Application of hanging drop technique for stem cell differentiation and cytotoxicity studies
Application of hanging drop technique for stem cell differentiation and cytotoxicity studies
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DOI：
10.1007/s10616-006-9001-z
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发表时间：
2006-01-01
期刊：
影响因子：
2.2
通讯作者：
Bhonde, Ramesh R.
中科院分区：
文献类型：
--
作者：
Banerjee, Meenal;Bhonde, Ramesh R.
本研究的目的是探索使用一种古老的培养技术-悬滴技术进行干细胞分化和细胞毒性测试的可能性。我们在这里展示了这种古老技术的各种新应用，不仅可以利用干细胞分化为特定谱系的潜力，还可以用于细胞毒性研究。在此，我们通过将20 μ 1微滴的营养培养基和含有感兴趣的细胞的10%胎牛血清（FCS）置于60 mm培养皿的盖子上来制备悬滴培养物。用无菌磷酸盐缓冲盐水（PBS）填充培养皿的底板，以避免样品干燥。然后将盖置于底板上以实现悬滴培养。我们利用此技术培养纤毛上皮细胞，研究骨髓基质细胞的细胞毒性和分化。最重要的是，本文提出的改良培养技术在所需时间和所需试剂方面是简单、经济和具有成本效益的，并且适合于目标特定的修改，例如细胞毒性测试。它在更长时间内保持活力和功能性以及提供三维生长微环境方面优于现有系统，使其可用于器官型培养和体内模拟。
The aim of our study is to explore the possibility of using an ancient method of culture technique- the hanging drop technique for stem cell differentiation and cytotoxicity testing. We demonstrate here a variety of novel applications of this age old technique not only to harness the differentiation potential of stem cells into specific lineages but also for cytotoxicity studies. Here we have prepared hanging drop cultures by placing 20 VI micro-drops of nutrient media and 10% Fetal Calf Serum (FCS) containing cells of interest on the lids of 60 mm dishes. Bottom plates of the dishes were filled with sterile Phosphate Buffer Saline (PBS) to avoid desiccation of samples. Lids were then placed on the bottom plates to achieve hanging drop cultures. We utilized this technique for cultivation of ciliated epithelia to study cytotoxicity and differentiation of bone marrow stromal cells. Most importantly the modified culture technique presented here is simple, economical and cost effective in terms of the time taken and the reagents required and are amenable to goal specific modification such as cytotoxicity testing. It is advantageous over the existing system in terms of retention of viability and functionality for longer duration and for providing three dimensional growth micro-environment making it useful for organotypic cultures and in vivo simulation.