Evaluation of flow cytometric methods for determining population potential doubling times using cultured cells.
Evaluation of flow cytometric methods for determining population potential doubling times using cultured cells.
复制标题
评估使用培养细胞确定群体潜在倍增时间的流式细胞术方法。
DOI：
10.1002/cyto.990120305
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Calkins,DP
中科院分区：
文献类型：
--
作者：
Terry,NH;White,RA;Meistrich,ML;Calkins,DP
已经提出了各种方法来确定潜在的倍增时间（Tpot）的哺乳动物细胞群体，通过使用流式细胞术技术后，标记的细胞与溴脱氧尿苷（BrdUrd）。我们在这里表明，在一个明确定义的体外系统中，可以进行多次测量，所有方法都给出了接近真实群体倍增时间的相似结果。然而，最终感兴趣的是从单个时间点测定Tpot的准确性。在本文中，我们比较了在标记后不同时间进行这种测定的方法的准确度和精密度。在测定时未分裂的BrdUrd标记细胞的相对运动（RM）允许计算S期的长度（Ts）。当使用量ν（BrdUrd标记的分裂细胞分数和BrdUrd标记的未分裂细胞分数的函数）来估计RM的初始截距时，Tss估计的精确度得到提高。此外，根据公式Tpot = ln（2）Ts/v计算Tpot，得到的值最接近观察到的群体倍增时间。建议使用RM作为根据单个时间点观察值计算Tpot的分析方法，最好在G2和M期（TG 2 M）和Ts之间的时间进行。
Various methods have been proposed for determining the potential doubling times (Tpot) of mammalian cell populations by using flow cytometric techniques after labeling the cells with bromodeoxyuridine (BrdUrd). We show here that, in a well‐defined in vitro system where multiple time measurements are possible, all the methods give similar results that are close to the true population doubling time. Of ultimate interest, however, is the accuracy of determination of Tpot from a single time point. In this paper we compare the accuracy and precision of the methods in making such determinations at different times after labeling.The relative movement (RM) of BrdUrd‐labeled cells that have not divided at the time of assay allows for computation of the length of S phase (Ts). The precision of estimation of Tswas enhanced when a quantity, ν (a function of the fraction of BrdUrd‐labeled divided and the fraction of BrdUrd‐labeled undivided cells), was used to estimate the initial intercept of RM. Furthermore, calculation of Tpot from the formula, Tpot = ln(2) Ts/v, gave values closest to the observed population doubling time. It is suggested that the use of RM withvbe the analytical method of choice for the calculation of Tpot from single time‐point observations, preferably made at times between the length of the G2and M phases (TG2M) and Ts.