Biophys J. 2012 Jul 3;103(1):L7-9. doi: 10.1016/j.bpj.2012.05.038.
NADH is a naturally fluorescent metabolite associated with cellular respiration. Exploiting the different fluorescence lifetime of free and bound NADH has the potential to quantify the relative amount of bound and free NADH, enhancing understanding of cellular processes including apoptosis, cancer pathology, and enzyme kinetics. We use the phasor-fluorescence lifetime image microscopy approach to spatially map NADH in both the free and bound forms of live undifferentiated and differentiated myoblast cells. The phasor approach graphically depicts the change in lifetime at a pixel level without the requirement for fitting the decay. Comparison of the spatial distribution of NADH in the nucleus of cells induced to differentiate through serum starvation and undifferentiated cells show differing distributions of bound and free NADH. Undifferentiated cells displayed a short lifetime indicative of free NADH in the nucleus and a longer lifetime attributed to the presence of bound NADH outside of the nucleus. Differentiating cells displayed redistribution of free NADH with decreased relative concentration of free NADH within the nucleus whereas the majority of NADH was found in the cytoplasm.
NADH 是一种与细胞呼吸相关的天然荧光代谢产物。利用游离和结合态 NADH 的不同荧光寿命，有可能定量测定结合态和游离态 NADH 的相对含量，从而增进对包括细胞凋亡、癌症病理学和酶动力学在内的细胞过程的理解。我们使用相图-荧光寿命图像显微镜方法，在活的未分化和分化成肌细胞中，以空间方式绘制游离态和结合态 NADH。相图方法以图形方式在像素水平上描绘了寿命的变化，而无需进行衰减拟合。通过血清饥饿诱导分化的细胞和未分化细胞中 NADH 的空间分布比较显示，结合态和游离态 NADH 的分布不同。未分化细胞的核内显示出短寿命，表明游离态 NADH 存在，而核外的长寿命归因于结合态 NADH 的存在。分化细胞显示出游离态 NADH 的重新分布，核内游离态 NADH 的相对浓度降低，而大部分 NADH 存在于细胞质中。
