI. In addition, we show that the BrdUrd labelling period should be kept as short as possible.