Department of Neuroscience, The Ohio State University, Columbus, OH 43210, United States.
J Neurosci Methods. 2012 Aug 15;209(2):344-50. doi: 10.1016/j.jneumeth.2012.06.015. Epub 2012 Jun 26.
FluoroMyelin™ Red is a commercially available water-soluble fluorescent dye that has selectivity for myelin. This dye is marketed for the visualization of myelin in brain cryosections, though it is also used widely to stain myelin in chemically fixed tissue. Here we have investigated the suitability of FluoroMyelin™ Red as a vital stain for live imaging of myelin in myelinating co-cultures of Schwann cells and dorsal root ganglion neurons. We show that addition of FluoroMyelin™ Red to the culture medium results in selective staining of myelin sheaths, with an optimal staining time of 2h, and has no apparent adverse effect on the neurons, their axons, or the myelinating cells at the light microscopic level. The fluorescence is bright and photostable, permitting long-term time-lapse imaging. After rinsing the cultures with medium lacking FluoroMyelin™ Red, the dye diffuses out of the myelin with a half life of about 130 min resulting in negligible fluorescence remaining after 18-24h. In addition, the large Stokes shift exhibited by FluoroMyelin™ Red makes it possible to readily distinguish it from popular and widely used green and red fluorescent probes such as GFP and mCherry. Thus FluoroMyelin™ Red is a useful reagent for live fluorescence imaging studies on myelinated axons.
FluoroMyelin™ Red 是一种商业上可获得的水溶性荧光染料，对髓鞘具有选择性。这种染料被用于脑冰冻切片中髓鞘的可视化，尽管它也被广泛用于化学固定组织中髓鞘的染色。在这里，我们研究了 FluoroMyelin™ Red 作为活髓鞘成像的示踪剂在施万细胞和背根神经节神经元共培养物中的适用性。我们表明，将 FluoroMyelin™ Red 添加到培养基中会导致髓鞘鞘选择性染色，最佳染色时间为 2 小时，并且在光镜水平上对神经元、其轴突或髓鞘细胞没有明显的不良影响。荧光明亮且稳定，允许进行长时间的延时成像。用不含 FluoroMyelin™ Red 的培养基冲洗培养物后，染料会从髓鞘中扩散出来，半衰期约为 130 分钟，在 18-24 小时后几乎没有荧光残留。此外，FluoroMyelin™ Red 表现出的大斯托克斯位移使其能够容易地区分它与 GFP 和 mCherry 等流行且广泛使用的绿色和红色荧光探针。因此，FluoroMyelin™ Red 是用于研究有髓轴突的活荧光成像的有用试剂。
