Long-term high-resolution imaging and culture of C. elegans in chip-gel hybrid microfluidic device for developmental studies.
Long-term high-resolution imaging and culture of C. elegans in chip-gel hybrid microfluidic device for developmental studies.
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DOI：
10.1039/c001986k
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发表时间：
2010-07-21
期刊：
影响因子：
6.1
通讯作者：
Lu H
中科院分区：
文献类型：
--
作者：
Krajniak J;Lu H
多细胞模型生物（例如秀丽隐杆线虫）的发育研究广泛依赖于在细胞或亚细胞水平上重复培养和成像动物的能力。然而，标准高分辨率成像技术需要使用麻醉剂进行固定，并且可能会对发育产生不良副作用。因此，此类技术对于允许相同动物在整个发育过程中生长和成像来说并不理想。在本文中，我们提出了一种旨在克服这些困难的微流体系统。该系统允许从 L1 幼虫阶段开始长期培养线虫，并在生理温度下重复高分辨率成像，而无需使用麻醉剂。我们使用市售的生物相容性聚合物 Pluronic F127 进行固定；这种聚合物能够在~1°C内发生可逆的热敏溶胶-凝胶转变，这在微流控芯片中得到了很好的控制。凝胶相足以固定动物。虽然动物没有成像，但它们在单独的培养室中用含有发育所需营养物质的培养基进行培养。我们在这里表明，这种方法有助于以高分辨率对单个动物进行延时研究，并适用于发育过程和动态事件的实时成像实验。
Developmental studies in multicellular model organisms such as Caernohabditis elegans rely extensively on the ability to cultivate and image animals repeatedly at the cell or subcellular level. However, standard high-resolution imaging techniques require the use of anesthetics for immobilization, and may have undesirable side effects on development. Thus such techniques are not ideal in allowing the same animals to grow and be imaged throughout development. In this paper, we present a microfluidic system designed to overcome these difficulties. The system allows for long-term culture of C. elegans starting at L1 larval stage and repeated high-resolution imaging at physiological temperatures without using anaesthetics. We use a commercially available biocompatible polymer, Pluronic F127 for immobilization; this polymer is capable of a reversible thermo-sensitive sol-gel transition within ~1 °C, which is well-controlled in the microfluidic chip. The gel phase is sufficient to immobilize the animals. While animals are not imaged, they are cultured in individual chambers in media containing nutrients required for devleopment. We show here that this method facilitates time-lapse studies of single animals at high-resolution and lends itself to live imaging experiments on developmental processes and dynamic events.