Morimoto K, Sato M, Koizumi A
Exp Cell Res. 1983 May;145(2):349-56. doi: 10.1016/0014-4827(83)90013-7.
A simple combination of autoradiography, to determine when a cell synthesized DNA, and sister chromatid differential staining, to determine how many times a cell has divided, was used to follow up the proliferating fate of human lymphocytes in culture. Cells were incubated continuously with 5-bromodeoxyuridine (BrdU) and pulse-labelled with 0.1 muCi/ml [3H]thymidine at various times after stimulation with phytohemagglutinin (PHA). The cells were then harvested at 4 h intervals up to 72 h, and the percentage of labelled mitoses was determined separately in first, second, or third division cells. The data showed that the cycling cells, whether they began cycling at earlier or later times after stimulation, had about the same generation times of 12--14 h. This confirms that the heterogeneity of cell generations seen in short-term lymphocyte cultures is in large part due to the difference in the times when cells began cell cycling in response to PHA.
采用一种简单的组合方法，即利用放射自显影术来确定细胞何时合成DNA，利用姐妹染色单体差异染色法来确定细胞分裂的次数，以此追踪培养的人淋巴细胞的增殖命运。用5-溴脱氧尿苷（BrdU）持续孵育细胞，并在植物血凝素（PHA）刺激后的不同时间用0.1μCi/ml [3H]胸腺嘧啶核苷进行脉冲标记。然后每隔4小时收获细胞，直至72小时，并分别测定第一代、第二代或第三代分裂细胞中标记有丝分裂的百分比。数据显示，循环细胞无论在刺激后的早期还是晚期开始循环，其世代时间大致相同，为12 - 14小时。这证实了在短期淋巴细胞培养中观察到的细胞世代异质性在很大程度上是由于细胞响应PHA开始细胞循环的时间不同所致。
