An Immobilization Technique for Long-Term Time-Lapse Imaging of Explanted Drosophila Tissues.
An Immobilization Technique for Long-Term Time-Lapse Imaging of Explanted Drosophila Tissues.
复制标题
DOI：
10.3389/fcell.2020.590094
复制
发表时间：
2020
影响因子：
5.5
通讯作者：
Fernandes VM
中科院分区：
文献类型：
--
作者：
Bostock MP;Prasad AR;Chaouni R;Yuen AC;Sousa-Nunes R;Amoyel M;Fernandes VM
时间推移成像是研究动态生物过程的重要工具，而动态生物过程不能仅从固定的样本中辨别出来。然而，如果生物体是不透明和/或可移动的，在完整动物身上对细胞和组织水平的过程进行成像会带来许多挑战。完整组织的外植体培养绕过了其中的一些挑战，但样本漂移仍然是一个相当大的障碍。我们使用了一种简单而有效的技术来固定在中浴琼脂糖凝胶中的组织。我们应用这项技术研究了从一龄幼虫到成虫不同方向的多个果蝇组织，没有证据表明各向异性压力或应力损伤。使用这种方法，我们能够在长达18小时的时间内成像精细的特征，并进行新颖的观察。具体地说，我们报告了静止的神经母细胞的纤维特征在重新激活期间由其基本子代继承；发育中的视觉系统中的板层大约一次组装2-3列；胶质板层在发育过程中是动态的；成年睾丸囊干细胞的核膜在有丝分裂期间不会完全分解。总之，我们证明了我们的方案非常适合组织固定和长期活体成像，使我们能够对果蝇的组织和细胞动力学有新的见解。
Time-lapse imaging is an essential tool to study dynamic biological processes that cannot be discerned from fixed samples alone. However, imaging cell- and tissue-level processes in intact animals poses numerous challenges if the organism is opaque and/or motile. Explant cultures of intact tissues circumvent some of these challenges, but sample drift remains a considerable obstacle. We employed a simple yet effective technique to immobilize tissues in medium-bathed agarose. We applied this technique to study multiple Drosophila tissues from first-instar larvae to adult stages in various orientations and with no evidence of anisotropic pressure or stress damage. Using this method, we were able to image fine features for up to 18 h and make novel observations. Specifically, we report that fibers characteristic of quiescent neuroblasts are inherited by their basal daughters during reactivation; that the lamina in the developing visual system is assembled roughly 2–3 columns at a time; that lamina glia positions are dynamic during development; and that the nuclear envelopes of adult testis cyst stem cells do not break down completely during mitosis. In all, we demonstrate that our protocol is well-suited for tissue immobilization and long-term live imaging, enabling new insights into tissue and cell dynamics in Drosophila.