Ng Kee W, Leong David T W, Hutmacher Dietmar W
Department of Surgery, National University of Singapore, Singapore.
Tissue Eng. 2005 Jan-Feb;11(1-2):182-91. doi: 10.1089/ten.2005.11.182.
Various assays, using different strategies, are available for assessing cultured cell proliferation. These include measurement of metabolic activity (tetrazolium salts and alamarBlue), DNA quantification using fluorophores (Hoechst 33258 and PicoGreen), uptake of radioactively-labeled DNA precursors such as [3H]thymidine, and physical counting (hemocytometer). These assays are well established in characterizing cell proliferation in two-dimensional (2D), monolayer cultures of low cell densities. However, increasing interest in 3D cultures has prompted the need to evaluate the effectiveness of using these assays in high cell density or 3D cultures. We show here that typical cell proliferation assays do not necessarily correlate linearly with increasing cell densities or between 2D and 3D cultures, and are either not suitable or only rough approximations in quantifying actual cell numbers in a culture. Prudent choice of techniques and careful interpretation of data are therefore recommended when measuring cell proliferation in high cell density and 3D cultures.
有多种采用不同策略的检测方法可用于评估培养细胞的增殖。这些方法包括代谢活性测定（四氮唑盐和alamarBlue）、使用荧光团进行DNA定量（Hoechst 33258和PicoGreen）、摄取放射性标记的DNA前体如[3H]胸腺嘧啶核苷以及物理计数（血细胞计数器）。这些检测方法在表征二维（2D）、低细胞密度单层培养中的细胞增殖方面已得到充分确立。然而，对三维（3D）培养的兴趣日益增加，促使人们需要评估在高细胞密度或3D培养中使用这些检测方法的有效性。我们在此表明，典型的细胞增殖检测方法不一定与细胞密度增加呈线性相关，也不一定在2D和3D培养之间呈线性相关，并且在量化培养物中的实际细胞数量时要么不适用，要么只是粗略的近似值。因此，在高细胞密度和3D培养中测量细胞增殖时，建议谨慎选择技术并仔细解读数据。
