Stewart G J, Wang Y, Niewiarowski S
Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA.
Biotechniques. 1995 Oct;19(4):598-604.
Preparation of suspension of anchorage-dependent cells growing in tissue cultures requires removal of the cells from their substrata by means of trypsin and/or EDTA. The purpose of the experiments was to investigate the effect of methylcellulose on the adhesive ability of cells removed from substrata by EDTA and trypsin. Human umbilical cord vein endothelial cells (HUVEC), after detachment from substrata, adhere well to fibronectin at 37 degrees C but not at 0 degrees C. During a 60-min incubation at 37 degrees C, these cells lose about 70% of their ability to adhere to fibronectin. The ability of cells to adhere was restored in the presence of 0.2% methylcellulose. Methylcellulose also prevented human skin fibroblasts, human melanoma cells and mouse lung fibroblasts from losing adhesive properties. By contrast, it did not affect the adhesive ability of B16F10 melanoma cells incubated at 37 degrees C. In conclusion, our study suggests that methylcellulose can be a useful reagent for preservation of cell function in suspension; it may also simplify some experimental procedures including radioiodination of cell surface components and cross-linking of radiolabeled ligands to the cell surface.
制备在组织培养中生长的贴壁依赖性细胞的悬液,需要通过胰蛋白酶和/或乙二胺四乙酸(EDTA)将细胞从其生长基质上分离下来。本实验的目的是研究甲基纤维素对经EDTA和胰蛋白酶从基质上分离下来的细胞黏附能力的影响。人脐静脉内皮细胞(HUVEC)从基质上脱落后,在37℃时能很好地黏附于纤连蛋白,但在0℃时则不能。在37℃孵育60分钟期间,这些细胞失去了约70%黏附于纤连蛋白的能力。在0.2%甲基纤维素存在的情况下,细胞的黏附能力得以恢复。甲基纤维素还能防止人皮肤成纤维细胞、人黑色素瘤细胞和小鼠肺成纤维细胞丧失黏附特性。相比之下,它对在37℃孵育的B16F10黑色素瘤细胞的黏附能力没有影响。总之,我们的研究表明,甲基纤维素可能是一种用于保存悬浮细胞功能的有用试剂;它还可能简化一些实验程序,包括细胞表面成分的放射性碘化以及放射性标记配体与细胞表面的交联。